Cloning Of A Novel Arabidopsis Thaliana Gene At234 And Its Functional Analysis In Abiotic And Biotic Stresses

In this study, we screened RNAi mutant library of Arabidopsis with high temperature stress, and got several high-temperature tolerant Arabidopsis mutants.The target silenced gene was amplified from these mutants and sequenced, which length is 206 bp and named as At234. By sequence alignment in Arabidopsis TAIR database website sequence, it indicated that At234 locates in Arabidopsis chromosome2 without introns, the full-length of At234 is 380 bp with 5’-UTR in length of 44 bp,3’-UTR in length of 102 bp and ORF in length of 234 bp. At234 encodes 77 amino acid residues, translation of protein molecular weight is 8.944 k Da, and the isoelectric point is 9.98. In order to further verify the function of this gene, we conducted biological and abiotic stress tests, such as disease tests of RNAi plants and plant by differentpathogens, heavy metal tolerance test, drought and high salt stress and so on.The main research results are as follows:Threshold temperature of high temperature tolerance of wild-type Arabidopsis was first determined. Then wild-type Arabidopsis and mutants of RNAi libraries(250and 350 library) were treated in threshold temperature(65℃) for 30 min, and several high-temperature tolerant Arabidopsis mutants were obtained. The target gene was cloned by PCR and the obtained mutants were all At234 gene mutants. The seeds of these mutants were screened by hygromycin and harvested in single plant. Finally, the homozygotes Arabidopsis RNAi mutants were obtained.The detection system of plant pathogen interaction in bottle, which was set up by our group, was used to analyze the reaction of At234 mutants to different pathogens. It was found that under tissue culture conditions, At234 mutants showed higher susceptibility to Verticillium dahliae and Sclerotinia sclerotiorum and showed higher resistance to B. cinerea and Fusarium solani, compared with wild-type Arabidopsis,and showed similar susceptibility to F. graminearum, compared with wild-type Arabidopsis.Under tissue culture conditions, At234 mutants showed high tolerance to Cd Cl2,Pb(NO3)2 and other heavy metal salts, showed significant tolerace to mannitol-mimicked drought. On medium containing 1.0% Na Cl, at234 root system developed normally, showing strong high salt tolerance, compared with wild-type Arabidopsis.In summary, At234 is a key gene involved in biotic and abiotic stress responses in Arabidopsis. The mechanism is not yet clear and needs further studied.