Basis Of Variable Sensitivity To Mesosulfuron-methyl In Different Tausch's Goatgrass (Aegilops Tauschii Coss.) Population

Tausch's goatgrass (Aegilops tauschii Coss.), an annual or perennial grass weed, is classified as Aegilops. In recent years, an estimated 330,000 ha are infested in winter wheat fields in Hebei, Shanxi, Shandong, Henan, Chongqing, Shaanxi, Inner Mongolia and Jiangsu Provinces. Increasing damage and rapid spreading made it a serious weed species in winter wheat fields. It is insensitive to most herbicides with the exception of mesosulfuron-methyl (Bayer CropScience). However, our previous research by whole-plant bioassay did observe that different tausch's goatgrass population had signi?cant variation in the level of sensitivity to mesosulfuron. We have studied the ALS gene sequence and ALS-mRNA expression level differences in different tausch's goatgrass population at the molecular level. Furthermore, at the physiological and biochemical levels, effects of mesosulfuron-methyl on target enzyme acetolactate synthase and metabolic enzyme glutathione-S-transferase activity were also systematically conducted. The research on mechanism of mesosulfuron-methyl sensitivity in different tausch's goatgrass population provide a theoretical basis for controlling tausch's goatgrass effectively and using mesosulfuron-methyl in a scientific, rational way. The main results are as follows:1. The response of different tausch's goatgrass population to mesosulfuron-methyl was determined by whole plant experiments in the greenhouse. The experiments demonstrated that GR50 of mesosulfuron-methyl to Hebei Baoding, Henan Anyang, Shanxi Linfen, Hebei Xingtai , Hebei Handan and Beijing population were 9.94, 5.84, 3.01, 2.79, 1.80 and 0.60 g a i·ha-1, respectively. The most unsusceptible Hebei Baoding population showed 16.57-fold more insensitive than the most susceptible Beijing population.2. Ampli?ed 1,720 base pairs of the acetolactate synthase (ALS) without any intron and derived 572 amino acid sequences from different tausch's goatgrass population were aligned and compared. The nucleotide sequences of Hebei Baoding, Beijing, Shanxi Linfen, Hebei Handan and Henan Anyang tausch's goatgrass populations were registered in the Genbank, with accession numbers FJ997627, FJ997628, FJ997629, FJ997630 and FJ997631, respectively. Seven base polymorphisms were observed and compared with the most susceptible Beijing population, four of which resulted in amino acid changes: Gln236Arg (Hebei Baoding, Hebei Handan, Shanxi Linfen and Henan Anyang populations), Ser285Pro (Hebei Baoding population), Met348Ile (Hebei Baoding population), and Pro611Leu (Hebei Handan population).3. Quantitative analysis was conducted by SYBR Green I real-time fluorescent quantitative PCR to detect the expression levels of ALS gene in Hebei Baoding and Beijing populations which were treated by recommended lower limit dose of mesosulfuron-methyl. The results indicated that the expression of ALS gene from Hebei Baoding population increased obviously at 1 d after treatment (DAT) and reached the peak value at 4 DAT which was 2.93 times higher than that of control, then it returned to its control level. Meanwhile, the expression of ALS gene from Beijing population in the first 2 days increased slightly, even though it also reached the peak value at 4 DAT which was only 1.59 times higher than that of control. However, it had a sustained decline and decreased to 42% of its control at 13 DAT.4. In vitro ALS enzyme assays showed that I50 value of mesosulfuron-methyl to Hebei Handan has the highest value 0.68μM; Henan Anyang was the lowest, its I50 was 0.22μM; I50 value of the most unsusceptible Hebei Baoding population and the most susceptible Beijing population were 0.50μM and 0.26μM respectively.5. In vivo ALS enzyme assays showed that Hebei Baoding and Beijing populations had similar ALS specific activity in the absence of mesosulfuron-methyl. However, in the presence of mesosulfuron-methyl, ALS specific activity of Hebei Baoding population increased 1 DAT and reached the peak value 58.2 n mol·h-1·mg-1 protein 3 DAT, which was 2.1 times greater than that of control. Then ALS specific activity returned to its normal level 5 DAT. Meanwhile, ALS specific activity of the Beijing population were all lower than that of contemporaneous control and decreased to 22% of its relative control 13 DAT.6. GSTs activity analysis for Hebei Baoding, Henan Anyang, Shanxi Linfen, Hebei Handan and Beijing populations demonstrated that the GSTs activity was similar among the five populations and OD340nm values were fluctuating between 0.31~0.33. In vivo, when treated with mesosulfuron-methyl, the GSTs relative activity of Hebei Baoding population had increased distinctly 1 DAT and reached the peak value 4 DAT, and maintained higher level until 9 DAT, then it returned to control level; GSTs relative activity of Beijing population reached its peak value 2 DAT with much lower value 1.13, and then decreased continuously, and fell to 0.13 13 DAT. The trend of changes in GSTs relative activities in other three populations were between Hebei Baoding and Beijing population.In summary, the sensitivity levels to mesosulfuron-methyl in tausch's goatgrass in winter wheat fields in North China was investigated for the first time. Basis of variable sensitivity to mesosulfuron-methyl in tausch's goatgrass was elucidated both at the enzyme level and gene level.