两个扶桑绵粉蚧细胞色素P450基因的克隆与分析

Cloning and analysis of two cytochrome P450 genes from Phenacoccus solenopsis

为了深入研究入侵生物扶桑绵粉蚧(Phenacoccus solenopsis Tinsley)对农药等外源物的作用机制,采用RACE的方法克隆了2个扶桑绵粉蚧细胞色素P450基因的开放阅读框序列.其中一个基因的开放阅读框包含1 536 bp的片段,编码512个氨基酸,其相对分子质量为59 843.5,理论等电点为8.79,推测属于稳定存在的蛋白,且没有跨膜区域;其具有WXXXR(WKNLR)、EXXR(ETLR)序列及FXXGXXXCXG/A(FGEGPRICIG)血红色素结合位点等细胞色素P450特有的氨基酸序列,其中ETLR属于CYP6家族所特有,且其blast比对均与昆虫体内CYP6A家族的相似性较高,将其命名为PsCYP6a1.另一个基因的开放阅读框包含1 575 bp的片段,编码525个氨基酸,相对分子质量为61 246.62,理论等电点为8.87,推测属于稳定存在的蛋白,且没有跨膜区域;与前者相似,也包含了相应的细胞色素P450基因的保守位点,将其命名为PsCYP6a2.2个扶桑绵粉蚧细胞色素P450基因的核苷酸序列的相似性为55.56%,氨基酸序列的相似性为39.41%,相似的残基占了44.12%.

In order to study the Phenacoccus solenopsis＇ degradation mechanism to pesticides and other xenobiotics,two Cytochrome P450 genes were cloned by the RACE method.One of them contained 1 536 bp Open reading frame（ORF） that encodes 512 amino acids.Its relative molecular mass was 59 843.5,and the isoelectric point was 8.79.The protein should be stable,with no transmembrane region.It got WXXXR（WKNLR）,EXXR（ETLR） and FXXGXXXCXG/A（FGEGPRICIG）,a heme binding site,which belongs to cytochrome P450 specific sequence.The ETLR was specific to CYP6 family.Also,the gene got the highest similarity with the insects of the CYP6 A family.The gene was named as PsCYP6a1 by the analysis.Another gene contained an ORF of 1 575 bp fragment,encoding 525 amino acids.The relative molecular mass was 61 246.62,while its theoretical isoelectric point was 8.87.The protein also was stable,with no trans membrane region.As to PsCYP6a1,it also contained the cytochrome P450 conserved sites.So it was named as PsCYP6a2.The nucleotide sequence similarity of the two cytochrome P450 gene was 55.56%,the amino acid sequence similarity of 39.41% residues similar accounted for44.12%.Two P.solenopsis cytochrome P450 genes that belong to CYP6 family were successfully cloned.