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ABC转运蛋白基因与植物的多抗耐药性密切相关,是植物抗农药机理研究的热点。克隆黄瓜ABC转运蛋白基因abca19,分析其编码蛋白的氨基酸序列、理化性质和结构,了解其在霜霉威胁迫条件下的表达规律,为abca19的研究奠定基础,为黄瓜抗农药霜霉威机理研究积累材料。以D0351为材料,根据黄瓜基因组数据库中Csa7M368750.1基因编码区全序列,应用Primer Premier 5.0软件设计引物,从黄瓜果实cDNA中克隆abca19基因的开放阅读框。定量PCR分析霜霉威胁迫处理条件下,黄瓜果实不同处理时间和黄瓜不同部位abca19的表达变化。abca19序列长度为921bp,注册到Gene Bank,登录号为KC123181。abca19编码306个氨基酸,与大豆(Glycine max)等植物abca19的同源性为80%。该蛋白是一个疏水蛋白,无跨膜结构,包含6个α-螺旋结构,无信号肽。具有蛋白激酶C磷酸化位点、N-豆蔻酰化位点、N-糖基化位点和酪蛋白激酶Ⅱ磷酸化位点等多个活性位点。霜霉威处理条件下,0.5~6h黄瓜果实处理组abca19相对表达量极显著高于对照,9~48h对照组abca19相对表达量高于处理组,但二者间差异不显著。不同处理时间点(3、9和24h),叶片中abca19相对表达量最高,果实中其次,茎中最低。成功克隆到黄瓜abca19,该基因具有已知物种abca19的特征。霜霉威处理条件下,黄瓜果实abca19参与农药霜霉威胁迫响应,反应迅速且具有时限性。在黄瓜叶片和果实中,abca19积极参与对农药霜霉威胁迫的响应,茎中响应较弱。
ABC transporter gene is closely related to multi-drug resistance in plants and is a hot spot in the study of plant anti-pesticide mechanism. Cloning cucumber ABC transporter gene abca19, analysis of its encoded protein amino acid sequence, physical and chemical properties and structure, to understand its expression in the conditions under the influence of downy mildew conditions laid the foundation for the study of abca19, Accumulate material. Based on the complete sequence of Csa7M368750.1 gene in cucumber genome database, Primer Premier 5.0 software was used to design the primers for D0351. The open reading frame of abca19 gene was cloned from cucumber cDNA. Quantitative PCR analysis was used to analyze the changes of abca19 in cucumber fruit at different treatment time and in different parts of cucumber under the conditions of downy mildew. The length of the abca19 sequence is 921 bp, registered with Gene Bank with accession number KC123181. abca19 encodes a polypeptide of 306 amino acids with a homology of 80% with plant abca19 such as Glycine max. This protein is a hydrophobin and has no transmembrane structure and contains 6 α-helical structures without signal peptide. It has many active sites such as phosphorylation site of protein kinase C, N-myristoylation site, N-glycosylation site and casein kinase Ⅱ phosphorylation site. Under the condition of propamocarb treatment, the relative expression level of abca19 in cucumber fruit treated with 0.5-6 h was significantly higher than that in control, and the relative expression level of abca19 in control group with 9-48 h was higher than that in control group, but the difference was not significant. At different treatment time points (3, 9 and 24h), the highest expression level of abca19 in leaves was observed, followed by the lowest in fruit and stems. The cucumber abca19 was successfully cloned, which has the characteristic of the known species abca19. Under the condition of propamocarb treatment, the cucumber fruit abca19 is involved in the response to the agrochemical downy mildew, and the response is rapid and time-limited. In cucumber leaves and fruits, abca19 actively participates in the response to pesticide downy mildew, with a weak response in stems.