大豆DNA脱甲基化酶相关基因的克隆及连作胁迫应答

Cloning of soybean DNA demethylase related genes and response to continuous cropping stress

利用大豆基因组数据库Phytozome v12.1.6获得大豆DNA脱甲基化相关基因Glyma03g34860和Glyma10g07601的CDS序列,以大豆叶片RNA为模板,RT-PCR克隆获得Glyma03g34860基因大小为5226 bp,Glyma10g07601基因大小为6045 bp。利用STRING在线软件预测这两个转录本的互作蛋白质完全一致,主要互作蛋白8个,包括一个AP位点裂解酶和2个RNA聚合酶亚基;采用qRT-PCR分析他们对大豆连作综合逆境胁迫的响应。结果表明:连作综合逆境胁迫使Glyma03g34860和Glyma10g07601的表达均不同程度上调,其中,Glyma10g07601基因在大豆品种安达农家、黑大豆、绥农14和黑农40达显著水平(P<0.05),分别增加1.37、1.22、1.98倍和1.67倍;Glyma03g34860基因在大豆品种垦丰16、绥农14达显著水平(P<0.05),分别增加1.62倍和1.65倍,因此推测他们可能通过使基因组DNA脱甲基化而参与大豆连作综合逆境胁迫的响应。

The soybean genome database phytozome v12.1.6 was used to obtain the CDS sequences of soybean DNA demethylase-related genes Glyma 03g34860 and Glyma 10g07601.Using soybean leaf RNA as template,5226 bp Glyma 03g34860 and 6045 bp Glyma10g07601 were successfully cloned by RT-PCR method.The interacting proteins predicted by using STRING online software were completely the same.There were 8 main interaction proteins that include one AP site lyase and two RNA polymerase subunits.The qRT-PCR results showed that the expressions of Glyma03g34860 and Glyma10g07601 were both up-regulated with different degree under continuous cropping stress.Of these,the level of Glyma10g07601 was significantly increased by 1.37,1.22,1.98 times,and 1.67 times in Andanongjia,Heidadou,Suinong 14,and heinong 40 varieties,respectively(P<0.05),while the level of Glyma03g34860 was significantly increased by 1.62 times and 1.65 times in Kenfeng 16 and Suinong 14 varieties,respectively(P<0.05),suggesting that they participated in the response on continuous cropping comprehensive stress through demethylating genomic DNA in soybean.