白纹伊蚊细胞色素P450 CYP6N亚家族新成员5‘—及3’—非翻译区序列功能分析

目的 获得白纹伊蚊细胞色素P450 CYP6N亚家族新成员5‘－及3‘－非翻译区（untranslated region,UTR）核苷酸序列,并进行功能分析。方法 以本文作者已获得的白纹伊蚊溴氰菊酯抗性株－新的细胞色素P450 CYP6基因cDNA片段,设计1对特异性引物,以反向引物,进行5‘-cDNA末端快速扩增（rapid amplification of cDNA ends,RACE）,以正向引物进行3‘-RACE。然后分别进行克隆、测序和鉴定。以获得5‘-UTR及3‘－UTR,并运用PC／GENE软件分析其功能。结果 获得CYP6N家族中两个新成员5‘－UTR及3‘－UTR;其结构特征包括：两个新成员前导序列均为46个核苷酸;起始密码子ATG两侧核苷酸－3位是A、＋4位是T;5‘－UTR区域无稳定性的发卡结构;终止密码子在CYP6N3为TZAA、在CYP6N4为TAG,紧挨其3‘端核苷酸分别为A和G,翻译产物羧基端均为亮氨酸。结论 成功地获得了白纹伊蚊细胞色素P450CYP6N亚家族两个新成员5‘－及3‘－非翻译区序列,进一步分析表明该两个新成员均能进行有效翻译,昆虫细胞色素P450基因翻译起始调控与翻译终止调控有部分类似于脊椎动物mRNA的翻译调控,但具有多态必性的特点。

FUNCTIONAL ANALYSIS OF 5' - AND 3' -UNTRANSLATED REGIONS OF NEW MEMBERS OF CYTOCHROME P450 CYP6N SUBFAMILY FROM AEDES ALBOPICTUS

Aim To obtain 5' and 3' untranslated region (UTR)sequences of new members of cytochrome P450 CYP6N subfamily from Aedes albopictus and analyze its function.Method A pair of specific primers according to an obtained cDNA fragment of cytochrome P450 CYP6 gene from deltamethrin-resistant Ae.albopictus were designed and used to amplify complete 5'-end and 3'-end by rapid amplification of cDNA ends (RACE)technique.The RACE products were cloned,sequenced and identified.The function of the obtained 5'- and 3'-UTRs were analyzed by PC/GENE software.Results The structural characters of 5'- and 3'-UTRs of two new members of CYP6N subfamily from Ae.albopictus include:both of 5'-UTRs have 46 nucleotides;the nucleotide at-3 position in franking sequences surrounding the start code is A,at 4 position is T;any stable hairpin structures in 5'-UTRs can be not found.The obtained 3'-UTRs have a typical stop code TAA or TAG,and there is a typical conserved A or G at the position following closely 3'-terminus of the stop code.The carboxy terminals of translated products are all leusine.Conclusions 5'- and 3'-UTRs of two new members of cytochrome P450 CYP6N subfamily from Ae.albopictus were successfully obtained.The mRNAs of the obtained new members of CYP6N subfamily can be effectively translated.The translation mechanisms of P450s in insects are partially similar to ones of most eukaryotic genes,but characterized by multiplicity.