Genetic transformation and pyramiding of aprotinin-expressing sugarcane with cry1Ab for shoot borer (Chilo infuscatellus) resistance
Genetic transformation and pyramiding of aprotinin-expressing sugarcane with cry1Ab for shoot borer (Chilo infuscatellus) resistance
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DOI：
10.1007/s00299-010-0829-5
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发表时间：
2010-04-01
影响因子：
6.2
通讯作者：
Subramonian, N.
中科院分区：
文献类型：
--
作者：
Arvinth, S.;Arun, S.;Subramonian, N.
在离体条件下，研究了Cry 1Aa、Cry 1Ab和Cry 1Ac毒素对甘蔗茎螟Chilo infuscatellus Snellen（鳞翅目：草螟科）幼虫的杀虫活性。在三种毒素中，Cry 1Ab的LC 50值最低，是最有效的。利用基因枪和农杆菌介导法，将玉米泛素启动子驱动的cry 1Ab基因导入甘蔗品种Co 86032和CoJ 64。利用cry 1Ab对转牛胰蛋白酶抑制剂（抑肽酶）基因的甘蔗植株进行转基因研究。Southern分析证实了在农杆菌介导的转化体中，在粒子轰击和单位点整合的情况下，转基因的多个整合。通过Western分析证明cry 1Ab的表达，并使用ELISA定量毒素。Cry 1Ab蛋白在不同转化体中的表达量在0.007 ~ 1.73%之间，农杆菌法转化的转化体表达量显著高于对照。在体内生物测定与新生幼虫的蛀虫，转基因产生了相当低的百分比deadhegarts尽管遭受取食损害的蛀虫相比，未转化的对照植物。表达的Cry 1Ab含量与死心损伤呈负相关。用cry 1Ab处理的表达抑肽酶的甘蔗也表现出损害的减少。根据所获得的结果，讨论了利用cry 1Ab和抑肽酶基因生产抗早梢螟转基因甘蔗的潜力。
We evaluated the insecticidal toxicity of Cry1Aa, Cry1Ab and Cry1Ac toxins against neonate larvae of sugarcane shoot borer Chilo infuscatellus Snellen (Lepidoptera: Crambidae) in vitro on diet surface. With the lowest LC50 value, Cry1Ab emerged as the most effective among the three toxins. Sugarcane cultivars Co 86032 and CoJ 64 were transformed with cry1Ab gene driven by maize ubiquitin promoter through particle bombardment and Agrobacterium-mediated transformation systems. Gene pyramiding was also attempted by retransforming sugarcane plants carrying bovine pancreatic trypsin inhibitor (aprotinin) gene, with cry1Ab. Southern analysis confirmed multiple integration of the transgene in case of particle bombardment and single site integration in Agrobacterium-mediated transformants. The expression of cry1Ab was demonstrated through Western analysis and the toxin was quantified using ELISA. The amount of Cry1Ab protein in different events varied from 0.007 to 1.73% of the total soluble leaf protein; the events transformed by Agrobacterium method showed significantly higher values. In in vivo bioassay with neonate larvae of shoot borer, transgenics produced considerably lower percentage of deadhearts despite suffering feeding damage by the borer compared with the untransformed control plants. Expressed Cry1Ab content was negatively related to deadheart damage. Aprotinin-expressing sugarcane pyramided with cry1Ab also showed reduction in damage. The potential of producing sugarcane transgenics with cry1Ab and aprotinin genes resistant to early shoot borer was discussed in the light of the results obtained.