Cloning And Functional Verification Of Chalcone Isomerase Gene GmCHI4A And GmCHI4B In Soybean

Soybean is an important cash crop in China.It is not only rich in protein and oil,but also rich in vitamins,isoflavones and other trace substances that are good for health.It has high nutritional value and economic value.However,drought,disease,low temperature,land salinization and other adverse environmental factors can seriously affect the normal growth and development of soybean,will affect production.Studies have shown that CHI chalcone isomerase plays an important role in plant response to abiotic stress and enhancing plant stress resistance.This experiment is based on soybean varieties Williams 82 as experimental material,cloned isomerase chalcone genes Gm CHI4 A and Gm CHI4 B,and carries on the functional verification,the main results were as follows:1.The Gm CHI4 A and Gm CHI4 B genes were cloned from soybean variety Willams 82 by RT-PCR method.The total length of ORF of Gm CHI4 A and Gm CHI4 B is 630 bp,encoding 209 amino acid residues,and the predicted relative molecular mass is 23.5KDa and 25.56 KDa.The predicted encoded proteins are located in the nucleus.Amino acid sequence alignment and phylogenetic tree analysis showed that Gm CHI4 A and Gm CHI4 B genes belong to the IV type chalcone isomerase gene.2.SDS-PAGE electrophoresis showed that both Gm CHI4 A and Gm CHI4 B genes were highly expressed in the prokaryotic system.3.The expression of Gm CHI4 A and Gm CHI4 B genes were analyzed using fluorescence quantitative PCR technology.The results showed that Gm CHI4 A and Gm CHI4 B expressed in root,stem,leaf and embryo,and the two genes had the highest expression level in roots.4.Gm CHI4 A and Gm CHI4 B were transformed into soybean cotyledon nodes to induce hairy roots by Agrobacterium rhizogenes.The transgenic soybean hairy roots was subjected to Na Cl salt stress.The results showed that,the salt tolerance of hairy roots of Gm CHI4 A and Gm CHI4 B increased significantly compared with the control,indicating that Gm CHI4 A and Gm CHI4 B could improve the salt tolerance of transgenic soybean roots.5.The expression of Gm TIP1:1,Gm SOD1,Gm SOS1 in the transgenic hair root of Gm CHI4 A and Gm CHI4 B increased significantly and the expression of Gm TIP1:3,Gm CAT1 and Gm POD decreased.6.Compared with the control,the content of isoflavones in the hair like roots of Gm CHI4 A and Gm CHI4 B was increased.