白纹伊蚊基因组中CYP6家族新成员基因片段的克隆与分析

目的获得白纹伊蚊CYP6基因家族中新成员的基因片段。方法根据家蝇CYP6A1、CYP6D1以及致倦库蚊CYP6E1同源区氨基酸序列，设计1对简并引物，分别以白纹伊蚊敏感品系和抗性品系基因组DNA为模板进行PCR，产物经T-A克隆转化至JM109大肠杆菌中，经筛选后测序，获得与设计的细胞色素P450基因片段大小相符合的5个基因片段，并将推导的氨基酸序列进行同源性分析。结果在白纹伊蚊敏感品系中获得了2个基因片段(AEDG-S1，AEDG-S2)，核苷酸序列长度分别为240bp和236bp；在白纹伊蚊溴氰菊酯抗性品系中获得了3个基因片段(AEDG-R2，AEDG-R3，AEDG-R4)，核苷酸序列长度分别为236bp、236bp和239bp。所得序列与昆虫CYP6家族的同源性在23.1%～53.8%之间，与CYP3家族的同源性在25.6%～43.9%之间，而与CYP4家族同源性较低，为13.9%～24.4%。结论所得5个序列为CYP6家族中5个新成员的基因片段。

CLONE AND ANALYSIS ON GENE FRAGMENTS OF NEW MEMBERS OF CYP6 FAMILY FROM GENOME OF AEDES ALBOPICTUS

Aim:To obtain new members'gene fragments of CYP6 family from Aedes albopictus. Method: A pair of degenerate primers according to the homologous amino acid sequences of CYP6A1,CYP6D1 in Musca domestica and CYP6E1 in Culex quinquefasciatus was designed and used to amplify CYP6 gene from genomic DNA of sensitive and deltamethrin resistant strains of Ae.albopictus by PCR technique. Five DNA fragments sized as designed were obtained. These fragments were recombined by T A ligation, and cloned into JM109.Five positive clones were selected to sequence. Homologous analysis on these deduced amino acid sequences were executed. Results: Five nucleotide sequences were obtained, of which the length is from 236bp to 240bp.The identities of these cloned sequences to other known members of CYP6 are from 23 1% to 53 8% ,to CYP3 are from 25 6% to 43 9% ,and to CYP4 are from 13 9% to 24 4% . Conclusion:These cloned sequences are gene fragments from new members of CYP6 family.