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Comparative Transcript Profiling and Multiplex qRT-PCR Analysis Between Salt-Tolerant and Sensitive Wheat Genotypes 耐盐小麦基因型与敏感小麦基因型的转录本比较分析和多重 qRT-PCR 分析
Pub Date : 2023-12-26 DOI: 10.12974/2311-858x.2023.11.7
Aysen Yumurtaci
Identification of candidate genes combined with gene expression profiling carries importance to facilitate the molecular basis of salt stress response in plants. Here, cDNA-AFLP was used to compare the transcribed sequences among two bread and two durum wheat genotypes with different levels of salt tolerance. Transcript derived fragments (TDFs) screened on polyacrylamide gels and 36 salt stress induced unique fragments were detected in salt tolerant bread wheat genotype (Alpu cv.). The fragment size of these 36 TDFs was ranged between 99bp to 252bp. Full sequence information of 14 TDFs were obtained after cloning, then GeXP analyzer-based multiplex qRT-PCR assay was performed on leaf tissue derived from 12 TDFs. Targeted gene expression levels of two TDFs (TDF4-GT066302 and TDF11-GT066301) were showed clear upregulation in salt tolerant bread wheat genotype (Alpu cv.) and they were matched with hypothetical proteins. Especially, gene expression level of GT066301 was increased as 3.28 fold at 27th hours of salt stress for salt tolerant genotype. According to blastx similarity results, out of 14 sequenced fragments, two TDFs were closely matched with “cytochrome P450 monooxygenase” protein while four of them matched with Oryza “hypothetical” and “unknown” proteins. Outputs of this study might ensure comparative data for hypothetical protein gene expression and new useful alleles in response to salt stress in wheat.
候选基因的鉴定与基因表达谱分析相结合,对促进植物盐胁迫响应的分子基础具有重要意义。在此,我们利用 cDNA-AFLP 比较了耐盐性不同的两种面包小麦和两种硬质小麦基因型的转录序列。在聚丙烯酰胺凝胶上筛选转录本衍生片段(TDFs),在耐盐面包小麦基因型(Alpu cv.)中检测到 36 个盐胁迫诱导的独特片段。这 36 个 TDFs 的片段大小在 99bp 到 252bp 之间。克隆后获得了 14 个 TDFs 的全序列信息,然后对 12 个 TDFs 的叶组织进行了基于 GeXP 分析仪的多重 qRT-PCR 检测。在耐盐面包小麦基因型(Alpu cv.)中,两个 TDFs(TDF4-GT066302 和 TDF11-GT066301)的靶向基因表达水平明显上调,并与假定蛋白相匹配。尤其是耐盐基因型在盐胁迫 27 小时后,GT066301 的基因表达水平增加了 3.28 倍。根据blastx相似性结果,在14个测序片段中,2个TDF与 "细胞色素P450单氧化酶 "蛋白密切匹配,4个与Oryza "假定 "和 "未知 "蛋白匹配。这项研究的成果可确保获得假说蛋白基因表达的比较数据,并在小麦应对盐胁迫时获得新的有用等位基因。
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Global Journal Of Botanical Science
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