| Helicoverpa armigera is one of the most important insect,which damage the cotton.The broadest uses of Bt(Bacillus thuringensis)cotton provide an effective protection against the cotton bollworm and reduce synthetic insecticide use.However,because pests are selected under the pressure of high-dose Bt toxin produced by Bt cotton for a long time,therefore the resistance evolution of pest to Bt cotton should not be ignored.The Bt’s mode of action has not been thoroughly understood and the resistance mechanism of Bt within the insect are complex and variable.The absence and changes of binding sites(cadherin,APN and ALP)in H.armigera,which in turn may be mainly responsible for its resistance to Bt.In order to fully understand the mechanism of resistance and the development of resistance in cotton bollworm,we did some work to study them.The results are as follows.1.The expression levels of seven reference genes(α-tubulin,α-tub;β-actin;actin2;Elongation factor 1 alpha,ef-1α;Glyceraldehyde-3-phosphate dehydrogenase,GAPDH;Trehalose 6-phosphate synthase,Tps and 18S ribosomal RNA,18SrRNA)were significantly different in different development periods,different tissues,different strains and different resistance levels cotton.The conditions of reference genes in q-PCR experiments was strict,the experimental results showed use of two or more reference genes at the same time is better than single one.2、We studied the expressions level of cadherin in different strains of cotton bollworm.Cadherin was distributed in the whole generation cycle of resistant strain and a strain that derived from resistant strain reared without exposure to Cry1Ac,and the lowest expression level occurred in pupal stage.The expression level of cadherin in resistant strain was higher than that in the strain that derived from resistant strain reared without exposure to Cry1Ac.In these two strains,the highest expression level was in second instar.Meanwhile,there are also some different expressions in different tissues.3、APN can express in the whole generation cycle of cotton bollworm,the lowest expression level was in pupal stage and the highest expression level was in third instar.Resistant cotton bollworm had a higher expression level of APN than the strain that derived from resistant strain reared without exposure to Cry 1 Ac.In the same stage,the resistance strain showed a lower expression level,but not in the third instar.The larva from the strain that derived from resistant strain reared without exposure to CrylAc had a no significant expression levels of APN in every stage.The trend of the expression level of APN was like a letter "V" in both of these two strains.4、ALP was very different with cadherin and APN,the expression levels of ALP were all showed significantly higher in resistant strain than that in the strain that derived from resistant strain reared without exposure to Cry1Ac,except in forth instar,which showed higher,but no significant difference.The highest expression levels of APN in the strain that derived from resistant strain reared without exposure to CrylAc occurred in fifth instar.The growth and development of resistant strain is earlier than that of the strain that derived from resistant strain reared without exposure to CrylAc,The highest expression levels of APN in resistant strain occurred in third instar.Both of these two strains displayed contrary tendency of APN’s expression in the first five instar.5、For cotton bollworm larvae from the strain that derived from resistant strain reared without exposure to CrylAc,the expression levels of CAD,APN and ALP were different in the foregut,midgut,hindgut,malpighian tube and cuticula,the trend is midgut>foregut>hindgut>malpighian tube>cuticula.Meanwhile,they all displayed different expression levels in different sex,different parts and different resistant strains.6、When knocking down of the endogenous ALP in susceptible strain,there were no significant differences in mortality,pupation rate and eclosion rate under the treatment without Bt,by comparing with control. |
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