Clone And Analysis Acetylcholinesterase1Gene Of Insecticide Resistant And Susceptible Plutella Xylostella,Gene Frequency Analysis And Properties

Acetyl cholinesterase (acetyl cholinesterase, AChE, EC3.1.1.7) is a kind of serinehydrolase, the substrate of which is acetylcholine in vivo. Acetylcholine is one of theseries of neurotransmitter, which is an important carrier of the action potentialtransmission between adjacent cells. The character of generation and degradation ofAChE is a necessary condition for organisms to maintain normal physiologicalfunction.The target of organ phosphorus insecticides is acetyl cholinesterase (AChE) invivo, which combined with acetyl cholinesterase.The function of pesticide in vivo isinhibition or reduced enzyme activity, which interferes the degradation of the enzymeacetylcholine. Therefore, Acetylcholine signaling is still have biological activity,which leads to the disorder of the organisms' normal physiological activity. So thepurpose of using pesticide is achievement. Nowadays, by virtue of the uniquecharacteristics of the growth and development of plutella xylostella (diamondbackmoth, DBM), intensive use of pesticide has led to the development of resistance inDBM, which seriously reduces agricultural production and farmers' income.In this paper, the sensitivity to chlorpyrifos in the insecticide-resistant (R) and-susceptible (S) DBM, the level of sensitivity of AChE to insecticide in the R and SDBM, the gene of ace-1cloning and analysis, and the detection of frequency ofresistance ace-1allele genotype were studied. This paper made a systematicalexpatiation on the reason why DBM become resistant to the insecticide, the level ofresistibility to chlorpyrifos in the R and S DBM, the change of the frequency of ace-1allele genotype without the insecticide stress.According to the ace-1gene fragment of DBM which is published in the NCBI,the full-length of ace-1from the R and S DBM were cloned and sequenced.Comparing the two deduced proteins, there mutation sites: Ala201Ser,Gly227Ala,Ala441Gly, were found, respectively. The two complete amino acid sequences ofAChE deduced from the cDNA consisted of19residues for the putative signal peptidein the N-terminal amino acid sequences, respectively. The presence of three mutation sites made its secondary structure change. This change in the secondary structure wasconsistent with that in the tertiary structure. By analysis, we found that the mutationsite of Ala201Ser and Ala441Gly doesn't affect the space configuration of AChE, andcontribute to the insecticide resistance little.The insecticide sensitivity and the level of sensitivity of AChE to insecticide inthe R and S DBM, as well as the detection of frequency of resistance ace-1allelegenotype were determined. The results showed that the R DBM (ace-1allele genotypecomposition:3.3%RR,92.5%RS,4.2%SS) displayed9.3-folds resistance to the SDBM (ace-1allele genotype composition:33.3%RS,66.7%SS). The sensitivity ofAChE to insecticide in the R DBM displayed0.268-folds to the S DBM. Thecomposition of ace-1allele genotype was consistent with the results of biochemicalbioassays.We found that the frequency of RS genotype showed a downward trend duringthe following generations, using the bi-PASA molecular technology to detect the fieldresistance of diamondback moth, which are selected under the insecticide pressure.On the contrary, the frequency of SS genotype showed a clear upward trend during thefollowing generations and both in the F7generations tend to be slower. We analyzedthat this was due to the Plutella xylostella, out of the selection pressure, having adecline in fitness. Therefore, in this condition, the advantage of the resistance gene inthe population disappeared, which result in the change of the frequency.