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  • High‐resolution expression profiling of selected gene sets during plant immune activation
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-27
    Pingtao Ding; Bruno Pok Man Ngou; Oliver J. Furzer; Toshiyuki Sakai; Ram Krishna Shrestha; Dan MacLean; Jonathan D. G. Jones

    The plant immune system involves detection of pathogens via both cell‐surface and intracellular receptors. Both receptor classes can induce transcriptional reprogramming that elevates disease resistance. To assess differential gene expression during plant immunity, we developed and deployed quantitative sequence capture (CAP‐I). We designed and synthesized biotinylated single‐strand RNA bait libraries targeted to a subset of defense genes, and generated sequence capture data from 99 RNA‐seq libraries. We built a data processing pipeline to quantify the RNA‐CAP‐I‐seq data, and visualize differential gene expression. Sequence capture in combination with quantitative RNA‐seq enabled cost‐effective assessment of the expression profile of a specified subset of genes. Quantitative sequence capture is not limited to RNA‐seq or any specific organism and can potentially be incorporated into automated platforms for high‐throughput sequencing.

    更新日期:2020-01-27
  • Use of synthetic biology tools to optimize the production of active nitrogenase Fe protein in chloroplasts of tobacco leaf cells
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-27
    Álvaro Eseverri; Gema López‐Torrejón; Xi Jiang; Stefan Burén; Luis M. Rubio; Elena Caro

    The generation of nitrogen fixing crops is considered a challenge that could lead to a new agricultural “green” revolution. Here we report the use of synthetic biology tools to achieve and optimize the production of active nitrogenase Fe protein (NifH) in the chloroplasts of tobacco plants. Azotobacter vinelandii nitrogen fixation genes nifH, M, U and S were re‐designed for protein accumulation in tobacco cells. Targeting to the chloroplast was optimized by screening and identifying minimal length transit peptides performing properly for each specific Nif protein. Putative peptidyl prolyl cis‐trans isomerase NifM proved necessary for NifH solubility in the stroma. Purified NifU, a protein involved in the biogenesis of NifH [4Fe‐4S] cluster, was found functional in NifH reconstitution assays. Importantly, NifH purified from tobacco chloroplasts was active in the reduction of acetylene to ethylene, with the requirement of NifU and NifS co‐expression. These results support the suitability of chloroplasts of photosynthetically active plants to host functional nitrogenase proteins, paving the way for future studies in the engineering of nitrogen fixation in higher plant plastids and describing an optimization pipeline that could also be used in other organisms and in the engineering of new metabolic pathways in plastids.

    更新日期:2020-01-27
  • A CRISPR‐Cas9‐mediated domain‐specific base‐editing screen enables functional assessment of ACCase variants in rice
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-27
    Xiaoshuang Liu; Ruiying Qin; Juan Li; Shengxiang Liao; Tiaofeng Shan; Rongfang Xu; Dexiang Wu; Pengcheng Wei

    The evolution and practical breeding of crops depend on genetic variations. Conventionally, these variations result from natural mutations or physical and chemical mutagenesis, both of which occur randomly and lack direction. To accelerate crop improvement, targeted mutagenesis methods are highly desired. Clustered regularly interspaced palindromic repeat (CRISPR)‐Cas9 systems have been engineered for genome‐targeted mutagenesis in eukaryotic organisms, including plants (Chen et al., 2019).

    更新日期:2020-01-27
  • SlGRAS4 mediates a novel regulatory pathway promoting chilling tolerance in tomato
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-25
    Yudong Liu; Yuan Shi; Ning Zhu; Silin Zhong; Mondher Bouzayen; Zhengguo Li

    Tomato (Solanum lycopersicum L.) plants are cold‐sensitive, and the fruit are susceptible to postharvest chilling injury when stored at low temperature. However, the mechanisms underlying cold stress responses in tomato are poorly understood. We demonstrate that SlGRAS4, encoding a transcription factor induced by low temperature, promotes chilling tolerance in tomato leaves and fruit. Combined genome‐wide ChIP‐seq and RNA‐seq approaches identified among cold stress‐associated genes those being direct targets of SlGRAS4 and protein studies revealed that SlGRAS4 forms a homodimer to self‐activate its own promoter. SlGRAS4 can also directly bind tomato SlCBF promoters to activate their transcription without inducing any growth retardation. The study identifies the SlGRAS4‐regulon as a new cold response pathway conferring cold stress tolerance in tomato independently of the ICE1‐CBF pathway. This provides new track for breeding strategies aiming to improve chilling tolerance of cultivated tomatoes and to preserve sensory qualities of tomato fruit often deteriorated by storage at low temperatures.

    更新日期:2020-01-26
  • miR156a‐targeted SBP‐Box transcription factor SlSPL13 regulates inflorescence morphogenesis by directly activating SFT in tomato
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-25
    Long Cui; Fangyan Zheng; Jiafa Wang; Chunli Zhang; Fangming Xiao; Jie Ye; Changxing Li; Zhibiao Ye; Junhong Zhang

    The inflorescences and lateral branches of higher plants are generated by lateral meristems. The structure of the inflorescence has a direct effect on fruit yield in tomato (Solanum lycopersicum). We previously demonstrated that miR156a plays important roles in determining the structures of the inflorescences and lateral branches in tomato by suppressing the expression of the SQUAMOSA PROMOTER BINDING PROTEIN LIKE (SPL) transcription factor gene family. However, information on regulatory pathways associated with inflorescence morphogenesis is still lacking. In this study, we demonstrate that SPL13 is the major SPL involved in miR156a‐regulated tomato inflorescence structure determination and lateral branch production. Suppressing the expression of SPL13 in tomato increases the number of inflorescences on vegetative branches and lateral branches, decreases the number of flowers and fruit, and reduces fruit size and yield. Genetic and biochemical evidence indicate that SPL13 controls inflorescence development by positively regulating the expression of the tomato inflorescence‐associated gene SINGLE FLOWER TRUSS (SFT) by directly binding to its promoter region. Thus, our findings provide a major advance to our understanding of the miR156a‐SlSPL‐based mechanism that regulates plant architecture and yield in tomato.

    更新日期:2020-01-26
  • Genetic and signalling pathways of dry fruit size: targets for genome editing‐based crop improvement
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-25
    Quaid Hussain; Jiaqin Shi; Armin Scheben; Jiepeng Zhan; Xinfa Wang; Guihua Liu; Guijun Yan; Graham J. King; David Edwards; Hanzhong Wang

    Fruit is seed‐bearing structures specific to angiosperm that form from the gynoecium after flowering. Fruit size is an important fitness character for plant evolution and an agronomical trait for crop domestication/improvement. Despite the functional and economic importance of fruit size, the underlying genes and mechanisms are poorly understood, especially for dry fruit types. Improving our understanding of the genomic basis for fruit size opens the potential to apply gene‐editing technology such as CRISPR/Cas to modulate fruit size in a range of species. This review examines the genes involved in the regulation of fruit size and identifies their genetic/signalling pathways, including the phytohormones, transcription and elongation factors, ubiquitin‐proteasome and microRNA pathways, G‐protein and receptor kinases signalling, arabinogalactan and RNA‐binding proteins. Interestingly, different plant taxa have conserved functions for various fruit size regulators, suggesting that common genome edits across species may have similar outcomes. Many fruit size regulators identified to date are pleiotropic and affect other organs such as seeds, flowers and leaves, indicating a coordinated regulation. The relationships between fruit size and fruit number/seed number per fruit/seed size, as well as future research questions, are also discussed.

    更新日期:2020-01-26
  • A polysaccharide deacetylase from Puccinia striiformis f. sp. tritici is an important pathogenicity gene that suppresses plant immunity
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-25
    Qiang Xu; Jianfeng Wang; Jinren Zhao; Shutian Sun; Huifei Zhang; JiaJie Wu; Chunlei Tang; Zhensheng Kang; Xiaojie Wang

    The cell wall of filamentous fungi, comprised of chitin, polysaccharide and glycoproteins, maintain the integrity of hyphae and protect them from defense responses by potential host plants. Here we report that one polysaccharide deacetylase of Puccinia striiformis f. sp. tritici (Pst), Pst_13661 suppresses Bax‐induced cell death in plants and Pst_13661 is highly induced during early stages of the interaction between wheat and Pst. Importantly, the transgenic wheat expressing the RNA interference (RNAi) construct of Pst_13661 exhibits high resistance to major Pst epidemic races CYR31, CYR32 and CYR33 by inhibiting growth and development of Pst, indicating that Pst_13661 is an available pathogenicity factor and is a potential target for generating broad‐spectrum resistance breeding material of wheat. It forms a homo‐polymer and has high affinity for chitin and germ tubes of Pst compared with the control. Besides, Pst_13661 suppresses chitin‐induced plant defense in plants. Hence, we infer that Pst_13661 may modify the fungal cell wall to prevent recognition by apoplastic surveillance systems in plants. This study opens new approaches for developing durable disease‐resistant germplasm by disturbing the growth and development of fungi and develop novel strategies to control crop diseases.

    更新日期:2020-01-26
  • Natural variation and CRISPR/Cas9‐mediated mutation in GmPRR37 affect photoperiodic flowering and contribute to regional adaptation of soybean
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-25
    Liwei Wang; Shi Sun; Tingting Wu; Luping Liu; Xuegang Sun; Yupeng Cai; Jicun Li; Hongchang Jia; Shan Yuan; Li Chen; Bingjun Jiang; Cunxiang Wu; Wensheng Hou; Tianfu Han

    Flowering time is a critical determinant of the geographic distribution and regional adaptability of soybean (Glycine max), and is strongly regulated by photoperiod and temperature. In this study, quantitative trait locus (QTL) mapping and subsequent candidate gene analysis revealed that GmPRR37, encoding a pseudo‐response regulator protein, is responsible for the major QTL qFT12‐2, which was identified from a population of 308 recombinant inbred lines (RILs) derived from a cross between a very late‐flowering soybean cultivar, ‘Zigongdongdou (ZGDD)’, and an extremely early‐flowering cultivar, ‘Heihe27 (HH27)’, in multiple environments. Comparative analysis of parental sequencing data confirmed that HH27 contains a nonsense mutation that causes the loss of the CCT domain in the GmPRR37 protein. CRISPR/Cas9‐induced Gmprr37‐ZGDD mutants in soybean exhibited early flowering under natural long‐day (NLD) conditions. Overexpression of GmPRR37 significantly delayed the flowering of transgenic soybean plants compared with wild type under long photoperiod conditions. In addition, both the knockout and overexpression of GmPRR37 in soybean showed no significant phenotypic alterations in flowering time under short‐day (SD) conditions. Furthermore, GmPRR37 downregulated the expression of the flowering‐promoting FT homologs GmFT2a and GmFT5a, and upregulated flowering‐inhibiting FT homolog GmFT1a expression under long‐day (LD) conditions. We analyzed haplotypes of GmPRR37 among 180 cultivars collected across China, and found natural Gmprr37 mutants flower earlier and enable soybean to be cultivated at higher latitudes. This study demonstrates that GmPRR37 controls soybean photoperiodic flowering and provides opportunities to breed optimized cultivars with adaptation to specific regions and farming systems.

    更新日期:2020-01-26
  • HvAKT2 and HvHAK1 confer drought tolerance in barley through enhanced leaf mesophyll H+ homoeostasis
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-24
    Xue Feng; Wenxing Liu; Cheng‐Wei Qiu; Fanrong Zeng; Yizhou Wang; Guoping Zhang; Zhong‐Hua Chen; Feibo Wu

    Plant K+ uptake typically consists low—affinity mechanisms mediated by Shaker K+ channels (AKT/KAT/KC) and high‐affinity mechanisms regulated by HAK/KUP/KT transporters, which are extensively studied. However, the evolutionary and genetic roles of both K+ uptake mechanisms for drought tolerance are not fully explored in crops adapted to dryland agriculture. Here, we employed evolutionary bioinformatics, biotechnological and electrophysiological approaches to determine the role of two important K+ transporters HvAKT2 and HvHAK1 in drought tolerance in barley. HvAKT2 and HvHAK1 were cloned and functionally characterized using barley stripe mosaic virus‐induced gene silencing (BSMV‐VIGS) in drought‐tolerant wild barley XZ5 and agrobacterium‐mediated gene transfer in the barley cultivar Golden Promise. The hallmarks of the K+ selective filters of AKT2 and HAK1 are both found in homologues from strepotophyte algae, and they are evolutionarily conserved in strepotophyte algae and land plants. HvAKT2 and HvHAK1 are both localized to the plasma membrane and have high selectivity to K+ and Rb+ over other tested cations. Overexpression of HvAKT2 and HvHAK1 enhanced K+ uptake and H+ homoeostasis leading to drought tolerance in these transgenic lines. Moreover, HvAKT2‐ and HvHAK1‐overexpressing lines showed distinct response of K+, H+ and Ca2+ fluxes across plasma membrane and production of nitric oxide and hydrogen peroxide in leaves as compared to the wild type and silenced lines. High‐ and low‐affinity K+ uptake mechanisms and their coordination with H+ homoeostasis play essential roles in drought adaptation of wild barley. These findings can potentially facilitate future breeding programs for resilient cereal crops in a changing global climate.

    更新日期:2020-01-24
  • The major‐effect quantitative trait locus Fnl7.1 encodes a late embryogenesis abundant protein associated with fruit neck length in cucumber
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-24
    Xuewen Xu; Chenxi Wei; Qianya Liu; Wenqing Qu; Xiaohua Qi; Qiang Xu; Xuehao Chen

    Fruit neck length (FNL) is an important quality trait in cucumber because it directly affects its market value. However, its genetic basis remains largely unknown. We identified a candidate gene for FNL in cucumber using a next‐generation sequencing‐based bulked segregant analysis in F2 populations, derived from a cross between Jin5‐508 (long necked) and YN (short necked). A quantitative trait locus (QTL) on chromosome 7, Fnl7.1, was identified through a genome‐wide comparison of single nucleotide polymorphisms between long and short FNL F2 pools, and it was confirmed by traditional QTL mapping in multiple environments. Fine genetic mapping, sequences alignment and gene expression analysis revealed that CsFnl7.1 was the most likely candidate Fnl7.1 locus, which encodes a late embryogenesis abundant protein. The increased expression of CsFnl7.1 in long‐necked Jin5‐508 may be attributed to mutations in the promoter region upstream of the gene body. The function of CsFnl7.1 in FNL control was confirmed by its overexpression in transgenic cucumbers. CsFnl7.1 regulates fruit neck development by modulating cell expansion. Probably, this is achieved through the direct protein–protein interactions between CsFnl7.1 and a dynamin‐related protein CsDRP6 and a germin‐like protein CsGLP1. Geographical distribution differences of the FNL phenotype were found among the different cucumber types. The East Asian and Eurasian cucumber accessions were highly enriched with the long‐necked and short‐necked phenotypes, respectively. A further phylogenetic analysis revealed that the Fnl7.1 locus might have originated from India. Thus, these data support that the CsFnl7.1 has an important role in increasing cucumber FNL.

    更新日期:2020-01-24
  • The genome of kenaf (Hibiscus cannabinus L.) provides insights into bast fiber and leaf shape biogenesis
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-23
    Liwu Zhang; Yi Xu; Xingtan Zhang; Xiaokai Ma; Lilan Zhang; Zhenyang Liao; Qing Zhang; Xuebei Wan; Yan Cheng; Jisen Zhang; Dongxu Li; Liemei Zhang; Jiantang Xu; Aifen Tao; Lihui Lin; Pingping Fang; Shuai Chen; Rui Qi; Xiuming Xu; Jianmin Qi; Ray Ming

    Kenaf is an ancient crop that is widely cultivated as a source of bast (phloem) fibers, the phytoremediation of heavy metal‐contaminated farmlands, and textile relevant compounds. Leaf shape played a unique role in kenaf improvement, due to the inheritance as a single locus and the association with fiber development in typical lobed‐leaf varieties. Here we report a high‐quality genome assembly and annotation for var. ‘Fuhong 952’ with 1,078 Mbp genome and 66,004 protein‐coding genes integrating single‐molecule real‐time sequencing, a high‐density genetic map, and high‐throughput chromosome conformation capture techniques. Gene mapping assists the identification of a homeobox transcription factor LATE MERISTEM IDENTITY 1 (HcLMI1) gene controlling lobed‐leaf. Virus‐induced gene silencing (VIGS) of HcLMI1 in a lobed‐leaf variety was critical to induce round‐like leaf formation. Candidate genes involved in cell wall formation were found in quantitative trait loci (QTL) for fiber yield and quality related traits. Comparative genomic and transcriptome analyses revealed key genes involved in bast fiber formation, among which there are twice as many cellulose synthase A (CesA) genes due to a recent whole‐genome duplication after divergence from Gossypium. Population genomic analysis showed two recent population bottlenecks in kenaf, suggesting domestication and improvement process have led to an increase in fiber biogenesis and yield. This chromosome‐scale genome provides an important framework and toolkit for sequence‐directed genetic improvement of fiber crops.

    更新日期:2020-01-24
  • CmBBX8 accelerates flowering by targeting CmFTL1 directly in summer chrysanthemum
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-22
    Lijun Wang; Jing Sun; Liping Ren; Min Zhou; Xiaoying Han; Lian Ding; Fei Zhang; Zhiyong Guan; Weimin Fang; Sumei Chen; Fadi Chen; Jiafu Jiang

    For a flowering plant, the transition from vegetative stage to reproductive growth is probably the most critical developmental switch. In the model plant Arabidopsis thaliana, the product of BBX7, group II member of BBX family, acts to delay floral transition. In this study, a presumed chrysanthemum homolog of a second group gene AtBBX8, designated CmBBX8, had been isolated and characterized. The transcription of CmBBX8 followed a diurnal rhythm as the chrysanthemum floral transition regulator. Overexpression of CmBBX8 accelerated flowering, while its (artificial microRNAs) amiR‐enabled knockdown delayed flowering in plants grown under both long‐ and short‐day conditions. Global expression analysis revealed that genes associated with photoperiod were down‐regulated in amiR‐CmBBX8 lines compared with the wild type, which were verified to be up‐regulated in overexpressing lines (OX‐CmBBX8) by RT‐PCR. A number of in vitro assays were used to show that CmBBX8 targets CmFTL1. Furthermore, the function of CmFTL1 as a floral inducer under long‐day conditions was confirmed by the behaviour of engineered summer‐flowering chrysanthemum plants. The conclusion is that the BBX8‐FT regulatory module is an important determinant of reproductive development in summer‐flowering chrysanthemum.

    更新日期:2020-01-23
  • DcMYB113, a root‐specific R2R3‐MYB, conditions anthocyanin biosynthesis and modification in carrot
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-22
    Zhi‐Sheng Xu; Qing‐Qing Yang; Kai Feng; Xiao Yu; Ai‐Sheng Xiong

    Purple carrots, the original domesticated carrots, accumulate highly glycosylated and acylated anthocyanins in root and/or petiole. Previously, a quantitative trait locus (QTL) for root‐specific anthocyanin pigmentation was genetically mapped to chromosome 3 of carrot. In this study, an R2R3‐MYB gene, namely DcMYB113, was identified within this QTL region. DcMYB113 expressed in the root of ‘Purple haze’, a carrot cultivar with purple root and nonpurple petiole, but not in the roots of two carrot cultivars with a purple root and petiole (Deep purple and Cosmic purple) and orange carrot ‘Kurodagosun’, which appeared to be caused by variation in the promoter region. The function of DcMYB113 from ‘Purple haze’ was verified by transformation in ‘Cosmic purple’ and ‘Kurodagosun’, resulting in anthocyanin biosynthesis. Transgenic ‘Kurodagosun’ carrying DcMYB113 driven by the CaMV 35S promoter had a purple root and petiole, while transgenic ‘Kurodagosun’ expressing DcMYB113 driven by its own promoter had a purple root and nonpurple petiole, suggesting that root‐specific expression of DcMYB113 was determined by its promoter. DcMYB113 could activate the expression of DcbHLH3 and structural genes related to anthocyanin biosynthesis. DcUCGXT1 and DcSAT1, which were confirmed to be responsible for anthocyanins glycosylation and acylation, respectively, were also activated by DcMYB113. The WGCNA identified several genes co‐expressed with anthocyanin biosynthesis and the results indicated that DcMYB113 may regulate anthocyanin transport. Our findings provide insight into the molecular mechanism underlying root‐specific anthocyanin biosynthesis and further modification in carrot and even other root crops.

    更新日期:2020-01-23
  • The gland localized CGP1 controls gland pigmentation and gossypol accumulation in cotton
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-21
    Wei Gao; Fu‐Chun Xu; Lu Long; Yang Li; Jun‐Li Zhang; Leelyn Chong; Jose Ramon Botella; Chun‐Peng Song

    Pigment glands, also known as black glands or gossypol glands, are specific for Gossypium spp. These glands strictly confine large amounts of secondary metabolites to the lysigenous cavity, leading to the glands’ intense colour and providing defence against pests and pathogens. This study performed a comparative transcriptome analysis of glanded versus glandless cotton cultivars. Twenty‐two transcription factors showed expression patterns associated with pigment glands and were characterized. Phenotypic screening of the genes, via virus‐induced gene silencing, showed an apparent disappearance of pigmented glands after the silencing of a pair of homologous MYB‐encoding genes in the A and D genomes (designated as CGP1). Further study showed that CGP1a encodes an active transcription factor, which is specifically expressed in the gland structure, while CGP1d encodes a non‐functional protein due to a fragment deletion, which causes premature termination. RNAi‐mediated silencing and CRISPR knockout of CGP1 in glanded cotton cultivars generated a glandless‐like phenotype, similar to the dominant glandless mutant Gl2e. Microscopic analysis showed that CGP1 knockout did not affect gland structure or density, but affected gland pigmentation. The levels of gossypol and related terpenoids were significantly decreased in cgp1 mutants, and a number of gossypol biosynthetic genes were strongly down‐regulated. CGP1 is located in the nucleus where it interacts with GoPGF, a critical transcription factor for gland development and gossypol synthesis. Our data suggest that CGP1 and GoPGF form heterodimers to control the synthesis of gossypol and other secondary metabolites in cotton.

    更新日期:2020-01-22
  • Metabolic rewiring of synthetic pyruvate dehydrogenase bypasses for acetone production in cyanobacteria
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-20
    Hyun Jeong Lee; Jigyeong Son; Sang Jun Sim; Han Min Woo

    Designing synthetic pathways for efficient CO2 fixation and conversion is essential for sustainable chemical production. Here we have designed a synthetic acetate‐acetyl‐CoA/malonyl‐CoA (AAM) bypass to overcome an enzymatic activity of pyruvate dehydrogenase complex. This synthetic pathway utilizes acetate assimilation and carbon rearrangements using a methyl malonyl‐CoA carboxyltransferase. We demonstrated direct conversion of CO2 into acetyl‐CoA‐derived acetone as an example in photosynthetic Synechococcus elongatus PCC 7942 by increasing the acetyl‐CoA pools. The engineered cyanobacterial strain with the AAM‐bypass produced 0.41 g L‐1 of acetone at 0.71 mmol L‐1 d‐1 of molar productivity. This work clearly shows that the synthetic pyruvate dehydrogenase bypass (AAM‐bypass) is a key factor for the high‐level production of an acetyl‐CoA‐derived chemical in photosynthetic organisms.

    更新日期:2020-01-22
  • A pathogenesis related protein GmPR08‐Bet VI promotes a molecular interaction between the GmSHMT08 and GmSNAP18 in resistance to Heterodera glycines
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-20
    Naoufal Lakhssassi; Sarbottam Piya; Sadia Bekal; Shiming Liu; Zhou Zhou; Catherine Bergounioux; Long Miao; Jonas Meksem; Aicha Lakhssassi; Karen Jones; My Abdelmajid Kassem; Moussa Benhamed; Abdelhafid Bendahmane; Kris Lambert; Adnane Boualem; Tarek Hewezi; Khalid Meksem

    Soybean cyst nematode (SCN, Heterodera glycines) is the most devastating pest affecting soybean production worldwide. SCN resistance requires both the GmSHMT08 and GmSNAP18 in “Peking” type resistance. Here we describe the molecular interaction between GmSHMT08 and GmSNAP18, which is potentiated by a pathogenesis‐related protein GmPR08‐Bet VI. Like GmSNAP18 and GmSHMT08, GmPR08‐Bet VI expression was induced in response to SCN and its overexpression decreased SCN cysts by 65% in infected transgenic soybean roots. Overexpression of GmPR08‐Bet VI did not have an effect on SCN resistance when the two‐cytokinin binding sites in GmPR08‐Bet VI were mutated, indicating a new role of GmPR08‐Bet VI in SCN resistance. GmPR08‐Bet VI was mapped to a QTL for resistance to SCN using different mapping populations. GmSHMT08, GmSNAP18, and GmPR08‐Bet VI localize to the cytosol and plasma membrane. GmSNAP18 expression and localization hyper‐accumulated at the plasma membrane and was specific to the root cells surrounding the nematode in SCN resistant soybeans. Genes encoding key components of the salicylic acid signaling pathway were induced under SCN infection. GmSNAP18 and GmPR08‐Bet VI were also induced under salicylic acid and cytokinin exogenous treatments, while GmSHMT08 was induced only when the resistant GmSNAP18 was present, pointing to the presence of a molecular crosstalk between SCN resistant genes and defense genes. Expression analysis of GmSHMT08 and GmSNAP18 identified the need of a minimum expression requirement to trigger the SCN resistance reaction. These results provide insight into a new response mechanism toward plant nematode resistance involving haplotype compatibility, gene dosage, and hormone signaling.

    更新日期:2020-01-22
  • Genetic modulation of RAP alters fruit coloration in both wild and cultivated strawberry
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-19
    Qi Gao; Huifeng Luo; Yongping Li; Zhongchi Liu; Chunying Kang

    Fruit colour affects consumer preference and is an important trait for breeding in strawberry. Previously, we isolated the Reduced Anthocyanins in Petioles (RAP) gene encoding a glutathione S‐transferase (GST) that binds anthocyanins to facilitate their transport from cytosol to vacuole in the diploid strawberry Fragaria vesca. The parent of rap was the F. vesca variety ‘Yellow Wonder’ that develops white fruit due to a natural mutation in the FveMYB10 gene. Here, we investigated the application potential of RAP in modulating fruit colours by overexpression of RAP in F. vesca and knockout of RAP in the cultivated strawberry Fragaria × ananassa. Unexpectedly, the RAP overexpression in Yellow Wonder background caused formation of red fruit. In addition, the red coloration occurs precociously at floral stage 10 and continues in the receptacle during early fruit development. Transcriptome analysis revealed that the anthocyanin biosynthesis genes were not up‐regulated in RAP‐ox; rap myb10 flowers at anthesis and largely inhibited at the turning stage in fruit, suggesting a coloration mechanism independent of FveMYB10. Moreover, we used CRISPR/Cas9 to knockout RAP in cultivated strawberry which is octoploid. Six copies of RAP were simultaneously knocked out in the T0 generation leading to the green stem and white‐fruited phenotype. Several T1 progeny have segregated away the CRISPR/Cas9 transgene but maintain the green stem trait. Our results indicate that enhancing the anthocyanin transport could redirect the metabolic flux from proanthocyanidin to anthocyanin production at early developmental stages of fruit and that RAP is one promising candidate gene in fruit colour breeding of strawberry.

    更新日期:2020-01-21
  • PINOID regulates floral organ development by modulating auxin transport and interacts with MADS16 in rice
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-17
    Hua‐Mao Wu; Dong‐Jiang Xie; Zuo‐Shun Tang; Dong‐Qiao Shi; Wei‐Cai Yang

    In rice (Oryza sativa L.), floral organ development is an important trait. Although a role for PINOID in regulating floral organ development was reported recently, the underlying molecular mechanism remains unclear. Here, we isolated and characterized an abnormal floral organ mutant and mapped the causative gene through an improved MutMap method. Molecular study revealed that the observed phenotype is caused by a point mutation in OsPINOID (OsPID) gene, therefore we named the mutation as ospid‐4. Our data demonstrate that OsPID interacts with OsPIN1a and OsPIN1b to regulate polar auxin transport as shown previously. Additionally, OsPID also interacts with OsMADS16 to regulate transcription during floral organ development in rice. Together, we propose a model that OsPID regulates floral organ development by modulating auxin polar transport and interaction with OsMADS16 and/or LAX1 in rice. These results provide a novel insight into the role of OsPID in regulating floral organ development of rice, especially in stigma development, which would be useful for genetic improvement of high‐yield breeding of rice.

    更新日期:2020-01-17
  • Overexpression of GmAAP6a enhances tolerance to low nitrogen and improves seed nitrogen status by optimizing amino acid partitioning in soybean
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-16
    Sheng Liu; Dan Wang; Yuanyuan Mei; Tongmei Xia; Wei Xu; Yuqing Zhang; Xiang You; Xiyu Zhang; Lei Li; Ning Ning Wang

    Amino acid transport via phloem is one of the major source‐to‐sink nitrogen translocation pathways in most plant species. Amino acid permeases (AAPs) play essential roles in amino acid transport between plant cells and subsequent phloem or seed loading. In this study, a soybean AAP gene, annotated as GmAAP6a, was cloned and demonstrated to be significantly induced by nitrogen starvation. Histochemical staining of GmAAP6a:GmAAP6a‐GUS transgenic soybean revealed that GmAAP6a is predominantly expressed in phloem and xylem parenchyma cells. Growth and transport studies using toxic amino acid analogs or single amino acids as a sole nitrogen source suggest that GmAAP6a can selectively absorb and transport neutral and acidic amino acids. Overexpression of GmAAP6a in Arabidopsis and soybean resulted in elevated tolerance to nitrogen limitation. Furthermore, the source‐to‐sink transfer of amino acids in the transgenic soybean was markedly improved under low nitrogen conditions. At the vegetative stage, GmAAP6a‐overexpressing soybean showed significantly increased nitrogen export from source cotyledons and simultaneously enhanced nitrogen import into sink primary leaves. At the reproductive stage, nitrogen import into seeds was greatly enhanced under both sufficient and limited nitrogen conditions. Collectively, our results imply that overexpression of GmAAP6a enhances nitrogen stress tolerance and source‐to‐sink transport and improves seed quality in soybean. Co‐expression of GmAAP6a with genes specialized in source nitrogen recycling and seed loading may represent an interesting application potential in breeding.

    更新日期:2020-01-17
  • Balancing the double‐edged sword effect of increased resistant starch content and its impact on rice texture: Its genetics and molecular physiological mechanisms
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-16
    Sabiha Parween; Joanne J. Anonuevo; Vito Butardo; Gopal Misra; Roslen Anacleto; Cindy Llorente; Ondrej Kosik; Marissa V. Romero; Evelyn H. Bandonill; Merlyn S. Mendioro; Alison Lovegrove; Alisdair R. Fernie; Yariv Brotman; Nese Sreenivasulu

    Resistant starch (RS) is the portion of starch that escapes gastrointestinal digestion and acts as a substrate for fermentation of probiotic bacteria in the gut. Aside from enhancing gut health, RS contributes to a lower glycemic index. A genome‐wide association study coupled with targeted gene association studies was conducted utilizing a diverse panel of 281 re‐sequenced Indica rice lines comprising of ~2.2 million single nucleotide polymorphisms. Low‐to‐intermediate RS phenotypic variations were identified in the rice diversity panel, resulting in novel associations of RS to several genes associated with amylopectin biosynthesis and degradation. Selected rice lines encoding superior alleles of SSIIa with medium RS and inferior alleles with low RS groups were subjected to detailed transcriptomic, metabolomic, non‐starch dietary fiber (DF), starch structural and textural attributes. The gene regulatory networks highlighted the importance of a protein phosphatase alongside multiple genes of starch metabolism. Metabolomics analyses resulted in the identification of several metabolite hubs (carboxylic acid, sugars and polyamines) in the medium RS group. Among DF, mannose and galactose from the water‐insoluble fraction were found to be highly associated with low and medium RS lines, respectively. Starch structural analyses revealed that a moderate increase in RS is also linked to an elevation of amylose 1 and amylose 2 fractions. Although rice lines with medium RS content negatively affected textural and viscosity properties in comparison to low RS, the textural property of medium RS lines was in the same acceptable range as IR64, a rice mega variety popular in Asia.

    更新日期:2020-01-17
  • BGAL1 depletion boosts the level of β‐galactosylation of N‐ and O‐glycans in N. benthamiana
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-11
    Ricarda Kriechbaum; Esmaiel Ziaee; Clemens Grünwald‐Gruber; Pierre Buscaill; Renier A. L. van der Hoorn; Alexandra Castilho

    Glyco‐design of proteins is a powerful tool in fundamental studies of structure–function relationship and in obtaining profiles optimized for efficacy of therapeutic glycoproteins. Plants, particularly Nicotiana benthamiana, are attractive hosts to produce recombinant glycoproteins, and recent advances in glyco‐engineering facilitate customized N‐glycosylation of plant‐derived glycoproteins. However, with exception of monoclonal antibodies, homogenous human‐like β1,4‐galactosylation is very hard to achieve in recombinant glycoproteins. Despite significant efforts to optimize the expression of β1,4‐galactosyltransferase, many plant‐derived glycoproteins still exhibit incomplete processed N‐glycans with heterogeneous terminal galactosylation. The most obvious suspects to be involved in trimming terminal galactose residues are β‐galactosidases (BGALs) from the glycosyl hydrolase family GH35. To elucidate the so far uncharacterized mechanisms leading to the trimming of terminal galactose residues from glycans of secreted proteins, we studied a N. benthamiana BGAL known to be active in the apoplast (NbBGAL1). Here, we determined the NbBGAL1 subcellular localization, substrate specificity and in planta biological activity. We show that NbBGAL1 can remove β1,4‐ and β1,3‐galactose residues on both N‐ and O‐glycans. Transient BGAL1 down‐regulation by RNA interference (RNAi) and BGAL1 depletion by genome editing drastically reduce β‐galactosidase activity in N. benthamiana and increase the amounts of fully galactosylated complex N‐glycans on several plant‐produced glycoproteins. Altogether, our data demonstrate that NbBGAL1 acts on galactosylated complex N‐glycans of plant‐produced glycoproteins.

    更新日期:2020-01-13
  • Metabolite‐based genome‐wide association study enables dissection of the flavonoid decoration pathway of wheat kernels
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-12
    Jie Chen; Xin Hu; Taotao Shi; Huanran Yin; Dongfa Sun; Hao Yuanfeng; Xianchun Xia; Jie Luo; Alisdair R. Fernie; Zhonghu He; Wei Chen

    The marriage of metabolomic approaches with genetic design has proven a powerful tool in dissecting diversity in the metabolome and has additionally enhanced our understanding of complex traits. That said, such studies have rarely been carried out in wheat. In this study, we detected 805 metabolites from wheat kernels and profiled their relative contents among 182 wheat accessions, conducting a metabolite‐based genome‐wide association study (mGWAS) utilizing 14,646 previously described polymorphic SNP markers. A total of 1,098 mGWAS associations were detected with large effects, within which 26 candidate genes were tentatively designated for 42 loci. Enzymatic assay of two candidates indicated they could catalyze glucosylation and subsequent malonylation of various flavonoids and thereby the major flavonoid decoration pathway of wheat kernel was dissected. Moreover, numerous high‐confident genes associated with metabolite contents have been provided, as well as more subdivided metabolite networks which are yet to be explored within our data. These combined efforts presented the first step towards realizing metabolomics‐associated breeding of wheat.

    更新日期:2020-01-13
  • Methylation of MdMYB1 locus mediated by RdDM pathway regulates anthocyanin biosynthesis in apple
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-12
    Shenghui Jiang; Nan Wang; Min Chen; Rui Zhang; Qingguo Sun; Haifeng Xu; Zongying Zhang; Yicheng Wang; Xiuqi Sui; Sufang Wang; Hongcheng Fang; Weifang Zuo; Mengyu Su; Jing Zhang; Zhangjun Fei; Xuesen Chen

    Methylation at the MdMYB1 promoter in apple sports has been reported as a regulator of the anthocyanin pathway, but little is known about how the locus is recognized by the methylation machinery to regulate anthocyanin accumulation. In this study, we analyzed three differently colored ‘Fuji' apples and found that differences in the transcript levels of MdMYB1, which encodes a key regulator of anthocyanin biosynthesis, control the anthocyanin content (and therefore, color) in fruit skin. The CHH methylation levels in the MR3 region (−1,246 to −780) of the MdMYB1 promoter were found to be negatively correlated with MdMYB1 expression. Thus, they were ideal materials to study DNA methylation in apple sports. The protein of RNA‐directed DNA methylation (RdDM) pathway responsible for CHH methylation, MdAGO4, was found to interact with the MdMYB1 promoter. MdAGO4s can interact with MdRDM1 and MdDRM2s to form an effector complex, fulfilling CHH methylation. When MdAGO4s and MdDRM2s were overexpressed in apple calli and Arabidopsis mutants, those proteins increase the CHH methylation of AGO4‐binding sites. In electrophoretic mobility shift assays, MdAGO4s were found to specifically bind to sequence containing ATATCAGA. Knock‐down of MdNRPE1 did not affect the binding of MdAGO4s to the c3 region of the MdMYB1 promoter in 35S::AGO4s calli. Taken together, our data show that the MdMYB1 locus is methylated through binding of MdAGO4s to the MdMYB1 promoter to regulate anthocyanin biosynthesis by the RdDM pathway.

    更新日期:2020-01-13
  • Expression of the CCCH‐tandem zinc finger protein gene OsTZF5 under a stress inducible promoter mitigates the effect of drought stress on rice grain yield under field conditions
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-12
    Michael Gomez Selvaraj; Asad Jan; Takuma Ishizaki; Milton Valencia; Beata Dedicova; Kyonoshin Maruyama; Takuya Ogata; Daisuke Todaka; Kazuko Yamaguchi‐Shinozaki; Kazuo Nakashima; Manabu Ishitani

    Increasing drought resistance without sacrificing grain yield remains an ongoing challenge in crop improvement. In this study, we report that Oryza sativa CCCH tandem zinc finger protein 5 (OsTZF5) can confer drought resistance and increase grain yield in transgenic rice plants. Expression of OsTZF5 was induced by abscisic acid, dehydration, and cold stress. Upon stress, OsTZF5‐GFP localized to the cytoplasm and cytoplasmic foci. Transgenic rice plants overexpressing OsTZF5 under the constitutive maize ubiquitin promoter exhibited improved survival under drought but also growth retardation. By introducing OsTZF5 behind the stress‐responsive OsNAC6 promoter in two commercial upland cultivars, Curinga and NERICA4, we obtained transgenic plants that showed no growth retardation. Moreover, these plants exhibited significantly increased grain yield compared to non‐transgenic cultivars in different confined field drought environments. Physiological analysis indicated that OsTZF5 promoted both drought tolerance and drought avoidance. Collectively, our results provide strong evidence that OsTZF5 is a useful biotechnological tool to minimize yield losses in rice grown under drought conditions.

    更新日期:2020-01-13
  • Genome‐wide analysis of epigenetic and transcriptional changes associated with heterosis in pigeonpea
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-10
    Pallavi Sinha; Vikas K Singh; Rachit K Saxena; Sandip M Kale; Yuqi Li; Vanika Garg; Meifang Tang; Aamir W Khan; Kyung Do Kim; Annapurna Chitikineni; KB Saxena; CV Sameer Kumar; Xin Liu; Xun Xu; Scott Jackson; Wayne Powell; Eviatar Nevo; Iain R Searle; Mukesh Lodha; Rajeev K Varshney

    Hybrids are extensively used in agriculture to deliver an increase in yield, yet the molecular basis of heterosis is not well understood. Global DNA methylation analysis, transcriptome analysis and small RNA profiling were aimed to understand the epigenetic effect of the changes in gene expression level in the two hybrids and their parental lines. Increased DNA methylation was observed in both the hybrids as compared to their parents. This increase DNA methylation in hybrids showed that majority of the 24‐nt sRNA clusters had higher expression in hybrids than the parents. Transcriptome analysis revealed that various phytohormones (auxin and salicylic acid) responsive hybrid‐MPV DEGs were significantly altered in both the hybrids in comparison to MPV. DEGs associated with plant immunity and growth were overexpressed whereas DEGs associated with basal defense level were repressed. This antagonistic patterns of gene expression might contribute to the greater growth of the hybrids. It was also noticed that some common as well as unique changes in regulatory pathways associated with heterotic growth in both the hybrids. Approximately 70% and 67% of down‐regulated hybrid‐MPV DEGs were found to be differentially methylated in ICPH 2671 and ICPH 2740 hybrid, respectively. This reflected the association of epigenetic regulation in altered gene expressions. Our findings also revealed that miRNAs might play important roles in hybrid vigor in both the hybrids by regulating their target genes, especially in controlling plant growth and development, defense and stress response pathways. The above finding provides an insight into the molecular mechanism of pigeonpea heterosis.

    更新日期:2020-01-13
  • Somatic embryogenesis critical initiation stage‐specific mCHH hypomethylation reveals epigenetic basis underlying embryogenic redifferentiation in cotton
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-10
    Huihui Guo; Yijie Fan; Haixia Guo; Jianfei Wu; Xiaoman Yu; Junmei Wei; Xian Lian; Li Zhang; Zhongyuan Gou; Yupeng Fan; Fanchang Zeng

    As a notable illustration of totipotency, somatic embryogenesis (SE) is the developmental reprogramming of somatic cells toward the embryogenesis pathway (Yang and Zhang, 2010). Investigations examining the totipotency process are of great fundamental and practical importance in crop biotechnology. Moreover, high‐frequency regeneration of SE has been limited due to the genotype‐dependent response. To date, the epigenetic molecular basis underlying embryogenic redifferentiation during SE remains largely unexplored.

    更新日期:2020-01-13
  • eRice: a refined epigenomic platform for japonica and indica rice
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-09
    Pingxian Zhang; Yifan Wang; Sadaruddin Chachar; Jian Tian; Xiaofeng Gu

    Epigenetic modifications including chromatin modifications and DNA methylation have emerged as essential marks that can influence various biological pathways in multicellular organisms. In plants, DNA methylation such as 5‐methylcytosine (5mC) and N6‐methyladenosine (6mA), which was discovered later, have crucial roles in the regulation of developmental cues and responses to environmental stresses. However, a database of epigenetic modifications, especially DNA methylation, was not yet available for rice (Oryza sativa). Here, we describe a species‐specific database of epigenomic annotations for rice, the eRice database (http://www.elabcaas.cn/rice/index.html), which we developed to facilitate efficient epigenomic annotation of both japonica and indica rice of two main cultivars (Nip and 93‐11) in Asia. This database integrates single‐base‐resolution 6mA and 5mC DNA methylation data at with annotations with artificial intelligence (AI) prediction data for 6mA modifications, as well as annotations for histone modifications, and genomic and transcriptomic resources in Nip and 93‐11 rice. This database will allow efficient inquiry and prediction of potential epigenetic modification sites (especially for 6mA) within targeted genes or regions. The epigenomic information in the eRice database about the distribution of epigenetic marks across rice genomes will help to provide a broader understanding of the epigenetic regulation of complex biological processes in plant development as well as a guide for future molecular design efforts.

    更新日期:2020-01-09
  • Targeted base editing in rice with CRISPR/ScCas9 system
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-08
    Meixia Wang; Ziyan Xu; Gokul Gosavi; Bin Ren; Yongsen Cao; Yongjie Kuang; Changyong Zhou; Carl Spetz; Fang Yan; Xueping Zhou; Huanbin Zhou

    The CRISPR/Cas system has rapidly become the preferred tool for genome engineering in various organisms due to high efficiency, specificity, simplicity and versatility. Currently, CRISPR/Cas‐mediated base editing, a novel genome editing strategy that enables irreversible nucleotide changes at target loci without double‐stranded DNA cleavage or any donor template, has been widely adopted for generating gain‐of‐function germplasms in functional genomics research and crop genetic improvement (Hua et al., 2019; Ren et al., 2018; Yan et al., 2018).

    更新日期:2020-01-09
  • Overexpression of GmWRI1b in soybean stably improves plant architecture and associated yield parameters, and increases total seed oil production under field conditions
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-07
    Wei Guo; Limiao Chen; Haifeng Chen; Hongli Yang; Qingbo You; Aili Bao; Shuilian Chen; Qingnan Hao; Yi Huang; Dezhen Qiu; Zhihui Shan; Zhonglu Yang; Songli Yuan; Chanjuan Zhang; Xiaojuan Zhang; Yongqing Jiao; Lam‐Son Phan Tran; Xinan Zhou; Dong Cao

    The functions of WRINKLED1 (WRI1) transcription factor in regulating fatty acid (FA) biosynthesis are highly conserved in crop plants, including maize (Zea mays) (Pouvreau et al., 2011), rapeseed (Brassica napus) (Li et al., 2015), oil palm (Elasis guineensis) (Ma et al., 2013), camelina (Camelina sativa) (An et al., 2017) and soybean (Glycine max) (Chen et al., 2018; Chen et al., 2020; Zhang et al., 2017). Recently, Kong et al. (2017) found that the WRI1 plays a role in root auxin homeostasis and affects root development in Arabidopsis, suggesting its involvement in root architecture, and potentially shoot architecture as well.

    更新日期:2020-01-08
  • Heritable temporal gene expression patterns correlate with metabolomic seed content in developing hexaploid oat seed
    Plant Biotech. J. (IF 6.840) Pub Date : 2020-01-04
    Haixiao Hu; Juan J. Gutierrez‐Gonzalez; Xinfang Liu; Trevor H. Yeats; David F. Garvin; Owen A. Hoekenga; Mark E. Sorrells; Michael A. Gore; Jean‐Luc Jannink

    Oat ranks sixth in world cereal production and has a higher content of health‐promoting compounds compared with other cereals. However, there is neither a robust oat reference genome nor transcriptome. Using deeply sequenced full‐length mRNA libraries of oat cultivar Ogle‐C, a de novo high‐quality and comprehensive oat seed transcriptome was assembled. With this reference transcriptome and QuantSeq 3′ mRNA sequencing, gene expression was quantified during seed development from 22 diverse lines across six time points. Transcript expression showed higher correlations between adjacent time points. Based on differentially expressed genes, we identified 22 major temporal co‐expression (TCoE) patterns of gene expression and revealed enriched gene ontology biological processes. Within each TCoE set, highly correlated transcripts, putatively commonly affected by genetic background, were clustered and termed genetic co‐expression (GCoE) sets. Seventeen of the 22 TCoE sets had GCoE sets with median heritabilities higher than 0.50, and these heritability estimates were much higher than that estimated from permutation analysis, with no divergence observed in cluster sizes between permutation and non‐permutation analyses. Linear regression between 634 metabolites from mature seeds and the PC1 score of each of the GCoE sets showed significantly lower p‐values than permutation analysis. Temporal expression patterns of oat avenanthramides and lipid biosynthetic genes were concordant with previous studies of avenanthramide biosynthetic enzyme activity and lipid accumulation. This study expands our understanding of physiological processes that occur during oat seed maturation and provides plant breeders the means to change oat seed composition through targeted manipulation of key pathways.

    更新日期:2020-01-04
  • My journey into the birth of plant transgenesis and its impact on modern plant biology
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-27
    Luis Herrera‐Estrella

    After finishing an M.Sc. degree in genetics and molecular biology at the Center for Research and Advanced Studies (Cinvestav) in Mexico City at the end of 1980, I decided to pursue a scientific career. Under the guidance of a great Mexican microbiologist, Dr. Jose Ruiz‐Herrera, I worked on the influence of light on cell wall biosynthesis in the phototropic response of the fungus Phycomyces blakesleanus.

    更新日期:2019-12-29
  • Overexpression of a developing xylem cDNA library in transgenic poplar generates high mutation rate specific to wood formation
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-25
    James Rauschendorfer; Yordan Yordanov; Petre Dobrev; Radomira Vankova; Robert Sykes; Carsten Külheim; Victor Busov

    We investigated feasibility of the Full‐length complementary DNA OvereXpression (FOX) system as a mutagenesis approach in poplar, using developing xylem tissue. The main goal was to assess the overall mutation rate and if the system will increase instances of mutants affected in traits linked to the xylem tissue. Indeed, we found a high mutation rate of 17.7%, whereas 80% of all mutants were significantly affected in cellulose, lignin and/or hemicellulose. Cell wall biosynthesis is a major process occurring during xylem development. Enrichment of mutants affected in cell wall composition suggests that the tissue source for the FOX library influenced the occurrence of mutants affected in a trait linked to this tissue. Additionally, we found that FLcDNAs from mutants affected in cell wall composition were homologous to genes known to be involved in cell wall biosynthesis and most recovered FLcDNAs corresponded to genes whose native expression was highest in xylem. We characterized in detail a mutant line with increased diameter. The phenotype was caused by a poplar homolog of LONELY GUY 1 (LOG1), which encodes an enzyme in cytokinin biosynthesis and significantly increased xylem proliferation. The causative role of LOG1 in the observed phenotype was further reaffirmed by elevated cytokinin concentration in the mutant and recapitulation overexpression experiment wherein multiple independent lines phenocopied the original FOX mutant. Our experiments show that the FOX approach can be efficiently used for gene discovery and molecular interrogation of traits specific to woody perennial growth and development.

    更新日期:2019-12-27
  • Dissection of genetic architecture for glucosinolate accumulations in leaves and seeds of Brassica napus by genome‐wide association study
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-25
    Sheng Liu; Huibin Huang; Xinqi Yi; Yuanyuan Zhang; Qingyong Yang; Chunyu Zhang; Chuchuan Fan; Yongming Zhou

    Glucosinolates (GSLs), whose degradation products have been shown to be increasingly important for human health and plant defence, compose important secondary metabolites found in the order Brassicales. It is highly desired to enhance pest and disease resistance by increasing the leaf GSL content while keeping the content low in seeds of Brassica napus, one of the most important oil crops worldwide. Little is known about the regulation of GSL accumulation in the leaves. We quantified the levels of 9 different GSLs and 15 related traits in the leaves of 366 accessions and found that the seed and leaf GSL content were highly correlated (r = 0.79). A total of 78 loci were associated with GSL traits, and five common and eleven tissue‐specific associated loci were related to total leaf and seed GSL content. Thirty‐six candidate genes were inferred to be involved in GSL biosynthesis. The candidate gene BnaA03g40190D (BnaA3.MYB28) was validated by DNA polymorphisms and gene expression analysis. This gene was responsible for high leaf/low seed GSL content and could explain 30.62% of the total leaf GSL variation in the low seed GSL panel and was not fixed during double‐low rapeseed breeding. Our results provide new insights into the genetic basis of GSL variation in leaves and seeds and may facilitate the metabolic engineering of GSLs and the breeding of high leaf/low seed GSL content in B. napus.

    更新日期:2019-12-27
  • Nested‐association mapping (NAM)‐based genetic dissection uncovers candidate genes for seed and pod weights in peanut (Arachis hypogaea)
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-25
    Sunil S. Gangurde; Hui Wang; Shasidhar Yaduru; Manish K. Pandey; Jake C. Fountain; Ye Chu; Thomas Isleib; C. Corley Holbrook; Alencar Xavier; Albert K. Culbreath; Peggy Ozias‐Akins; Rajeev K. Varshney; Baozhu Guo

    Multiparental genetic mapping populations such as nested‐association mapping (NAM) have great potential for investigating quantitative traits and associated genomic regions leading to rapid discovery of candidate genes and markers. To demonstrate the utility and power of this approach, two NAM populations, NAM_Tifrunner and NAM_Florida‐07, were used for dissecting genetic control of 100‐pod weight (PW) and 100‐seed weight (SW) in peanut. Two high‐density SNP‐based genetic maps were constructed with 3341 loci and 2668 loci for NAM_Tifrunner and NAM_Florida‐07, respectively. The quantitative trait locus (QTL) analysis identified 12 and 8 major effect QTLs for PW and SW, respectively, in NAM_Tifrunner, and 13 and 11 major effect QTLs for PW and SW, respectively, in NAM_Florida‐07. Most of the QTLs associated with PW and SW were mapped on the chromosomes A05, A06, B05 and B06. A genomewide association study (GWAS) analysis identified 19 and 28 highly significant SNP–trait associations (STAs) in NAM_Tifrunner and 11 and 17 STAs in NAM_Florida‐07 for PW and SW, respectively. These significant STAs were co‐localized, suggesting that PW and SW are co‐regulated by several candidate genes identified on chromosomes A05, A06, B05, and B06. This study demonstrates the utility of NAM population for genetic dissection of complex traits and performing high‐resolution trait mapping in peanut.

    更新日期:2019-12-27
  • Towards coeliac‐safe bread
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-24
    Zhiyong Zhang; Yiting Deng; Wei Zhang; Yongrui Wu; Joachim Messing

    Gluten‐free foods cannot substitute for products made from wheat flour. When wheat products are digested, the remaining peptides can trigger an autoimmune disease in 1% of the North American and European population, called coeliac disease. Because wheat proteins are encoded by a large gene family, it has been impossible to use conventional breeding to select wheat varieties that are coeliac‐safe. However, one can test the properties of protein variants by expressing single genes in coeliac‐safe cereals like maize. One source of protein that can be considered as coeliac‐safe and has bread‐making properties is teff (Eragrostis tef), a grain consumed in Ethiopia. Here, we show that teff α‐globulin3 (Etglo3) forms storage vacuoles in maize that are morphologically similar to those of wheat. Using transmission electron microscopy, immunogold labelling shows that Etglo3 is almost exclusively deposited in the storage vacuole as electron‐dense aggregates. Of maize seed storage proteins, 27‐kDa γ‐zein is co‐deposited with Etglo3. Etglo3 polymerizes via intermolecular disulphide bonds in maize, similar to wheat HMW glutenins under non‐reducing conditions. Crossing maize Etglo3 transgenic lines with α‐, β‐ and γ‐zein RNA interference (RNAi) lines reveals that Etglo3 accumulation is only dramatically reduced in γ‐zein RNAi background. This suggests that Etglo3 and 27‐kDa γ‐zein together cause storage vacuole formation and behave similar to the interactions of glutenins and gliadins in wheat. Therefore, expression of teff α‐globulins in maize presents a major step in the development of a coeliac‐safe grain with bread‐making properties.

    更新日期:2019-12-25
  • Manipulation of β‐carotene levels in tomato fruits results in increased ABA content and extended shelf life
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-24
    Gianfranco Diretto; Sarah Frusciante; Claudia Fabbri; Nicolas Schauer; Lucas Busta; Zhonghua Wang; Antonio J. Matas; Alessia Fiore; Jocelyn K.C. Rose; Alisdair R. Fernie; Reinhard Jetter; Benedetta Mattei; Jim Giovannoni; Giovanni Giuliano

    Tomato fruit ripening is controlled by the hormone ethylene and by a group of transcription factors, acting upstream of ethylene. During ripening, the linear carotene lycopene accumulates at the expense of cyclic carotenoids. Fruit‐specific overexpression of LYCOPENE β‐CYCLASE (LCYb) resulted in increased β‐carotene (provitamin A) content. Unexpectedly, LCYb‐overexpressing fruits also exhibited a diverse array of ripening phenotypes, including delayed softening and extended shelf life. These phenotypes were accompanied, at the biochemical level, by an increase in abscisic acid (ABA) content, decreased ethylene production, increased density of cell wall material containing linear pectins with a low degree of methylation, and a thicker cuticle with a higher content of cutin monomers and triterpenoids. The levels of several primary metabolites and phenylpropanoid compounds were also altered in the transgenic fruits, which could be attributed to delayed fruit ripening and/or to ABA. Network correlation analysis and pharmacological experiments with the ABA biosynthesis inhibitor, abamine, indicated that altered ABA levels were a direct effect of the increased β‐carotene content and were in turn responsible for the extended shelf life phenotype. Thus, manipulation of β‐carotene levels results in an improvement not only of the nutritional value of tomato fruits, but also of their shelf life.

    更新日期:2019-12-25
  • Cytokinin dehydrogenase: a genetic target for yield improvement in wheat
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-22
    Lei Chen; Jiqiang Zhao; Jiancheng Song; Paula E. Jameson

    The plant hormone group, the cytokinins, is implicated in both qualitative and quantitative components of yield. Cytokinins have opposing actions in shoot and root growth—actions shown to involve cytokinin dehydrogenase (CKX), the enzyme that inactivates cytokinin. We revise and provide unambiguous names for the CKX gene family members in wheat, based on the most recently released wheat genome database, IWGSC RefSeq v1.0 & v2.0. We review expression data of CKX gene family members in wheat, revealing tissue‐specific gene family member expression as well as sub‐genome‐specific expression. Manipulation of CKX in cereals shows clear impacts on yield, root growth and orientation, and Zn nutrition, but this also emphasizes the necessity to unlink promotive effects on grain yield from negative effects of cytokinin on root growth and uptake of mineral nutrients, particularly Zn and Fe. Wheat is the most widely grown cereal crop globally, yet is under‐research compared with rice and maize. We highlight gaps in our knowledge of the involvement of CKX for wheat. We also highlight the necessity for accurate analysis of endogenous cytokinins, acknowledging why this is challenging, and provide examples where inadequate analyses of endogenous cytokinins have led to unjustified conclusions. We acknowledge that the allohexaploid nature of bread wheat poses challenges in terms of uncovering useful mutations. However, we predict TILLING followed by whole‐exome sequencing will uncover informative mutations and we indicate the potential for stacking mutations within the three genomes to modify yield components. We model a wheat ideotype based on CKX manipulation.

    更新日期:2019-12-23
  • Scaffold protein GhMORG1 enhances the resistance of cotton to Fusarium oxysporum by facilitating the MKK6‐MPK4 cascade
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-20
    Chen Wang; Hongbin Guo; Xiaowen He; Shuxin Zhang; Jiayu Wang; Lijun Wang; Dezheng Guo; Xingqi Guo

    In eukaryotes, MAPK scaffold proteins are crucial for regulating the function of MAPK cascades. However, only a few MAPK scaffold proteins have been reported in plants, and the molecular mechanism through which scaffold proteins regulate the function of the MAPK cascade remains poorly understood. Here, we identified GhMORG1, a GhMKK6‐GhMPK4 cascade scaffold protein that positively regulates the resistance of cotton to Fusarium oxysporum. GhMORG1 interacted with GhMKK6 and GhMPK4, and the overexpression of GhMORG1 in cotton protoplasts dramatically increased the activity of the GhMKK6‐GhMPK4 cascade. Quantitative phosphoproteomics was used to clarify the mechanism of GhMORG1 in regulating disease resistance, and thirty‐two proteins were considered as the putative substrates of the GhMORG1‐dependent GhMKK6‐GhMPK4 cascade. These putative substrates were involved in multiple disease resistance processes, such as cellular amino acid metabolic processes, calcium ion binding and RNA binding. The kinase assays verified that most of the putative substrates were phosphorylated by the GhMKK6‐GhMPK4 cascade. For functional analysis, nine putative substrates were silenced in cotton, respectively. The resistance of cotton to F. oxysporum was decreased in the substrate‐silenced cottons. These results suggest that GhMORG1 regulates several different disease resistance processes by facilitating the phosphorylation of GhMKK6‐GhMPK4 cascade substrates. Taken together, these findings reveal a new plant MAPK scaffold protein and provide insights into the mechanism of plant resistance to pathogens.

    更新日期:2019-12-21
  • Disruption of miRNA sequences by TALENs and CRISPR/Cas9 induces varied lengths of miRNA production
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-20
    Honghao Bi; Qili Fei; Riqing Li; Bo Liu; Rui Xia; Si Nian Char; Blake C. Meyers; Bing Yang

    MicroRNAs (miRNAs) are 20‐24 nucleotides (nt) small RNAs functioning in eukaryotes. The length and sequence of miRNAs are not only related to the biogenesis of miRNAs but are also important for downstream physiological processes like ta‐siRNA production. To investigate these roles, it is informative to create small mutations within mature miRNA sequences. We used both TALENs (transcription activator‐like effector nucleases) and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR‐associated protein 9 (Cas9) to introduce heritable base pair mutations in mature miRNA sequences. For rice, TALEN constructs were built targeting five different mature miRNA sequences and yielding heritable mutations. Among the resulting mutants, mir390 mutant showed a severe defect in the shoot apical meristem (SAM), a shootless phenotype, which could be rescued by the wild‐type MIR390. Small RNA sequencing showed the two base pair deletion in mir390 substantially interfered with miR390 biogenesis. In Arabidopsis, CRISPR/Cas9‐mediated editing of the miR160* strand confirmed that the asymmetric structure of miRNA is not a necessary determinant for secondary siRNA production. CRISPR/Cas9 with double‐guide RNAs successfully generated mir160a null mutants with fragment deletions, at a higher efficiency than a single‐guide RNA. The difference between the phenotypic severity of miR160a mutants in Col‐0 versus Ler backgrounds highlights a diverged role for miR160a in different ecotypes. Overall, we demonstrated that TALENs and CRISPR/Cas9 are both effective in modifying miRNA precursor structure, disrupting miRNA processing and generating miRNA null mutant plants.

    更新日期:2019-12-20
  • Increased Rubisco content in maize mitigates chilling stress and speeds recovery
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-20
    Coralie E. Salesse‐Smith; Robert E. Sharwood; Florian A. Busch; David B. Stern

    Many C4 plants, including maize, perform poorly under chilling conditions. This phenomenon has been linked in part to decreased Rubisco abundance at lower temperatures. An exception to this is chilling‐tolerant Miscanthus, which is able to maintain Rubisco protein content under such conditions. The goal of this study was to investigate whether increasing Rubisco content in maize could improve performance during or following chilling stress. Here, we demonstrate that transgenic lines overexpressing Rubisco large and small subunits and the Rubisco assembly factor RAF1 (RAF1‐LSSS), which have increased Rubisco content and growth under control conditions, maintain increased Rubisco content and growth during chilling stress. RAF1‐LSSS plants exhibited 12% higher CO2 assimilation relative to nontransgenic controls under control growth conditions, and a 17% differential after 2 weeks of chilling stress, although assimilation rates of all genotypes were ~50% lower in chilling conditions. Chlorophyll fluorescence measurements showed RAF1‐LSSS and WT plants had similar rates of photochemical quenching during chilling, suggesting Rubisco may not be the primary limiting factor that leads to poor performance in maize under chilling conditions. In contrast, RAF1‐LSSS had improved photochemical quenching before and after chilling stress, suggesting that increased Rubisco may help plants recover faster from chilling conditions. Relatively increased leaf area, dry weight and plant height observed before chilling in RAF1‐LSSS were also maintained during chilling. Together, these results demonstrate that an increase in Rubisco content allows maize plants to better cope with chilling stress and also improves their subsequent recovery, yet additional modifications are required to engineer chilling tolerance in maize.

    更新日期:2019-12-20
  • Exome association analysis sheds light onto leaf rust (Puccinia triticina) resistance genes currently used in wheat breeding (Triticum aestivum L.)
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-20
    Fang Liu; Yusheng Zhao; Sebastian Beier; Yong Jiang; Patrick Thorwarth; C. Friedrich H. Longin; Martin Ganal; Axel Himmelbach; Jochen C. Reif; Albert W. Schulthess

    Resistance breeding is crucial for a sustainable control of leaf rust (Puccinia triticina) in wheat (Triticum aestivum L.) while directly targeting functional variants is the Holy Grail for efficient marker‐assisted selection and map‐based cloning. We assessed the limits and prospects of exome association analysis for severity of leaf rust in a large hybrid wheat population of 1574 single‐crosses plus their 133 parents. After imputation and quality control, exome sequencing revealed 202 875 single‐nucleotide polymorphisms (SNPs) covering 19.7% of the high‐confidence annotated gene space. We performed intensive data mining and found significant associations for 2171 SNPs corresponding to 50 different loci. Some of these associations mapped in the proximity of the already known resistance genes Lr21, Lr34‐B, Lr1 and Lr10, while other associated genomic regions, such as those on chromosomes 1A and 3D, harboured several annotated genes putatively involved in resistance. Validation with an independent population helped to narrow down the list of putative resistance genes that should be targeted by fine‐mapping. We expect that the proposed strategy of intensive data mining coupled with validation will significantly influence research in plant genetics and breeding.

    更新日期:2019-12-20
  • Robust CRISPR/Cas9 mediated genome editing and its application in manipulating plant height in the first generation of hexaploid Ma bamboo (Dendrocalamus latiflorus Munro)
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-19
    Shanwen Ye; Gang Chen; Markus V. Kohnen; Wenjia Wang; Changyang Cai; WenSha Ding; Chu Wu; Lianfeng Gu; Yushan Zheng; Xiangqing Ma; Chentao Lin; Qiang Zhu

    Bamboo covers over 30 million hectares of land across the world, and around 2.5 billion people are directly producing and consuming bamboo. However, probably it is the only major agronomic crop that is nearly impossible to make improvement by traditional breeding as the flowering time of most bamboos exceeds 70 years. Bamboo is the fastest growing plant on this planet, whearas, our knowledge on the underlying mechanisms is quite limited mainly due to the lack of efficient genetic manipulation tools. Therefore, the application of genetic engineering technology on bamboo is extremely important for its breeding and basic research. CRISPR (Clustered Regularly Interspaced Short Palindromic repeat)/Cas9 system provides straight forward ways for genome editing in many plants, but was never used in bamboo. In this study, we optimized the CRISPR/Cas9 elements in bamboo protoplast, and for the first time we established an efficient protocol for artificially producing the hexaploid bamboo mutant in its first generation by simultaneously targeting all homo‐alleles or specifically targeting one allele of a putative phytoene synthase gene through CRISPR/Cas9 technology. Furthermore, we produced a bamboo mutant with increased plant height by knocking out one Gibberelline‐responsive gene that we identified previously, which provides a good material for the future study on the molecular mechanism that controls the fast growth of bamboo.

    更新日期:2019-12-20
  • MeCIPK23 interacts with Whirly transcription factors to activate abscisic acid biosynthesis and regulate drought resistance in cassava
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-19
    Yu Yan; Wen Liu; Yunxie Wei; Haitao Shi

    As one of the most important food and energy crops in the world, cassava (Manihot esculenta) is high tolerant to drought and poor nutritional environment. However, the key regulators of drought response in cassava remain elusive. In this study, we found that MeWHYs could physically interact with MeCIPK23 in vivo and in vitro, as revealed by yeast two‐hybrid, biomolecular fluorescence complementation (BiFC), luciferase (LUC) complementation, and pull‐down assays. Moreover, we highlighed their roles in regulating drought response in cassava. Under drought stress conditions, the expression of MeCIPK23 and MeWHYs are up‐regulated. In addition, MeCIPK23 interacts with MeWHYs, which directly bind to the PB element in the promoter of MeNCED1 and activate its transcription. Then the up‐regulated expression of MeNCED1 results in elevated ABA biosynthesis and enhanced drought stress response. Therefore, this study provides new insight into the drought‐resistance mechanism in cassava and potential strategies for further crop breeding and germplasm enhancement.

    更新日期:2019-12-20
  • Learning from methylomes: epigenomic correlates of Populus balsamifera traits based on deep learning models of natural DNA methylation
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-18
    Marc J. Champigny; Faride Unda; Oleksandr Skyba; Raju Y. Soolanayakanahally; Shawn D. Mansfield; Malcolm M. Campbell

    Epigenomes have remarkable potential for the estimation of plant traits. This study tested the hypothesis that natural variation in DNA methylation can be used to estimate industrially important traits in a genetically diverse population of Populus balsamifera L. (balsam poplar) trees grown at two common garden sites. Statistical learning experiments enabled by deep learning models revealed that plant traits in novel genotypes can be modelled transparently using small numbers of methylated DNA predictors. Using this approach, tissue type, a nonheritable attribute, from which DNA methylomes were derived was assigned, and provenance, a purely heritable trait and an element of population structure, was determined. Significant proportions of phenotypic variance in quantitative wood traits, including total biomass (57.5%), wood density (40.9%), soluble lignin (25.3%) and cell wall carbohydrate (mannose: 44.8%) contents, were also explained from natural variation in DNA methylation. Modelling plant traits using DNA methylation can capture tissue‐specific epigenetic mechanisms underlying plant phenotypes in natural environments. DNA methylation‐based models offer new insight into natural epigenetic influence on plants and can be used as a strategy to validate the identity, provenance or quality of agroforestry products.

    更新日期:2019-12-19
  • Genome‐wide association study identifies an NLR gene that confers partial resistance to Magnaporthe oryzae in rice
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-15
    Ming‐Hao Liu; Houxiang Kang; Yucheng Xu; Ye Peng; Dan Wang; Lijun Gao; Xuli Wang; Yuese Ning; Jun Wu; Wende Liu; Chengyun Li; Bin Liu; Guo‐Liang Wang

    Because of the frequent breakdown of major resistance (R) genes, identification of new partial R genes against rice blast disease is an important goal of rice breeding. In this study, we used a core collection of the Rice Diversity Panel II (C‐RDP‐II), which contains 584 rice accessions and are genotyped with 700 000 single‐nucleotide polymorphism (SNP) markers. The C‐RDP‐II accessions were inoculated with three blast strains collected from different rice‐growing regions in China. Genome‐wide association study identified 27 loci associated with rice blast resistance (LABRs). Among them, 22 LABRs were not associated with any known blast R genes or QTLs. Interestingly, a nucleotide‐binding site leucine‐rich repeat (NLR) gene cluster exists in the LABR12 region on chromosome 4. One of the NLR genes is highly conserved in multiple partially resistant rice cultivars, and its expression is significantly up‐regulated at the early stages of rice blast infection. Knockout of this gene via CRISPR‐Cas9 in transgenic plants partially reduced blast resistance to four blast strains. The identification of this new non‐strain specific partial R gene, tentatively named rice blast Partial Resistance gene 1 (PiPR1), provides genetic material that will be useful for understanding the partial resistance mechanism and for breeding durably resistant cultivars against blast disease of rice.

    更新日期:2019-12-17
  • The control of red colour by a family of MYB transcription factors in octoploid strawberry (Fragaria × ananassa) fruits
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-12
    Hua Wang; Hui Zhang; Yuan Yang; Maofu Li; Yuntao Zhang; Jiashen Liu; Jing Dong; Jie Li; Eugenio Butelli; Zhen Xue; Aimin Wang; Guixia Wang; Cathie Martin; Wanmei Jin

    Octoploid strawberry (Fragaria × ananassa Duch.) is a model plant for research and one of the most important non‐climacteric fruit crops throughout the world. The associations between regulatory networks and metabolite composition were explored for one of the most critical agricultural properties in octoploid strawberry, fruit colour. Differences in the levels of flavonoids are due to the differences in the expression of structural and regulatory genes involved in flavonoid biosynthesis. The molecular mechanisms underlying differences in fruit colour were compared between red and white octoploid strawberry varieties. FaMYB genes had combinatorial effects in determining the red colour of fruit through the regulation of flavonoid biosynthesis in response to the increase in endogenous ABA at the final stage of fruit development. Analysis of alleles of FaMYB10 and FaMYB1 in red and white strawberry varieties led to the discovery of a white‐specific variant allele of FaMYB10, FaMYB10‐2. Its coding sequence possessed an ACTTATAC insertion in the genomic region encoding the C‐terminus of the protein. This insertion introduced a predicted premature termination codon, which suggested the loss of intact FaMYB10 protein playing a critical role in the loss of red colour in white octoploid strawberry.

    更新日期:2019-12-13
  • Multiplex CRISPR/Cas9‐mediated metabolic engineering increases soya bean isoflavone content and resistance to soya bean mosaic virus
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-09
    Peipei Zhang, Hongyang Du, Jiao Wang, Yixiang Pu, Changyun Yang, Rujuan Yan, Hui Yang, Hao Cheng, Deyue Yu

    Isoflavonoids, which include a variety of secondary metabolites, are derived from the phenylpropanoid pathway and are distributed predominantly in leguminous plants. These compounds play a critical role in plant–environment interactions and are beneficial to human health. Isoflavone synthase (IFS) is a key enzyme in isoflavonoid synthesis and shares a common substrate with flavanone‐3‐hydroxylase (F3H) and flavone synthase II (FNS II). In this study, CRISPR/Cas9‐mediated multiplex gene‐editing technology was employed to simultaneously target GmF3H1, GmF3H2 and GmFNSII‐1 in soya bean hairy roots and plants. Various mutation types and frequencies were observed in hairy roots. Higher mutation efficiencies were found in the T0 transgenic plants, with a triple gene mutation efficiency of 44.44%, and these results of targeted mutagenesis were stably inherited in the progeny. Metabolomic analysis of T0 triple‐mutants leaves revealed significant improvement in isoflavone content. Compared with the wild type, the T3 generation homozygous triple mutants had approximately twice the leaf isoflavone content, and the soya bean mosaic virus (SMV) coat protein content was significantly reduced by one‐third after infection with strain SC7, suggesting that increased isoflavone content enhanced the leaf resistance to SMV. The isoflavone content in the seeds of T2 triple mutants was also significantly increased. This study provides not only materials for the improvement of soya bean isoflavone content and resistance to SMV but also a simple system to generate multiplex mutations in soya bean, which may be beneficial for further breeding and metabolic engineering.

    更新日期:2019-12-11
  • Understanding sheath blight resistance in rice: the road behind and the road ahead
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-07
    Kutubuddin A. Molla, Subhasis Karmakar, Johiruddin Molla, Prasad Bajaj, Rajeev K. Varshney, Swapan K. Datta, Karabi Datta

    Rice Sheath blight disease, caused by the basidiomycetous necrotrophy Rhizoctonia solani, became one of the major threats to the rice cultivation worldwide, especially after the adoption of high yielding varieties. The pathogen is challenging to manage because of its extensively broad host range and high genetic variability and also due to the inability to find any satisfactory level of natural resistance among the available rice germplasm. It is high time to find remedies to combat the pathogen for reducing rice yield losses and subsequently to minimize the threat to global food security. The development of genetic resistance is the only alternative means to avoid the use of hazardous chemical fungicides. This review mainly focuses on the effort of better understanding the host‐pathogen relationship, finding the gene loci/markers imparting resistance response, and modifying the host genome through transgenic development. The latest development and trend in the R. solani‐ rice patho‐system research with gap analysis is provided.

    更新日期:2019-12-07
  • A membrane‐associated NAC transcription factor OsNTL3 is involved in thermotolerance in rice
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-06
    Xue‐Huan Liu, Yu‐Shu Lyu, Weiping Yang, Zheng‐Ting Yang, Sun‐Jie Lu, Jian‐Xiang Liu

    Heat stress induces misfolded protein accumulation in endoplasmic reticulum (ER), which initiates the unfolded protein response (UPR) in plants. Previous work has demonstrated the important role of a rice ER membrane‐associated transcription factor OsbZIP74 (also known as OsbZIP50) in UPR. However, how OsbZIP74 and other membrane‐associated transcription factors are involved in heat stress tolerance in rice is not reported. In the current study, we discovered that OsNTL3 is required for heat stress tolerance in rice. OsNTL3 is constitutively expressed and up‐regulated by heat and ER stresses. OsNTL3 encodes a NAC transcription factor with a predicted C‐terminal transmembrane domain. GFP‐OsNTL3 relocates from plasma membrane to nucleus in response to heat stress and ER stress inducers. Loss‐of‐function mutation of OsNTL3 confers heat sensitivity while inducible expression of the truncated form of OsNTL3 without the transmembrane domain increases heat tolerance in rice seedlings. RNA‐Seq analysis revealed that OsNTL3 regulates the expression of genes involved in ER protein folding and other processes. Interestingly, OsNTL3 directly binds to OsbZIP74 promoter and regulates its expression in response to heat stress. In turn, up‐regulation of OsNTL3 by heat stress is dependent on OsbZIP74. Thus, our work reveals the important role of OsNTL3 in thermotolerance, and a regulatory circuit mediated by OsbZIP74 and OsNTL3 in communications among ER, plasma membrane and nucleus under heat stress conditions.

    更新日期:2019-12-06
  • Vascular‐Specific Expression of Gastrodia Antifungal Protein Gene Significantly Enhanced Cotton Verticillium Wilt Resistance
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-05
    Yiqin Wang, Chengzhen Liang, Shenjie Wu, Guiliang Jian, Xueyan Zhang, Huanyang Zhang, Jiuyou Tang, Jing Li, Gaili Jiao, Fuguang Li, Chengcai Chu

    Verticillium wilt caused by Verticillium dahliae Kleb is the most destructive disease in cotton‐growing areas around the world, and about 50% of the cotton planting area in China suffers from Verticillium wilt infection, which causes huge economic losses annually. Breeding of broad‐spectrum resistant cotton cultivars through genetic engineering is considered to be one of the most effective approaches to control this fungal disease. We previously reported that a member of Gastrodia antifungal protein (GAFP) family, GAFP4, showed strong antifungal activity to cotton Verticillium wilt. To further enhance the resistance level of transgenic cotton, we supposes that vascular‐specific promoter should be more effective than CaMV 35S promoter. Then we made detailed analysis of the gdcsP promoter from C3‐C4 plant Flaveria anomala and found it contains not only the cis‐acting regulatory elements required for vascular‐ and root‐specific expression, but also disease and wounding inducible elements. The vascular‐specific expression pattern of gdcsP promoter was further confirmed in the transgenic Arabidopsis and cotton expressing β‐glucuronidase (GUS). Moreover, the transgenic cotton plants harboring GAFP4 under control of gdcsP promoter were generated and evaluated for their disease resistance level in Verticillium wilt nursery together with the non‐transgenic control and CaMV 35S:GAFP4 transgenic plants. We demonstrated that with the similar GAFP4 expression, the vascular‐specific expression of GAFP4 plants shows much enhanced resistance to Verticillium wilt. Current data suggested that the vascular‐specific gdcsP promoter is an effective tool not only for genetic engineering of cotton Verticillium wilt resistance, but also for many other applications in genetic engineering.

    更新日期:2019-12-06
  • TaDA1, a conserved negative regulator of kernel size, has an additive effect with TaGW2 in common wheat (Triticum aestivum L.)
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-04
    Hong Liu, Huifang Li, Chenyang Hao, Ke Wang, Yamei Wang, Lin Qin, Diaoguo An, Tian Li, Xueyong Zhang

    Kernel size is an important trait determining cereal yields. In this study, we cloned and characterized TaDA1, a conserved negative regulator of kernel size in wheat (Triticum aestivum). The overexpression of TaDA1 decreased the size and weight of wheat kernels, while its down‐regulation using RNA interference (RNAi) had the opposite effect. Three TaDA1‐A haplotypes were identified in Chinese wheat core collections, and a haplotype association analysis showed that TaDA1‐A‐HapI was significantly correlated with the production of larger kernels and higher kernel weights in modern Chinese cultivars. The haplotype effect resulted from a difference in TaDA1‐A expression levels between genotypes, with TaDA1‐A‐HapI resulting in lower TaDA1‐A expression levels. This favourable haplotype was found having been positively selected during wheat breeding over the last century. Furthermore, we demonstrated that TaDA1‐A physically interacts with TaGW2‐B. The additive effects of TaDA1‐A and TaGW2‐B on kernel weight were confirmed not only by the phenotypic enhancement arising from the simultaneous down‐regulation of TaDA1 and TaGW2 expression, but also by the combinational haplotype effects estimated from multi‐environment field data from 348 wheat cultivars. A comparative proteome analysis of developing transgenic and wild‐type grains indicated that TaDA1 and TaGW2 are involved in partially overlapping but relatively independent protein regulatory networks. Thus, we have identified an important gene controlling kernel size in wheat and determined its interaction with other genes regulating kernel weight, which could have beneficial applications in wheat breeding.

    更新日期:2019-12-04
  • Genome editing in wheat microspores and haploid embryos mediated by delivery of ZFN proteins and cell‐penetrating peptide complexes
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-03
    Andriy Bilichak, Lakshmi Sastry‐Dent, Shreedharan Sriram, Matthew Simpson, Pon Samuel, Steve Webb, Fengying Jiang, Francois Eudes

    Recent advances in genome engineering technologies based on designed endonucleases (DE) allow specific and predictable alterations in plant genomes to generate value‐added traits in crops of choice. The EXZACT Precision technology, based on zinc finger nucleases (ZFN), has been successfully used in the past for introduction of precise mutations and transgenes to generate novel and desired phenotypes in several crop species. Current methods for delivering ZFNs into plant cells are based on traditional genetic transformation methods that result in stable integration of the nuclease in the genome. Here, we describe for the first time, an alternative ZFN delivery method where plant cells are transfected with ZFN protein that eliminates the need for stable nuclease genomic integration and allows generation of edited, but not transgenic cells or tissues. For this study, we designed ZFNs targeting the wheat IPK1 locus, purified active ZFN protein from bacterial cultures, complexed with cell‐penetrating peptides (CPP) and directly transfected the complex into either wheat microspores or embryos. NGS analysis of ZFN‐treated material showed targeted edits at the IPK1 locus in independent experiments. This is the first description of plant microspore genome editing by a ZFN when delivered as a protein complexed with CPP.

    更新日期:2019-12-04
  • Deciphering the High Quality Genome Sequence of Coriander that Causes Controversial Feelings
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-04
    Xiaoming Song, Jinpeng Wang, Nan Li, Jigao Yu, Fanbo Meng, Chendan Wei, Chao Liu, Wei Chen, Fulei Nie, Zhikang Zhang, Ke Gong, Xinyu Li, Jingjing Hu, Qihang Yang, Yuxian Li, Chunjin Li, Shuyan Feng, He Guo, Jiaqing Yuan, Qiaoying Pei, Tong Yu, Xi Kang, Wei Zhao, Tianyu Lei, Pengchuan Sun, Li Wang, Weina Ge, Di Guo, Xueqian Duan, Shaoqi Shen, Chunlin Cui, Ying Yu, Yangqin Xie, Jin Zhang, Yue Hou, Jianyu Wang, Jinyu Wang, Xiu‐Qing Li, Andrew H. Paterson, Xiyin Wang

    Coriander (Coriandrum sativum L. 2n = 2x = 22), a plant from the Apiaceae family, also called cilantro or Chinese parsley, is a globally important crop used as vegetable, spice, fragrance, and traditional medicine. Here, we report a high‐quality assembly and analysis of its genome sequence, anchored to 11 chromosomes, with total length of 2,118.68 Mb and N50 scaffold length 160.99 Mb. We found that two whole‐genome duplication events, respectively dated to ~ 45‐52 and ~ 54‐61 million years ago, were shared by the Apiaceae family after their split from lettuce. Unbalanced gene loss and expression observed between duplicated copies produced by these two events. Gene retention, expression, metabolomics and comparative genomic analyses of Terpene synthase (TPS) gene family, involved in terpenoid biosynthesis pathway contributing to coriander’s special flavor, revealed that tandem duplication contributed to coriander TPS gene family expansion, especially compared to their carrot counterparts. Notably, a TPS gene highly expressed in all 4 tissues and 3 development stages studied, is likely a major‐effect gene encoding linalool synthase and myrcene synthase. The present genome sequencing, transcriptome, metabolome and comparative genomic efforts provide valuable insights into the genome evolution and spice trait biology of Apiaceae and others related plants, and facilitated further research into important gene functions and crop improvement.

    更新日期:2019-12-04
  • Engineering bacteriocin‐mediated resistance against the plant pathogen Pseudomonas syringae
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-12-03
    William M. Rooney, Rhys W. Grinter, Annapaula Correia, Julian Parkhill, Daniel C. Walker, Joel J. Milner

    The plant pathogen, Pseudomonas syringae (Ps), together with related Ps species, infects and attacks a wide range of agronomically important crops, including tomato, kiwifruit, pepper, olive and soybean, causing economic losses. Currently, chemicals and introduced resistance genes are used to protect plants against these pathogens but have limited success and may have adverse environmental impacts. Consequently, there is a pressing need to develop alternative strategies to combat bacterial disease in crops. One such strategy involves using narrow‐spectrum protein antibiotics (so‐called bacteriocins), which diverse bacteria use to compete against closely related species. Here, we demonstrate that one bacteriocin, putidacin L1 (PL1), can be expressed in an active form at high levels in Arabidopsis and in Nicotiana benthamiana in planta to provide effective resistance against diverse pathovars of Ps. Furthermore, we find that Ps strains that mutate to acquire tolerance to PL1 lose their O‐antigen, exhibit reduced motility and still cannot induce disease symptoms in PL1‐transgenic Arabidopsis. Our results provide proof‐of‐principle that the transgene‐mediated expression of a bacteriocin in planta can provide effective disease resistance to bacterial pathogens. Thus, the expression of bacteriocins in crops might offer an effective strategy for managing bacterial disease, in the same way that the genetic modification of crops to express insecticidal proteins has proven to be an extremely successful strategy for pest management. Crucially, nearly all genera of bacteria, including many plant pathogenic species, produce bacteriocins, providing an extensive source of these antimicrobial agents.

    更新日期:2019-12-03
  • Optimizing plant adenine base editor systems by modifying the transgene selection system
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-11-29
    Juan Li, Ruiying Qin, Yuandi Zhang, Shanbin Xu, Xiaoshuang Liu, Jianbo Yang, Xiuqing Zhang, Pengcheng Wei

    Classical CRISPR‐Cas systems introduce a DNA double‐strand break (DSB) at target genomic loci. In plant, DSBs are typically repaired through the error‐prone nonhomologous end joining (NHEJ) pathway and result in small InDels (Chen et al., 2019). Recently, base editors (BEs), including cytosine BEs (CBEs) and adenine BEs (ABEs), were developed to introduce precise nucleotide substitutions by combining the CRISPR‐Cas system with engineered nucleotide deaminases (Gaudelli et al., 2017; Komor et al., 2016). These BEs enable precise conversions between A∙T and G∙C pairs in the eukaryotic genome without introducing DSBs.

    更新日期:2019-11-30
  • Rice microtubule‐associated protein IQ67‐DOMAIN14 regulates grain shape by modulating microtubule cytoskeleton dynamics
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-11-20
    BaoJun Yang, Jos R. Wendrich, Bert De Rybel, Dolf Weijers, Hong‐Wei Xue

    Cortical microtubule (MT) arrays play a critical role in plant cell shape determination by defining the direction of cell expansion. As plants continuously adapt to ever‐changing environmental conditions, multiple environmental and developmental inputs need to be translated into changes of the MT cytoskeleton. Here, we identify and functionally characterize an auxin‐inducible and MT‐localized protein OsIQ67‐DOMAIN14 (OsIQD14), which is highly expressed in rice seed hull cells. We show that while deficiency of OsIQD14 results in short and wide seeds and increases overall yield, overexpression leads to narrow and long seeds, caused by changed MT alignment. We further show that OsIQD14‐mediated MT reordering is regulated by specifically affecting MT dynamics, and ectopic expression of OsIQD14 in Arabidopsis could change the cell shape both in pavement cells and in hypocotyl cells. Additionally, OsIQD14 activity is tightly controlled by calmodulin proteins, providing an alternative way to modify the OsIQD14 activity. Our results indicate that OsIQD14 acts as a key factor in regulating MT rearrangements in rice hull cells and hence the grain shape, and allows effective local cell shape manipulation to improve the rice yield trait.

    更新日期:2019-11-21
  • The dual benefit of a dominant mutation in Arabidopsis IRON DEFICIENCY TOLERANT1 for iron biofortification and heavy metal phytoremediation
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-11-20
    Reena Sharma, Kuo‐Chen Yeh

    One of the goals of biofortification is to generate iron‐enriched crops to combat growth and developmental defects especially iron (Fe) deficiency anaemia. Fe‐fortification of food is challenging because soluble Fe is unstable and insoluble Fe is nonbioavailable. Genetic engineering is an alternative approach for Fe‐biofortification, but so far strategies to increase Fe content have only encompassed a few genes with limited success. In this study, we demonstrate that the ethyl methanesulfonate (EMS) mutant, iron deficiency tolerant1 (idt1), can accumulate 4–7 times higher amounts of Fe than the wild type in roots, shoots and seeds, and exhibits the metal tolerance and iron accumulation (Metina) phenotype in Arabidopsis. Fe‐regulated protein stability and nuclear localisation of the upstream transcriptional regulator bHLH34 were uncovered. The C to T transition mutation resulting in substitution of alanine to valine at amino acid position 320 of bHLH34 (designated as IDT1A320V) in a conserved motif among mono‐ and dicots was found to be responsible for a dominant phenotype that possesses constitutive activation of the Fe regulatory pathway. Overexpression of IDT1A320V in Arabidopsis and tobacco led to the Metina phenotype; a phenotype that has escalated specificity towards optimising Fe homeostasis and may be useful in Fe‐biofortification. Knowledge of the high tolerance and accumulation of heavy metals of this mutant can aid the development of tools for phytoremediation of contaminants.

    更新日期:2019-11-21
  • Integrated QTL mapping, gene expression and nucleotide variation analyses to investigate complex quantitative traits: a case study with the soybean‐Phytophthora sojae interaction
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-11-20
    Maxime de Ronne, Caroline Labbé, Amandine Lebreton, Humira Sonah, Rupesh Deshmukh, Martine Jean, François Belzile, Louise O’Donoughue, Richard Bélanger

    Soybean (Glycine max (L.) Merr.) is the most important legume in the world(Dorff, 2007; Shoemaker et al., 1996). However, the rapid expansion of its cultivated areas, has created new ecological niches for many pathogens. Among them, Phytophthora sojae (Kaufmann and Gerdemann) ‐ ranks as one of the most damaging soybean pests in the world. The most common method to control it is the introgression of resistance genes termed Rps (Resistance to P. sojae) into elite cultivars. This imposes a high selection pressure on P. sojae leading to the development of new virulent pathotypes.

    更新日期:2019-11-21
  • Arabidopsis GDSL1 overexpression enhances rapeseed Sclerotinia sclerotiorum resistance and the functional identification of its homolog in Brassica napus
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-11-20
    Li‐Na Ding, Ming Li, Xiao‐Juan Guo, Min‐Qiang Tang, Jun Cao, Zheng Wang, Rui Liu, Ke‐Ming Zhu, Liang Guo, Sheng‐Yi Liu, Xiao‐Li Tan

    Sclerotinia stem rot (SSR) caused by Sclerotinia sclerotiorum is a devastating disease of rapeseed (Brassica napus L.). To date, the genetic mechanisms of rapeseed’ interactions with S. sclerotiorum are not fully understood, and molecular‐based breeding is still the most effective control strategy for this disease. Here, Arabidopsis thaliana GDSL1 was characterized as an extracellular GDSL lipase gene functioning in Sclerotinia resistance. Loss of AtGDSL1 function resulted in enhanced susceptibility to S. sclerotiorum. Conversely, overexpression of AtGDSL1 in B. napus enhanced resistance, which was associated with increased reactive oxygen species (ROS) and salicylic acid (SA) levels, and reduced jasmonic acid levels. In addition, AtGDSL1 can cause an increase in lipid precursor phosphatidic acid levels, which may lead to the activation of downstream ROS/SA defence‐related pathways. However, the rapeseed BnGDSL1 with highest sequence similarity to AtGDSL1 had no effect on SSR resistance. A candidate gene association study revealed that only one AtGDSL1 homolog from rapeseed, BnaC07g35650D (BnGLIP1), significantly contributed to resistance traits in a natural B. napus population, and the resistance function was also confirmed by a transient expression assay in tobacco leaves. Moreover, genomic analyses revealed that BnGLIP1 locus was embedded in a selected region associated with SSR resistance during the breeding process, and its elite allele type belonged to a minor allele in the population. Thus, BnGLIP1 is the functional equivalent of AtGDSL1 and has a broad application in rapeseed S. sclerotiorum‐resistance breeding.

    更新日期:2019-11-20
  • PpGST1, an anthocyanin‐related glutathione S‐transferase gene, is essential for fruit coloration in peach
    Plant Biotech. J. (IF 6.840) Pub Date : 2019-11-19
    Yun Zhao, Weiqi Dong, Yongchao Zhu, Andrew C. Allan, Kui Lin‐Wang, Changjie Xu

    Anthocyanins have crucial biological functions and affect quality of horticultural produce. Anthocyanins accumulate in ripe peach fruit; differential accumulation is observed in deep coloured cultivar ‘Hujingmilu’ and lightly pigmented cultivar ‘Yulu’. The difference was not fully explained by accumulation of total flavonoids and expression of anthocyanin biosynthetic genes. Expression analysis was conducted on a glutathione S‐transferase gene (PpGST1), and it was found that the expression correlated well with anthocyanin accumulation in peach fruit tissues. Functional complementation of the Arabidopsis tt19 mutant indicated that PpGST1 was responsible for transport of anthocyanins but not proanthocyanidins. PpGST1 was localized in nuclei and the tonoplast, including the sites at which anthocyanin vacuolar sequestration occurred. Transient overexpression of PpGST1 together with PpMYB10.1 in tobacco leaves and peach fruit significantly increased anthocyanin accumulation as compared with PpMYB10.1 alone. Furthermore, virus‐induced gene silencing of PpGST1 in a blood‐fleshed peach not only resulted in a reduction in anthocyanin accumulation but also a decline in expression of anthocyanin biosynthetic and regulatory genes. Cis‐element analysis of the PpGST1 promoter revealed the presence of four MYB binding sites (MBSs). Dual‐luciferase assays indicated that PpMYB10.1 bound to the promoter and activated the transcription of PpGST1 by recognizing MBS1, the one closest to the ATG start codon, with this trans‐activation being stronger against the promoter of deep coloured ‘Hujingmilu’ compared with lightly coloured cultivar ‘Yulu’. Altogether, our data provided molecular evidence supporting coordinative regulatory roles of PpGST1 and PpMYB10.1 in anthocyanin accumulation in peach.

    更新日期:2019-11-20
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