当前位置: X-MOL 学术J. Berry Res. › 论文详情
Our official English website, www.x-mol.net, welcomes your feedback! (Note: you will need to create a separate account there.)
Transcriptome sequencing and de novo assembly in red raspberry fruit development to elucidates the secondary metabolite pathways
Journal of Berry Research ( IF 1.7 ) Pub Date : 2020-07-31 , DOI: 10.3233/jbr-200552
Xiaojun Kang 1 , Wenxin Li 1 , Xuemei Zhang 2 , Yiwei Tang 3 , Zhilei Zhao 4 , Yuhong Gu 1 , Guohui Qi 2 , Suping Guo 2
Affiliation  

BACKGROUND:Red raspberry (Rubus idaeus L.), known as “golden fruit”, has excellent potential for immune‘regulation, anti-inflammation and anti-cancer due to its health-promoting secondary metabolites. The lack of genetic information in public databases has been a constraint for the genetic improvement of red raspberry. OBJECTIVE:The primary aim of the work was to find the key genes relating with the secondary metabolite pathways. METHODS:De novo assembly transcriptome sequencing of red raspberry (‘Heritage’ variety) fruit in different development stages was performed using an Illumina Hiseq platform. Transcriptome was obtained by the de novo assembly through Trinity assembler. Coding sequences were successfully characterized using databases including non-redundant protein (NR), euKaryotic Ortholog Groups of proteins (KOG), Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). Fragments Per Kilobase of transcript sequence per Million base pairs sequenced (FPKM) method was used to calculate the differentially expressed unigenes. RESULTS:In total, 205,880 unigenes with an average length of 1120 bp and an N50 of 2005 bp were obtained, of which 182,443 unigenes were annotated. Non redundant (NR) annotation showed that a majority of the best hits (58.6%) are wild strawberry (Fragaria vesca). Additionally, the unigenes were also annotated in euKaryotic Ortholog Groups of proteins database and Gene ontology database, and mapped the KEGG pathways. We predicted that 8331 TFs from the unigenes database and these TFs were classified into 94 different common families. The major families were associated with the C2H2 (9.19%), followed by the C3H (4.60%), MYB-related group (4.23%), bZIP (4.13%) and B3 (4.03%). These results were considered to be involved in the regulation of metabolic and secondary metabolic biosynthesis. Totally, 3,369, 3,461 and 441 differentially expressed genes (DEGs) were found in period 2 vs period 1, period 3 vs period 2 and period 4 vs period 3 paired comparisons, respectively. These DEGs were analyzed based on BLASTx, which were mapped to 22 KEGG pathways associating with secondary metabolites during red raspberry fruit ripening, involving anthocyanin biosynthesis, flavonoid biosynthesis, sesquiterpenoid and triterpenoid biosynthesis, etc. To validate the high-throughput sequencing results, six target genes involved in secondary metabolite pathways of red raspberry fruit were tested by qRT-PCR. The results of qRT-PCR assay were generally consistent with the results of RNA sequencing. CONCLUSION:The transcriptome sequencing of the red raspberry fruit at different development stage in this study enriched the genetic information resources of this variety, and will discover the genes relating with secondary metabolic pathways, benefiting to engineer high-quality plants with enhanced active ingredients.

中文翻译:

红树莓果实发育中的转录组测序和从头组装,阐明了次级代谢产物途径

背景:被称为“金果”的红树莓(Rubus idaeus L.)具有促进健康的次生代谢产物,具有极好的免疫调节,抗发炎和抗癌的潜力。公共数据库中缺乏遗传信息一直是红树莓遗传改良的制约因素。目的:这项工作的主要目的是寻找与次生代谢途径相关的关键基因。方法:使用Illumina Hiseq平台在不同发育阶段对红树莓(“ Heritage”品种)果实进行从头组装转录组测序。转录组是通过Trinity汇编程序从头开始获得的。使用非冗余蛋白(NR),真核蛋白直链同源蛋白组(KOG),基因本体论(GO)和《京都基因与基因组百科全书》(KEGG)。每百万碱基对测序的转录物序列的每千碱基片段数(FPKM)方法用于计算差异表达的单基因。结果:获得205,880个单基因,平均长度为1120 bp,N50为2005 bp,其中注释了182,443个单基因。非冗余(NR)注释显示,大多数最佳匹配(58.6%)是野草莓(Fragaria vesca)。此外,在基因数据库的真核直系同源组和基因本体数据库中还注释了单基因,并绘制了KEGG通路。我们从unigenes数据库中预测了8331个TF,并将这些TF分为94个不同的常见家族。主要家庭与C2H2(9.19%)相关,其次与C3H(4.60%)相关,MYB相关组(4.23%),bZIP(4.13%)和B3(4.03%)。这些结果被认为与代谢和次级代谢生物合成的调节有关。在第2期对第1期,第3期对第2期以及第4期对第3期配对比较中,共发现了3,369、3,461和441个差异表达基因(DEG)。基于BLASTx对这些DEG进行了分析,并将其映射到红树莓果实成熟过程中与次级代谢产物相关的22条KEGG途径,涉及花色苷生物合成,类黄酮生物合成,倍半萜和三萜生物合成等。为验证高通量测序结果,共有六个靶标通过qRT-PCR测试了红树莓果实次生代谢途径中涉及的基因。qRT-PCR分析的结果通常与RNA测序的结果一致。结论:本研究在不同发育阶段对红树莓果实进行转录组测序,丰富了该品种的遗传信息资源,并将发现与次生代谢途径有关的基因,有利于提高植物活性成分的品质。
更新日期:2020-08-04
down
wechat
bug