猪殃殃对AHAS抑制剂靶标抗性的快速分子检测

Rapid molecular detection of the resistance of Galium aparine var.tenerum to AHAS inhibitors

为建立猪殃殃靶标抗性快速检测方法,并明确小麦田猪殃殃Galium aparine var.tenerum对AHAS抑制剂靶标的突变类型及分布,从河南、陕西、安徽、江苏和山东5省不同田块采集疑似对AHAS抑制剂产生抗性的猪殃殃植株,采用特异性引物PCR扩增靶标酶AHAS基因保守区片段,并以直接测序法检测采集样品,通过与拟南芥AHAS基因序列比对分析后明确其突变位点。结果显示,在5省25个农田的样品中共有19个农田检测到AHAS突变,分布在河南、安徽和江苏3省;在检测样品中发现突变发生在2个位点,共有3种突变类型,分别是197位脯氨酸(CCC)突变为丙氨酸(GCC)或丝氨酸(TCC),或者是574位色氨酸(TGG)突变为亮氨酸(TTG),检测结果与田间药效反应基本一致。这种用特异性引物扩增目的片段测序的方法,由于其可以在生长当季进行检测,适用于田间靶标突变抗性猪殃殃的快速检测与监测。

In order to establish a rapid method for detecting target resistance to AHAS inhibitors,and confirm the resistant mutant patterns and distributions of Galium aparine var. tenerum in wheat,the suspected AHAS-resistant G. aparine var. tenerum seedlings were selected from different fields in Henan,Shaanxi,Anhui,Jiangsu and Shandong provinces. The primers were designed to amplify the fragments of AHAS gene sequence in G. aparine var. tenerum corresponding to AHAS gene in Arabidopsis thaliana.PCR products were directly sequenced to detect the mutations. Among 25 populations collected from five provinces,19 AHAS-resistant mutant populations were detected from Henan, Anhui and Jiangsu provinces. There were totally two mutant sites and three mutant patterns being detected,which were pro-197-ser,pro-197-ala,and try-574-leu. The results of target-mutant detection were consistent with the bioassay efficacy in the field. The method of target mutant detection by PCR amplification and sequencing could detect the resistance in season,and it was an economical,quick,convenient and accurate way for detecting AHAS-target resistant G. aparine var. tenerum. So it＇s very important and necessary to detect and monitor the AHAS herbicide-resistance in the field.