【作者】 李光涛；

【导师】 何康来；

【作者基本信息】 中国农业科学院 ， 农业昆虫与害虫防治， 2004， 硕士

【摘要】 亚洲玉米螟Ostrinia furnacalis（Guen（?）e）是我国转Cry1Ab杀虫蛋白基因Bt抗虫玉米的主要靶标害虫，大面积种植转单一基因的Bt玉米有可能会引发亚洲玉米螟对Bt杀虫蛋白的抗性。只有实施合理的抗性治理对策，才能使这一现代化技术成果得以持久的利用。深入研究抗性种群的生物学和遗传规律，是制定抗性治理计划、实施合理的抗性治理措施的基础。本论文从生物学和遗传学的角度，首次就亚洲玉米螟对Cry1Ab杀虫蛋白抗性的种群生物学和抗性遗传规律以及与Cry1Ac杀虫蛋白之间的交互抗性关系作了系统的研究。主要结果如下： 通过饲料混合法以Cry1Ab杀虫蛋白对亚洲玉米螟进行室内汰选38代，与敏感种群相比，抗性种群的幼虫发育历期明显延长，平均蛹重明显下降。用表达Cry1Ab杀虫蛋白的MON810玉米对Cry1Ab抗性种群连续4代生测结果表明，亚洲玉米螟初孵幼虫不能在MON810玉米的心叶和苞叶上存活，但是能够在花丝上完成幼虫期并化蛹和繁殖后代，首次用转基因作物离体生测的方法发现了室内汰选的亚洲玉米螟抗性种群可以在MON810花丝上完成幼虫发育历期。在羽化期相同的条件下，抗、感种群雄蛾间交尾竞争力没有显著差异，抗、感雌蛾对敏感雄蛾的竞争力差异不显著，但对抗性雄蛾的竞争力，抗性雌蛾显著强于敏感雌蛾。当环境中抗性雄蛾的种群密度相对高时，发生抗性种群内交尾的概率要高，将增加亚洲玉米螟对Bt玉米产生抗性的风险。抗、感种群的交尾日活动节律一致。抗、感种群非滞育老熟幼虫的过冷却点没有显著差异；而抗性种群滞育幼虫的过冷却点显著高于敏感种群及F₁代约1～2℃。 用饲料混合法测定了亚洲玉米螟Cry1Ab抗性种群、敏感种群及其正反交F₁和Cry1Ac抗性种群对Cry1Ab的毒力回归曲线；同时测定了Cry1Ab、Cry1Ac抗性种群以及敏感种群对Cry1Ac的敏感水平。从LC₅₀计算正反交的D_LC（抗性基因显性水平）分别为0.22和0.63，这表明目前亚洲玉米螟对杀虫蛋白Cry1Ab的抗性遗传为受母性影响的不完全隐性遗传。以Bt玉米花丝作为衡量标准，对抗性种群和对照种群及其F₁代进行生测，结果表明其有效显性为完全隐性遗传(D_ML=0)。MON810花丝中Coy1Ab蛋白表达量足以杀死杂合个体，符合高剂量加庇护所的要求。比较不同种群对Cry1Ab和Cry1Ac的毒力回归曲线和LC₅₀可以看出，以Cry1Ab汰选的对Cry1Ab产生了107倍抗性的Cry1Ab抗性种群，对Cry1Ac产生了10.4倍的抗性；以Cry1Ac汰选对Cry1Ac产生了14.0倍抗性的Cry1Ac抗性种群，对Cry1Ab产生了约6倍的抗性。这说明亚洲玉米螟对Cry1Ac和Cry1Ab存在着很强的交互抗性。Cry1Ab抗性种群去掉选择压后，在普通饲料上连续饲养3代后，幼虫发育历期开始下降，6代以后基本上与敏感种群一致，而且蛹重也有不断增加的趋势。MON810花丝生测表明，亚洲玉米螟对Cry1Ab杀虫蛋白的抗性不能稳定遗传，失去选择压后，其抗性会迅速丢失。更多还原

【Abstract】 Asian corn borer, Ostrinia furnacalis Guenee, (ACB), is the primary insect pest targeted by transgenic Bt com in China. Bt corn can provide season long protection from this insect, but its success will be short-lived if ACB adapt to Bt corn. The primary strategy for delaying ACB resistance to Bt com is to provide refuges of non Bt corn and other host plants. However, the feasibility of refuge strategy depends on the biology of resistant colony and inheritance of resistance. We investigated the biology of a laboratory-selected CrylAb resistant colony of ACB and inheritance of resistance, which included the survival of resistant colony fed on diet incorporated with CrylAb protein and transgenic Bt corn event MON810, mating attractiveness between resistant and susceptible colonies, cold tolerance among resistant, susceptible and their hybrid progeny F1 colonies, resistant gene dominant level, mater effect, stability, and the cross resistance between CrylAb and Cryl Ac.There were 70% survival of larvae and 60% pupation rate when resistant colony fed on the diet incorporated with 100ng/g CryAb. The larval developmental time of resistant colony was significantly longer (5 days) than susceptible. The average pupal weight of resistant colony was 42.0% less than susceptible. When the resistant colony was exposed to the high dosage of CrylAb protein diet, the survival and the pupation decreased in a few generations and then increased. This indicated that ACB would evolve resistant to CrylAb when it was selected by the CrylAb protein.Bioassays with neonate of resistant and susceptible colonies infested on whorl leaves, fresh husks and silks of Bt corn Event MON810 for 4 generations, showed that the resistant colony could not survive more than 7 days on the whorl leaves and husks of Bt corn, but it could complete the development of larva and average pupation rate was 14.5%. This was the first report that laboratory-selected Bt resistant colony could complete the development of larva on fresh silk of Bt maize. The expression of CrylAb protein in the silk of Event MON810 was 585.46ng/g (fresh weight) according to enzyme-linked immunosorbent assays.Among four type of mating combinations (RR SS/RR, SS SS/RR, SS/RR SS, SS/RR RR), the inter-population mating was 43-62.5% in the combinations of RR SS/RR, SS SS/RR, and SS/RR SS, no significantly was observed between the inter-population and intro-population mating, i.e. the mating between resistant and susceptible moths was random. However, the inter-population mating was only 28.0% in the combinations of SS/RR RR, which was significantly lower than intro-population mating. This indicated that the resistant males were more attracted to the resistant females, i.e. the mating between the inter- and intro-population. In this case, when the population density of resistant males was higher, the mating within resistant population could be higher, and the resistant homozygous of offspring were increase. Both resistant and susceptible moths had the same daily mating activity rhythm. This indicates that if resistant and susceptible moths emerge synchronizing well, the daily mating activity rhythm would not affect the random mating between resistant and susceptible colonies.The suppercooling points were tested for diapaused and non-diapaused larvae of resistant and susceptible colonies and their F1 progeny. There was no significant difference in suppercooling points and freezing points between resistant (-11.5 C) and susceptible (-11.4 C) colonies of non-diapausedlarvae. The suppercooling point of diapause larvae from resistant colony was significant higher than those from susceptible and FI progeny of resistant and susceptible colonies. There were no significant different among susceptible and F1 progeny of resistant and susceptible colonies. Although the suppercooling point was only 1.5 C higher for resistant colony than susceptible, this could not impact the overwintering of resistant larvae.Susceptibility to the CrylAb protein was determined by dose mortality response for resi更多还原