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Cloning and expression pattern research under abiotic stress of the WRKY2 in potato

LI Li-Qin;WANG Xi-Yao;College of Agronomy,Sichuan Agricultural University;  
Function of St WRKY2 has not been reported. Plant WRKY proteins are found as an important class of transcription factors in recent years,which are widely involved in biotic stress,abiotic stress and growth development regulation,and they are mainly divided into three groups. WRKY2 was cloned from potato by homology cloning,with length of the coding region 1 065 bp,encoding 355 amino acids by DNA sequencing. With the amino acid sequence of St WRKY2 blasting in NCBI,19 homology amino acid sequences were selected to analysis conservative domain. Amino acid analysis showed that the St WRKY2 contained 1 conserved WRKY domain( TTGACC) and belonged to the second group of WRKY family members,and its zinc-finger structure was C-X5-C-X23H-X-H. Phylogeny tree results showed St WRKY2 was the most closest to Sl WRKY7( Solanum lycopersicum) with 96% similarity,and the similarity of a WRKY protein of tobacco was 86%. The protein-GST( glutathione-S-transferase) complex was obtained in Escherichia coli by using the method of prokaryotic expression. St WRKY2-GST could combine W-box in vivo by Electrophoretic Mobility Shift Assay analysis,but only GST protein could not combine W-box. St WRKY2-GST combination with W-box could be competed by cold probe( unlabeled probe). And the experiment results also showed that St WRKY2-GST could not combine mutation W-box,this suggested St WRKY2-GST combined W-box with specificity. Analysis of St WRKY2 expression level in the root,stem and leaf tissue through the real-time fluorescence quantitative PCR,the results showed that the gene was mainly expressed in the root. Next was leaf and stem. In order to further study the possible function of the gene,potato seeding from tissue culture were treated under 10 μmol / L low phosphorus,10 μmol / L low potassium,200 mmol / L Na Cl,400 mmol / L PEG solution and 4 ℃ low-temperature treatment for 6 h,real-time fluorescence quantitative PCR showed that this gene expression level decreased significantly after low phosphorus treatment,expression of St WRKY2 increased significantly after 200 mmol / L Na Cl and 400 mmol / L treatment. Expression level of St WRKY2 had no obvious change after low potassium and 4 ℃ low temperature treatment. The results indicated that St WRKY2 could in response to low phosphorus,Na Cl and PEG three kinds of abiotic stresses. These results would lay a solid foundation for further study of potato WRKY2 gene function.
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