Molecular Mechanisms Of Metabolic Resistance To Thiamethoxam Of An Alien Invasive Pest: B-biotype Bemisia Tabaci

The B-biotype of the whitefly, Bemisia tabaci (Gennadius) (Homoptera:Aleyrodidae) is an agricultural pest that distributes extensively and causes serious economical damage. Thiamethoxam has good effects for controlling the sucking pest, including the whitefly, as the model of the second new neonicotinoid insecticides and also the first s-neonicotinoid insecticide. In order to reveal the resistance mechanism of whitefly to thiamethoxam, this thesis cloned the P450 gene, CYP6CM1, from the resistant and susceptible strains of whitefly, and the sequences were aligned and analyzed. The mRNA levels of the CYP6CM1 gene from resistant and susceptible strains were compared through the real-time PCR technique. The RNAi was carried out preliminarily. The results were as follows.(1) The P450 gene, CYP6CM1, was cloned successfully from the thiamethoxam resistant and susceptible B. tabaci. A 1591bp sequence had been achieved from the susceptible strains B. tabaci, including an open reading frame of 1563 nucleotides, encoding a protein of 521 amino acids. The nucleotide length of CYP6CM1 gene from the resistant strain was 1591bp. Three nucleotides and two amino acids were found to be different between the resistant and susceptible strains. The positions of replaced nucleotides were 1048, 1357, and 1377 bp, resulting in 2 amino acids replacement of E350G and P452T. The similarity of nucleotides and identity of amino acids of sequences were 99.81% and 99.62%.(2) The mRNA expression levels of CYP6CM1 gene were compared between the resistant and susceptible strain of B. tabaci using real-time quantitative PCR. The results showed that the mRNA expression of CYP6CM1 in resistant strain was 3.66-fold higher than the susceptible strain, with 86.91 and 23.72 respectively. The mRNA expression levels preliminarily suggested that the over-expression of CYP6CM1 was probably associated with high resistance to thiamethoxam.(3) RNAi experiment was carried out preliminarily. Two CYP6CM1 cDNA fragments have been designed. One fragment was a length of 223bp sequence located at 1185bp-1408bp and another was 218bp located at 750bp- 967bp, which were the template of synthesizing the corresponding dsRNA, providing a basis for the next work.