Study On Dynamic Expression Of As-C,WG,DLL Genes Under Heat Stress Between Insecticide-resistant And-susceptible Plutella Xylostella

Diamondback moth(DBM)(Plutella xylostella L.)is a worldwide pest that harmful to cruciferous vegetables.At present,the control methods of DBM is still concentrated on chemical pesticides,but due to the rapid evolution of drug resistance and unreasonable use of pesticides,DBM has a high resistance to all kinds of insecticides.and DBM has a strong adaptability to the environment,which makes it difficult to control.In our previous study,significantly injury of wing vein showed in chlorpyrifos-resistant(Rc)DBM than chlorpyrifos-susceptible(Sm)DBM under heat stress.This article inherits the above findings,sequenced the wing development-related genes and studied differences of mRNA expression between resistance and sensitive strains under heat stress,and lays a foundation for the physiological and morphological differences of DBM between chlorpyrifos-resistant and chlorpyrifos-susceptible,the main results were as follows:1.According to the wing development patterns of Drosophila and Bombyx mori,the full cDNA sequence of As-C and the partial fragment of DLL,WG were obtained by RACE method.The full-length cDNA of ASH1 is 873 bp,and its ORF is 564 bp,which encodes 187 amino acids,the estimated molecular mass is 20.47 kDa,and an estimated isoelectric point(PI)is 8.81 in DBM;The full-length cDNA of ASH2 is 809 bp,and its ORF is 696 bp,which encodes 231 amino acids,the estimated molecular mass is 25.51 kDa,and the PI is 9.02;The full-length cDNA of ASH3 is 1012 bp,and its ORF is 672 bp,which encodes 223 amino acids,The estimated molecular mass is 24.85 kDa,and an estimated PI of 7.02.The full-length cDNA of ASE is 1799 bp,and its ORF is 1191 bp,which encodes 397 amino acids,the estimated molecular mass is 44.78 kDa and an estimated PI of 4.89.The 4 genes belong to the As-C family gene,which contains the bHLH structure domain.The full-length cDNA of DLL is 1541 bp,and its ORF is 897 bp,which encodes 299 amino acids,the results of multiple sequence alignment show that the structure domain of Hox were conserved in DLL.2.The dynamic expression of the six genes in four heat stress treatment groups were studied with qRT-PCR technique by using laboratory screening resistant and sensitive strains of DBM.The results showed that in the temperature(℃)-treated time(h)groups of(25 ℃-8 h,42℃-4 h,42℃-8 h)and(25℃-16 h,40℃-8 h,40℃-16 h)which contain 4 h-16 h,The basal levels(at 25℃)of mRNA expression of the wing development-related genes in both Sm and Rc DBM pupae were high and no significant difference in most cases.However,all wing development-related genes of Rc pupae under heat stress were significantly down-regulated-expression compared to Sm pupae.In the other two temperatures(℃)-treated time(h)groups of(25℃-1 h,44℃-1 h)and(25℃-48 h,38℃-48 h),since the processing time is too short or too long,the mRNA expression levels of wing development-related genes were low both in Sm and Rc DBM pupae.Meanwhile the mRNA of wing development-related genes was down-regulation expressed significantly in Rc DBM pupae under heat stress compared with Sm DBM pupae.