Cloning, Sequence Analysis And Expression Vector Construction Of A Novel Deltamethrin-resistance Associated Gene NYD-MLC2 From Culex Pipiens Pallens

[Objective] To identify differential expression levels of NYD-MLC2 gene in deltamethrin-resistance and susceptible strain of Culex pipiens pallens, to clone full length cDNA of NYD-MLC2 and construct prokaryotic and insect expression plasmids of NYD-MLC2.[Methods] Expression levels of NYD-MLC2 in the resistant and the susceptible strains were identified by real time quantitative RACE. The full length cDNA of NYD-MLC2 was cloned by RACE, then sequenced, and analyzed with bioinformatic softwares. The prokaryotic and insect expression plasmids of NYD-MLC2 were constructed by gene recombination methods.[Results] NYD-MLC2 was found to express 4.08 fold higher in the resistant strain than in the susceptible strain by real time quantitative RT-PCR. Full length cDNA of NYD-MLC2 was obtained (GenBank/NCBI DQ140391, 2005). The sequence of opening reading frame (ORF) sequence is 630bp. The deduced protein with 210 amino acids shared more than 25 % homology with proteins of myosin light chain 2 of many other species. The prokaryotic expression plasmids pET32a/NYD-MLC2 and insect expression plasmids pIB/V5 His-TOPO/NYD-MLC2 were constructed successsfully.[Conclusion] NYD-MLC2 expressed higher in the resistant strain than in the susceptible strain of the Culex mosquito, indicating that NYD-MLC2 may be associated with the deltamethrin resistance of Cx. pipiens pallens. The results in this work are valuable for further study of the relationship between NYD-MLC2 gene and insect resistance.