[Cloning and identification of cytochrome P450 resistance related genes in the mosquito, Culex pipiens pallens].

ABSTRACT

OBJECTIVE: To inquire into the relationship between cytochrome P450 and deltamethrin resistance. METHODS: 24 new cDNA sequences encoding cytochrome P450 were amplified respectively from deltamethrin-susceptible and -resistant strains of Culex pipiens pallens with a pair of degenerate primers according to the conservative amino acid sequences of CYP4 in insects by RT-PCR and the Direct Cloning Method, and then were identified by cDNA chip and reverse Northern. RESULTS: 112 positive clones were obtained, of which 24 were shown to be new sequences encoded for cytochrome P450. They have been lodged in GenBank and were appraised by the Nomenclature Committee of Cytochrome P450, belonging to the subfamily CYP4C, CYP4D, CYP4H and CYP4J in CYP4 family. The hybrid signal values of 6 P450 sequences (NYDS3, NYDS5, NYDR6, NYDR9, NYDR15 and NYDR17) were 3.1-9.7 times higher in the resistance probe than in the susceptible probe, and NYDR17 only reacted with the resistance probe. The result of reverse Northern in NYDR15 was similar to that of cDNA chip. CONCLUSION: CYP4 is related to deltamethrin resistance and the specific expression caused by point mutation of cytochrome P450 gene may exist in deltamethrin-resistant Cx. pipiens pallens.