Gene Clone Bcl-2 And Bax And Expression Under Heat Stress In The Resistant And Susceptible Plutella Xylostella

Plutella xylostella is a worldwide pest,major harm cruciferous vegetables and is thought to be an important model pest for studying the resistance level to insecticides.In order to reveal the relationship of the fitness between the chlorpyrifos-resistant(R)and susceptible(S)DBM(larva,pupa and adult)under heat stress,effects of heat stress on expression levels of mitochondrial apoptosis-related bcl-2 and bax genes.This article clones gene by homology cloning method and has been cloned DBM mitochondrial apoptosis gene bcl-2 and bax.The full length cDNA of bcl-2(Gene Bank:KP271439)was 1309 bp cluding a 978 bp ORF encoding a putative protein of 325 amino acids.The calculated molecular mass and isoelectric point of deduced protein were 35.48 kDa and 9.60,respectively.The full length cDNA of bax(Gene Bank:KT427406)was 1037 bp cluding a 738 bp ORF encoding a putative protein of 245 amino acids.The calculated molecular mass and isoelectric point of deduced protein were 28.24 kDa and 8.50,respectively.By protein structure prediction,bcl-2 and bax genes have been found to contain bcl-2 family function structure domain.Through the multiple sequence alignment,it found that bcl-2 and bax have high homology with lepidoptera insects.By constructing the genetic evolutionary tree,the result found lepidoptera insects other close relatives.Based on biological information analysis,bcl-2 and bax genes are highly conserved during evolution.It lays the theoretical basis to further research of mitochondrial apoptosis gene expression under heat shock and signal transmission mechanism in resistance and sensitive DBM.By applying qRT-PCR technology,the relative mRNA expression of bcl-2 and bax genes was determined in resistance(resistance heterozygous and homozygous resistant)and sensitive Plutella xylostella.Expressions of bax were found in DBM larvae and adults treated with 37 ? and 42 ?for 3 h,and the extent of up-regulated expression of these genes was higher in R DBM than that in S DBM,On the whole.the relative mRNA expression of bcl-2 was lower.As compared to 25 ?treatment,38 ? for 48 h,40 ? for 8 or 16 h,42 ? for 4 or 8 h and 44 ? for 1 h resulted in up-regulated expression of gene in DBM pupae,and significantly higher up-regulated expression of bax observed in R DBM than in S DBM.And the mRNA expression of bcl-2 was downward trend.High temperatures(33.5-50 ? for 3 h)could induce up-regulated expression of bax in DBM adults.With the increase of temperature,the expression of bax increased and reached at a plateau at 47 ?.However,high temperatures(33.5-50 ? for 3 h)could reduce expression of bcl-2 in DBM adults.The results show that expression level of bax will rise,while expression level of bcl-2 decreased,after high temperature treatment.It may be the high temperature to promote the expression of pro-apoptotic gene,and reduce expression of antiapoptotic gene.Differential expression mechanism of bax and bcl-2 in the resistance and sensitive Plutella xylostella populations,mRNA expression of bax and bcl-2 may play an important role.in the regulation and sensitive Plutella xylostella.