The Functional Analysis Of P450 Genes In The Brown Planthopper (Nilaparvata Lugens)

Cytochrome P450 enzymes are a complex family of hemi-containing enzymes found in virtually all aerobic organisms. It can regulate and control metabolism and transformation in endogenous substances or exogenous substance. The genome of each insect species may carry a hundred or so different P450 genes. P450 metabolism of certain plant chemicals is often the key to insects resistant to insecticides and the adaptation of plant poisonous substances. Furthermore, P450 enzymes are found in the biosynthetic and metabolic pathways of ecdysteroids and juvenile homnones.In this study, we identified 68 P450 genes via searching the genomic and transcriptomic sequences. Detecting the gene of CYP18A1, CYP304H1, CYP303A1 growth specificity expression and tissue specific expression. QRT-PCR analysis showed that CYP18A1, CYP304H1 genes within the whole BPH nymphae ages are expressed and the expression increased little by little, the CYP303A1 gene also have expressed throughout the larva instar,but in contrast to the former two genes, its expression reduced gradually. Interestingly, all the genes in BPH adult hardly expressed. QRT-PCR analysis showed that CYP18A1 was most expressed in wing bud,the following is Ovary and integument; CYP304H1 was most expressed in fat body and salivary glands; CYP303A1 was most expressed in integument and wing bud. After microinjecting to the fifth instar BPH with dsRNA of CYP18A1,CYP3O4H1, CYP303A1, all the genes expression levels was significantly down-regulated. The integument of 5th instar nymphs appeared deformity and could not normally stretch. The molting appeared abnormal in 3rd,4th or 5th instars, and all finally died. We amplified CYP304H1 gene with 522bp in length from BPH cDNA by PCR, which is potential to encode a predicted 22.3 kDa protein containing 174 amino acid residues. CYP304H1 gene was successfully expressed in Escherichia coli and expressed proteins was used to prepared polyclonal antibodies.In this study, we first reported the new functions of CYP304H1 found in insect molting. Our results laid a basis for the further study of P450 and for RNAi-based control of this important rice pest.