利用密码子优化提高Bt cry1Ah基因在转基因玉米(Zea mays L.)中的表达

Improvement of Bt cry1Ah Gene Expression in Transgenic Maize (Zea mays L.) Through Codon Optimization

Bt cry1Ah基因是本实验室从苏云金芽胞杆菌(Bacillus thuringiensis,Bt)菌株BT8中克隆的一种新型杀虫蛋白基因。在前期工作中已根据植物密码子偏好性对cry1Ah基因进行过一次优化。根据玉米密码子偏好性对cry1Ah基因进行第二次优化,并将两次优化的cry1Ah基因分别构建了pAhmG和pmAhb植物表达载体,利用农杆菌介导法转化玉米自交系综31,分别得到10株和9株阳性转基因玉米植株。通过ELISA、Westernblot检测及生物活性检测,比较不同优化的基因在玉米中的蛋白表达量及抗虫情况。初步结果表明外源基因在玉米植株中可以稳定表达并可以遗传。二次优化基因比一次优化基因在玉米中的蛋白表达量更高,抗虫性更好。表明密码子优化有利于提高外源基因的表达。

The crylAh gene is one of novel insecticidal genes cloned from Bacillus thuringiensis isolate BTS. The sequences of crylAh gene had been optimized according to the codon bias of plant in former research. In this study it was optimized again according to the codon bias of maize. Two plant expression vectors pAhmG and pmAhb harboring crylAh gene by different codon optimization were constructed and transferred into maize （Zea mays L. ） mediated by Agrobacterium. Ten and nine regenerated plants were PCR positive, respectively. Comparison of gene expi＇ession and insect-resistance in transgenic maize with different codon optimization was carried out through ELISA, Western blot detection and bioassay. The preliminary results showed that the foreign gene crylAh could be expressed and inherited stably in maize. The protein expression and insect-resistance in transgenic plants harboring the second codon optimization of crylAh gene were better than the first codon optimization. So, codon optimization can improve expression of foreign genes.