淡色库蚊抗溴氰菊酯品系CYP4家族新成员基因克隆及序列分析  

CLONING AND SEQUENCE ANALYSIS OF CYTOCHROME FAMILY 4 NEW GENES FROM THE DELTAMETHRIN-RESISTANT MOSQUITO,CULEX PIPIENS PALLENS

作　　者：滕达[1] 李秀兰[1] 公茂庆[1] 马磊[1] 陆洵蔚[1] 孙艳[1] 朱昌亮[1] 

机构地区：[1]南京医科大学病原生物学系,江苏南京210029

出　　处：《中国寄生虫病防治杂志》2004年第2期65-67,共3页Chinese Journal of Parasitic Disease Control

基　　金：国家自然科学基金资助项目 (No .30 1 70 835) ;江苏省教育厅自然科学基金资助项目 (No.0 1KJB31 0 0 0 6) ;南京师范大学生命科学学院江苏省资源生物技术重点实验室开放研究课题资助项目

摘　　要：目的　获取淡色库蚊CYP4家族新基因。　方法　根据CYP4氨基酸保守序列设计一对简并引物 ,采用RT PCR的方法 ,从淡色库蚊抗溴氰菊酯品系四龄幼虫总RNA中扩增出与设计相符的基因片段。利用T/A克隆方法 ,将获得的基因片段克隆入 pGEM Teasy载体 ,经PCR鉴定重组成功 ,测序并进行比对。　 结果　共克隆出 3 6个阳性克隆 ,将其中 9个随机挑取的阳性克隆测序及同源性分析 ,表明为CYP4家族中 9个新的cDNA序列 ,由GeneBank登录上网 ;经国际细胞色素P45 0命名委员会鉴定 ,属CYP4家族CYP4H、CYP4J亚家族。为进一步研究细胞色素P45 0的多样性及其与抗药性的关系打下基础。　结论　克隆 9个淡色库蚊基因部分序列 ,被鉴定属于CYP4家族CYP4H、CYP4J亚家族。Objective To obtain the partial cDNA sequences encoding cytochrome P450 from the deltamethrin-resistant mosquito,Culex pipiens pallens . Methods The conserved region in the alignment of insect P450 family 4(CYP4) proteins served as a guide for the synthesis of degenerate oligonucleotide primers to clone P450 cDNA. Primers were used in the reverse transcription-polymerase chain reaction (RT-PCR) of RNA from the 4th instar larvae of deltamethrin-resistant Cx. pipiens pallens . PCR products purified were cloned into pGEM R-T Easy Vector and sequenced. Results 9 of 36 positive clones obtained shown to be new sequences encoded cytochrome P450 and appraised belonging the subfamily CYP4H and CYP4J by the Nomenclature Committee of Cytochrome P450 (GeneBank CB074944-CB074951;CB270837,2003). Conclusion 9 new CYP4 subfamily genes were obtained from Cx. pipiens pallens .

关 键 词：淡色库蚊 细胞色素P450 基因克隆 CYP4 

分 类 号：R384.1[医药卫生—医学寄生虫学]