抗性家蝇para基因的选择性剪接和位点突变

ALTER SPLICING AND POINT MUTATION IN PARA-LIKE GENE FROM PYRETHROID RESISTANT HOUSEFLY (MUSCA DOMESTICA)

为了在分子水平上研究拟除虫菊酯抗性家蝇靶标抗性机制 ,根据昆虫para型钠通道Ⅱ区S4至S6及Ⅱ与Ⅲ连接区的保守区域 ,设计简并引物 ,利用降落RT PCR ,克隆拟除虫菊酯kdr (R品系 )家蝇para型钠通道 70 4bp的cDNA ,根据此序列用PrimerPremier5 0设计特异引物 ,分别扩增敏感 (SS品系 )、super kdr(RR品系 )家蝇各 6 5 1bp的cDNA。结果SS无有义突变 ,R和RR在ⅡS6CTT到TTT的替换导致Leu 10 14到Phe的保守性突变 ,RR在ⅡS 4～ 5细胞内片段接头处没有与超 -kdr抗性相关的Met到Thr突变 ;另外首次发现kdr (R品系 )家蝇在Ⅱ与Ⅲ连接区出现 39bp的选择性剪接 ,构成家蝇para钠通道亚型 (命名为xych ,GenBank登陆号为AF4 114 5 2 )。

To investigate the molecular mechanism of target resistance to pyrethroid in housefly, cDNA fragments of para like gene locating between domain Ⅱ and Ⅲ in susceptible (SS), resistant (R) and super resistant (RR) strains were cloned by RT PCR with general primers. Sequencing analysis showed that the conserved point mutation of C to T,which results in an amino acid change from 1014 Leu to Phe, appeared in cDNA fragment from both R and RR strains. However, no other mutations that may cause super Kdr were found. Otherwise, We first found 39bp alter splicing base from kdr housefly in the connecting linkers between Ⅱ and Ⅲ domains, which composes of subtype of sodium channel (named xych, GenBank accession number: AF411452).