Screening of Interacting Proteins in the Coat Protein of Areca Palm Yellow-ing Leaf Virus 1
In order to identify the pathogenesis of Areca palm leaf yellowing virus 1(APLYV 1)and explore the interaction between APLYV 1 and its host protein,a areca leaf yeast two-hybrid cDNA library was constructed.Using coat protein(CP)of APLYV 1 as bait,the decoy vector pGBKT7-CP was constructed,and the interacting proteins were screened from areca cDNA library by yeast two-hybridization.The results showed that the areca yeast library was successfully constructed with a library capacity of more than 1.1 ×107,a recombination rate of 100％,and an average insert length of more than 1 000 bp.The bait vector pGBKT7-CP was successfully con-structed.Through the self-activation test,the positive control could grow bacteria normally and show blue color on the four-deficiency medium plate,while the negative control and experimental group could grow bacteria normally on the two-deficiency medium plate,but did not grow bacteria on the three-deficiency and four-deficiency medium plate,indicating that the successfully constructed recombinant bait vector had no self-activation activity and could be used in the screen library experiment.A total of 13 positive clones were screened,and 10 candidate areca pro-teins,including sHSP(small Heat Shock Protein)and DnaJ protein,were obtained by comparison between NCBI website and areca transcriptome library.DnaJ protein was selected and predicted by physical and chemical proper-ties analysis and secondary structure analysis.The total length of the aele gene was 1 035 bp,the theoretical molecu-lar weight of the protein was 37.9 kD,the isoelectric point pI was about 5.05,and the secondary structure was com posed of random curl(49.13％)and α-helix(25.87％),without any signal peptide.There is no transmembrane do-main,mainly located in the nucleus.The results of this study provide a basis for further research on the function and expression regulation mechanism of APLYV 1 protein.
Areca palm leaf yellowing virus 1Y east libraryProtein interactionCoat proteinHeat shock protein
万方数据
维普
