家蚕核型多角体病毒P10基因的克隆及核苷酸序列分析

CLONING AND SEQUENCING OF PIO GENE FROM THE BOMBYX MORI NUCLEAR POLYHEDROSIS VIRUS

杆状病毒P10基因与多角体蛋白基因一样,都是由强启动子控制的高效表达晚期基因,且非病毒复制所必需,故在构建杆状病毒载体表达系统方面受到研究工作者的重视。苜蓿尺蠖核多角体病毒(Autographa californica nuclear polyhedrosis virus,AcMNPV)和黄杉毒蛾核型多角体病毒(Orgyia pseudotsugate nuclear polyhedrosis virus,OpMNPV)

A32P-labeled cloned DNA fragment ( AcMNPV EcoR I-P) containing the P10 gene from Autographa californica multicapsid nuclear polyhedrosis virus ( AcMNPV ) was used to probe Southern blots containing restriction endonuclease digests of Bombyx mori nuclear polyhedrosis virus DNA.The fragments of 0.5 and 1.1kb were positive and cloned into pUC18. The 1.1kb fragment was mapped with restriction endonucleases, and reprobed with the AcMNPV EcoR I-P fragment. Two small regions of about 200bp and 700bp were cross-hybridized. DNA sequencing in these regions showed an open reading frame of 285bp which shared detectable homology with the P10 gene of AcMNPV. The 5'-and 3'-flanking regions had been sequenced. It was revealed that the BmNPV P10 gene coulden code the P10 protein which has 93 amino acid residues. The homology of the nuc leotide sequences of P10 structural gene and the amino acids sequences of P10 protein between BmNPV and AcMNPV reached 95% and 86%, respectively. A comparison between the 5'-and 3'-flanking sequences of the AcMNPV and BmNPV P10 genes was made. This study may provided the basis for further understanding the function of the BmNPV P10 gene promoter, and for constructing the expression vectors with the P10 gene promoter.