Cloning, Sequence Analysis And Construction Of Expression Plasmids Of Deltamethrin

Objective To identify expression levels of deltamethrin-resistance associated opsin gene ( NYD-0P7 ) from the resistant strain of Culex pipiens pal lens, to clone ORF of NYD-0P7 and construct prokaryotic and eukaryotic expression plasmids of NYD-0P7.Methods Expression levels of NYD-0P7 between the resistant and the susceptible strains were identified by real time quantitative RT-PCR. The open reading frame (ORF) sequence of NYD-0P7 was cloned by RT-PCR, sequenced in an automatic DNA sequencer (ABI-100) and analyzed with bioinformatic softwares. The prokaryotic and eukaryotic expression plasmids of NYD-0P7 were constructed by gene re-construction methods.Results NYD-0P7 was found to be expressed 9. 78 fold in the resistant strain than in the susceptible strain by real time quantitative RT-PCR. The ORF sequence of 1116bp was obtained for the cDNAs, named NYD-0P7 (GenBank/NCBI AY749413,2004).The deduced protein with 371 amino acid shared 91 ~93% homology respectively, aligned with deduced proteins of NYD-0P1 ~ 6 of Cx. pipiens pallens. The expressed plasmids PGEX-6P-1/NYD-OP7 and pIEl-3/NYD-OP7 were re-constructed successsfully.Conclusion NYD-0P7 expressed higher in the resistant strain than in the susceptible strain of the Culex mosquito, indicating that NYD-OP7 may be associated with the deltamethrin resistance of Cx. pipiens pallens . The results in this work...