关键词: 梨小食心虫, 化学感受蛋白, 分子克隆, 序列分析, 原核表达, 融合蛋白

Abstract: In order to clarify the physiological function of chemosensory proteins (CSPs) in the olfactory system of Grapholita molesta, the full-length cDNA encoding a chemosensory protein was isolated from G. molesta by using reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends-PCR (RACE-PCR), and named as GmolCSP (GenBank accession no. JQ821389). The results of sequence analysis indicated that the open reading frame (ORF) of GmolCSP is 384 bp in length, encoding 127 amino acid residues, with the deduced molecular weight (MW) of 12.80 kD and isoelectric point (pI) of 8.33. The deduced amino acid sequence of GmolCSP showed a high identity to CPSs of other lepidopteran insects. RT-PCR analysis revealed that GmolCSP was expressed in antennae, head, thorax, abdomen, wings and legs of the test moth. GmolCSP was then constructed into the expression vector pET-32a and expressed in Escherichia coli BL21 (DE3) after induction with IPTG.-SDS-PAGE and Western Blot analysis confirmed the molecular weight of the recombinant GmolCSP is 29 kD, consistent with the predicted result. The results in this study are helpful for further research on molecular structure and function of GmolCSP.

Key words: Grapholita molesta, chemosensory protein, molecular cloning, sequence analysis, prokaryotic expression, fusion protein