Freezing temperatures are an abiotic stress that has a serious impact on plant growth and development in temperate regions and even threatens plant survival. The wild apple tree (Malus sieversii) needs to undergo a cold acclimation process to enhance its freezing tolerance in winter. Changes that occur at the molecular level in response to low temperatures are poorly understood in wild apple trees. Phytohormone and physiology profiles and transcriptome analysis were used to elaborate on the dynamic response mechanism. We determined that JA, IAA, and ABA accumulated in the cold acclimation stage and decreased during freezing stress in response to freezing stress. To elucidate the molecular mechanisms of freezing stress after cold acclimation, we employed single molecular real-time (SMRT) and RNA-seq technologies to study genome-wide expression profiles in wild apple. Using the PacBio and Illumina platform, we obtained 20.79G subreads. These reads were assembled into 61,908 transcripts, and 24,716 differentially expressed transcripts were obtained. Among them, 4410 transcripts were differentially expressed during the whole process of freezing stress, and these were examined for enrichment via GO and KEGG analyses. Pathway analysis indicated that “plant hormone signal transduction”, “starch and sucrose metabolism”, “peroxisome” and “photosynthesis” might play a vital role in wild apple responses to freezing stress. Furthermore, the transcription factors DREB1/CBF, MYC2, WRKY70, WRKY71, MYB4 and MYB88 were strongly induced during the whole stress period. Our study presents a global survey of the transcriptome profiles of wild apple trees in dynamic response to freezing stress after two days cold acclimation and provides insights into the molecular mechanisms of freezing adaptation of wild apple plants for the first time. The study also provides valuable information for further research on the antifreezing reaction mechanism and genetic improvement of M. sieversii after cold acclimation.

中文翻译：

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冷驯化后冰冻胁迫下海棠的转录组分析

低温是一种非生物胁迫，严重影响温带地区植物的生长发育，甚至威胁植物的生存。野苹果树（苹果）需要进行冷驯化过程，以增强其冬季的抗冻能力。在野生苹果树中，人们对低温在分子水平上发生的变化知之甚少。使用植物激素和生理学概况以及转录组分析来阐述动态响应机制。我们确定 JA、IAA 和 ABA 在冷驯化阶段积累，并在冷冻胁迫期间减少以响应冷冻胁迫。为了阐明冷驯化后冷冻应激的分子机制，我们采用单分子实时 (SMRT) 和 RNA-seq 技术来研究野生苹果的全基因组表达谱。使用 PacBio 和 Illumina 平台，我们获得了 20.79G 的 subreads。这些读数被组装成 61,908 个转录本，并获得了 24,716 个差异表达的转录本。其中，4410个转录本在整个冷冻胁迫过程中差异表达，并通过GO和KEGG分析对这些转录本进行了富集检查。通路分析表明，“植物激素信号转导”、“淀粉和蔗糖代谢”、“过氧化物酶体”和“光合作用”可能在野苹果对冷冻胁迫的反应中起重要作用。此外，转录因子 DREB1/CBF、MYC2、WRKY70、WRKY71、MYB4 和 MYB88 在整个胁迫期间都受到强烈诱导。我们的研究对野苹果树在冷驯化两天后对冷冻胁迫的动态响应的转录组谱进行了全球调查，并首次提供了对野苹果植物冷冻适应的分子机制的见解。该研究也为进一步研究冷驯化后的筛子菌抗冻反应机理和遗传改良提供了有价值的信息。