论文部分内容阅读
为探索棉铃虫Helicoverpa armigera液泡型ATP酶(vacuolar-type proton ATPase,V-ATP酶)亚基A对棉铃虫生长发育的影响及其在Bt杀虫机制中的作用,采用PCR结合RACE技术克隆了棉铃虫V-ATP酶亚基A基因序列,通过实时荧光定量PCR技术测定了其在棉铃虫不同发育历期和幼虫肠道不同组织中的表达量;并比较了4龄幼虫取食含Cry1Ac蛋白饲料后中肠中该基因表达量的变化。结果显示,棉铃虫V-ATP酶亚基A基因全长2 578 bp(Gen Bank登录号KP090287),开放阅读框1 863 bp,编码621个氨基酸。V-ATP酶亚基A高度保守,不同物种间氨基酸序列同源性大于90%。V-ATP酶亚基A在棉铃虫整个生育期都有表达,在4龄幼虫体内表达量最高,是卵期的3.00倍;在幼虫肠道不同组织中,中肠中表达量最高,是后肠中的2.65倍。4龄棉铃虫幼虫取食含Cry1Ac蛋白的人工饲料后,中肠V-ATP酶亚基A的表达受到抑制,表达量为对照的0.39~0.81倍。表明V-ATP酶亚基A基因可能参与棉铃虫的生长发育和代谢过程,并可能与抵御Cry1Ac的毒杀作用有一定关系。
In order to explore the effect of subunit A of Helicoverpa armigera vacuolar-type proton ATPase on the growth and development of cotton bollworm (Helicoverpa armigera) and its role in Bt insecticidal mechanism, we cloned The expression of V-ATPase subunit A gene of Helicoverpa armigera was detected by real-time fluorescence quantitative PCR in different developmental stages of larvae and larvae. The 4th instar larvae were fed with Cry1Ac Changes in expression of the gene in the midgut after feeding. The results showed that the subunit A gene of cotton bollworm V-ATPase was 2 578 bp in length (Gen Bank Accession No. KP090287). The gene was 1 863 bp in open reading frame and encoded 621 amino acids. V-ATPase subunit A is highly conserved, with amino acid sequence homology greater than 90% among different species. V-ATPase subunit A was expressed throughout the growth period of H. armigera, and was the highest in the 4th instar larvae, 3.00 times that of the egg stage. The expression of V-ATPase subunit A was the highest in the midgut of different tissues of larvae, 2.65 times in the intestine. After 4-day-old cotton bollworm fed artificial diet containing Cry1Ac protein, the expression of V-ATPase subunit A in the midgut was inhibited, and the expression level was 0.39 ~ 0.81 times that of the control. The results indicated that the subunit A gene of V-ATPase may be involved in the growth and metabolism of cotton bollworm and may be related to its resistance to the toxic effects of Cry1Ac.