Screening Of Drought-resistant Soybean Varieties And CircRNA Analysis In Response To Drought Stress

Under natural conditions,plants are often affected by abiotic stresses such as drought,chilling injury,high temperature,salt damage and so on.Soybean is an important economic crop in China,and drought stress is one of the important abiotic stresses affecting soybean yield.The lack of water resources and the frequent occurrence of drought have seriously affected the increase of soybeanyield.The cultivation of drought-tolerant soybean varieties plays an important role in increasing the yield of soybean.Drought resistance is a complex quantitative trait,which is affected by a variety of genes and environment.Therefore,it is of great significance to screen soybean drought resistance related circRNA and drought resistance genes to develop soybean drought resistance related circRNA molecular marker assisted breeding technology.In this study,20 soybean varieties were selected for the experiment.The germination rate,physiological index,chlorophyll fluorescence,agronomic characters and other items were counted in the bud stage,seedling stage and flowering stage,by PEG-6000 simulating drought and planting potted soybean in a drought-resistant shed.Using the method of calculating drought resistance coefficient and membership value function to conduct a comprehensive analysis.Finally,the drought resistant varieties were selected.Using six drought-resistant varieties selected as experimental materials,44 circRNA were detected by PCR.The expression differenciation of different circRNA in different drought resistant varieties was analyzed,and the key circRNA,responding to drought stress was selected.The drought-resistant function of a key parent gene corresponding to circRNA was further discussed by using soybean hairy root,which laid a foundation for further study on the function of circRNA and for using circRNA as a molecular marker to identify drought-resistant varieties and to explore drought-resistant genes.Main research results are as follows:(1)PEG simulated drought experiment was carried out with 20 soybean varieties at bud stage,and pot experiment at seedling stage was carried out in 2017 and 2018.According to physiological indexes,yield characters,four varieties with strong drought resistance were selected: Tiefeng 8,Jindou 21,Aika 166 and Suinong 14,and four varieties with moderate drought resistance were Longpin 03-311,L-10,Zaoshu 18,Heinong 44.Three drought sensitive varieties were identified as Cateye,Dongnong 594 and Hefeng 47.(2)Six drought resistant varieties,Tiefeng 8,Jindou 21,Aika 166,Suinong 14,Longpin 03-311 and L-10,and six drought sensitive varieties were used as experimental materials to identify 44 circRNA by PCR,and 26 circRNA,differentially expressed under control and drought conditions were obtained.Among them,6 circRNA were the key drought resistant circRNA.(3)The target gene Glyma.07G2457,corresponding to gma-circ R29 was cloned to construct the recombinant vector pCAMBIA3300-Glyma.07G2457.(4)The function of Glyma.07G2457 gene was predicted to be lysyl tRNA synthetase,and the relative expression of Arabidopsis thaliana homologue AT3G11710 was predicted to increase at first and then decrease under drought stress.The results of fluorescence quantitative PCR on Glyma.07G2457 showed that under drought stress,the expression of Glyma.07G2457 was affected by drought stress and positively regulated,which was similar to the prediction of homologous genes in Arabidopsis thaliana.(5)The overexpressed pCAMBIA3300-Glyma.07G2457 vector and the control empty vector were transformed into Agrobacterium tumefaciens.The results of PEG simulated drought experiment showed that the overexpressed root length increased by twice as much as that of the control.The parent gene Glyma.07G2457 corresponding to gma-circ R29 is involved in the drought resistance response of soybean,and it is a drought resistance gene.Gma-circ R29 is up-regulated in response to plant water stress,and is an important soybean drought resistance circRNA.