Potential novel bZIP-like gene for resistance to Erysiphe necator identified in Chinese wild Vitis pseudoreticulata

Abstract

In this study, a novel bZIP-like gene was isolated from Chinese wild Vitis pseudoreticulata W. T. acc. Baihe-35-1. The full-length complementary deoxyribonucleic acid (cDNA) sequence of the gene was 1583 bp including 159 bp 5’ untranslated region (UTR), 365 bp 3’ UTR and a 1083 bp ORF which encodes a polypeptide of 360 amino acids with a molecular weight of 38.662 kDa. The deduced amino acid sequence shares an overall 46 to 69.8% sequence similarity with bZIP from other plants. Therefore, we designated this gene as V. pseudoreticulata bZIP (VpbZIP-like). The expression of VpbZIP-like was induced 12 h post inoculation (hpi) by Erysiphe necator, but transiently decreased, then increased in these two genotypes and its expression was lower in highly resistant genotype Baihe-35-1 than in susceptible genotype Hunan-1 at 24, 48 and 72 hpi. We further tested whether the expression was also a response to plant signaling molecules. Results indicate that the susceptible genotype Hunan-1 showed higher expression of VpbZIP-like than the highly resistant genotype Baihe-35-1 after exogenous application of methyl jasmonate (MeJA), salicylic acid (SA) and ethephon (Eth). Moreover, tissue specific expression pattern of VpbZIP-like was analyzed by reverse transcription-polymerase chain reaction (RT-PCR). Results reveal that it was in lower lever in flower than in leaf, stem, tendril and fruit. The CDS of VpbZIP-like was inserted into the prokaryotic expression construct pGEX-4T-1, and then transformed into Escherichia coli BL21-code induced by isopropyl-b-D-thiogalactopyranoside (IPTG) which resulted in the production of a Mr. 64 kDa of GST- VpbZIP-like fusion protein displayed in sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE).

Key words: Chinese wild Vitis, bZIP, gene expression, signaling molecules, fusion protein expression.