Diagnosis of different genotypes of the mutations using allele-specific PCR.
Five individual forth instar larvae from both JA and TH populations were detected using both susceptible-specific (W) and resistant-specific (M) primers, respectively. Samples from the JA population are all homozygous wild type at all three mutation sites, while the sample TH-2 are heterozygous at Q4594L and G4946E and TH-34 are heterozygous at E1338D, homozygous mutation genotype at Q4594L and homozygous wild genotype at G4946E. TH-38, 39 and 40 are all homozygous mutation genotypes at all three sites. The products of PCR were examined on a 2.5% agarose gel and stained with ethidium bromide after electrophoresis. The length of the products for E1338D, Q4594L and G4946E detection are 116 bp, 168 bp and 71 bp, respectively. L represents the DL 2000 DNA Marker (Takara Biotechnology, Dalian, China).