非生物胁迫诱导的GmMYB基因克隆与表达分析

摘要

Response to external environment is the outcome of stress-induced gene expression. In this paper, based on one stress-induced EST sequence, we cloned four R2R3-MYB genes from soybean cultivar Dongnong 42, whose genomic sequences consisted of three exons and two introns. Three of them corresponding to Gm02gl300, Gm03g38040, and Gml0g01340 are respectively consistent with the sequences of Williams 82. A mutation at the 375th single nucleotide in the sequence of Gml9g40650 from Dongnong 42 caused a synonymous amino acid substitution (E125-D125). To test the relationship of four MYB genes with stress resistance, we treated the seedlings of cultivar Dongnong 42 with abiotic stresses including salt, alkali, drought and low temperature in the artificial climate chamber. Quantitative PCR analysis indicated that all of the four genes were transient down-regulated or up-regulated when subjected to the stresses, but different in the expression time, level and tendency. Gm02g01300 was induced by drought stress while Gm03g38040 was strongly induced by multiple stresses, indicating that they play important roles in responding to external stresses. There were also differences in the expression of individual gene between cotyledons and embryos. These results under a variety of abiotic stress conditions suggest that the four R2R3-MYB genes are different not only in the expression patterns, but also in the regulation modes.%本研究室根据一段抗逆的EST序列,从栽培大豆东农42中克隆到4个开放阅读框均是外显子和内含子间隔构成的R2R3 -MYB基因,其中Gm02g01300、Gm03g38040和Gm10g01340与已公布的Williams 82基因组序列完全一致,Gm19g40650第375位的单核苷酸突变导致多肽链第125位的氨基酸发生置换(GAG375→GAC375,E125→D125).以人工气候箱内模拟非生物胁迫(盐、碱、干旱和低温)处理栽培大豆东农42芽期,选择适宜时间点,采用荧光定量PCR技术,检测R2R3 -MYB基因的表达.结果表明,4个基因的表达水平都存在明显波动,呈诱导后短暂上调或下调两种表达模式,但表达时间、强度和趋势存在明显差异；Gm02g01300受干旱诱导明显,Gm03g38040受多种胁迫条件诱导表达强烈,推测这些基因在大豆非生物胁迫的调控中起到重要作用；另外,在子叶与胚间,单个基因的表达也存在差异；多种非生物胁迫条件下,基因的表达不仅存在时空差异,可能也具有调控模式的差异.