Cloning And Functional Verification Of Cysteine Protease Gene SaCP1 In Sophora Alopecuroides L.

The earth's water resources are about 1.386 billion cubic kilometers,of which salt water accounts for 97.5%,and available fresh water resources are relatively scarce.With the development of the human economy and population expansion,water shortages have become more and more serious,which has led to the expansion of arid areas and the increase of drought levels,thus aggravating agricultural drought.In China,drought is the most harmful to agriculture,and the degree of damage is increasing year by year.Therefore,improving the drought tolerance of crops has become an urgent problem to be solved in crop breeding.With the advancement of transgenic technology in molecular biology,the method of using transgenic technology to improve plant drought tolerance has been widely recognized.For the selection of drought-tolerant genes in transgenic technology,it is one of the effective methods to isolate and clone genes that significantly increase the drought tolerance of plants in drought-tolerant plants.Sophora alopecurides L.is a perennial shrub of the genus Leguminosae,widely distributed in the arid desert areas of northwestern China.It is drought-tolerant,salt-tolerant and cold-tolerant,and is a gene resource pool rich in stress-resistant genes.In this experiment,a drought-tolerant gene cysteine protease SaCP1 was screened from the yeast yeast expression library of Sophora alopecuroides seedlings,and the preliminary structural analysis and functional verification of this gene were carried out.Below are key research findings :1.Identification of drought tolerance index of bitter bean in seedling stage,the results showed that bitter bean has high drought tolerance.2.The SaCP1 gene was cloned from the yeast yeast expression library of Sophora alopecuroides seedlings.The SaCP1 's ORF is 534 bp in length and encodes 177 amino acid residues with a predicted relative molecular mass of 19.07.5 KDa.The prediction of protein tertiary structure indicates that the gene belongs to the papain family.3.We analysis the SaCP1 by real-time PCR.The results showed that SaCP1 was expressed in the leaves,roots and steams.The highest expression is in roots.4.After salt,alkali and drought treatment of Sophora alopecuroides,the transcriptome sequencing of Sophora alopecuroides seedlings was carried out.The results showed that compared with the control,the transfer of SaCP1 significantly improved the drought tolerance of soybean hairy roots.SaCP1 is involved in the process of apoptosis.5.The high osmotic stress treatment of S.cerevisiae INVSC1 Ura deficiency type transferred to SaCP1 gene showed that SaCP1 gene increased the high osmotic stress tolerance of S.cerevisiae INVSC1 Ura deficiency type.6.Using the legume model transformation system,the SaCP1 gene was transformed into Agrobacterium rhizogenes K599,infecting the soybean cotyledonary node,and the transgenic soybean hairy roots were subjected to sorbitol drought stress and NaCl salt stress.The results showed that compared with the control,the transfer of SaCP1 The drought tolerance and salt tolerance of soybean hairy roots was significantly improved,indicating that SaCP1 can improve the drought and salt tolerance of transgenic soybean roots.