State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, PR China.
J Invertebr Pathol. 2013 Sep;114(1):31-3. doi: 10.1016/j.jip.2013.05.005. Epub 2013 May 24.
A pooled clone method was developed to screen for cry2A genes. This metagenomic method avoids the need to analyse isolated Bacillus thuringiensis strains by performing gene specific PCR on plasmid-enriched DNA prepared from a pooled soil sample. Using this approach the novel holotype gene cry2Ah1 was cloned and characterized. The toxin gene was over-expressed in Escherichia coli Rosetta (DE3) and the expressed toxin accumulated in both the soluble and insoluble fractions. The soluble Cry2Ah1 was found to have a weight loss activity against Ostrinia furnacalis, and a growth inhibitory activity to both Cry1Ac-susceptible and resistant Helicoverpa armigera populations.
我们开发了一种用于筛选 cry2A 基因的克隆池方法。这种宏基因组方法通过对从混合土壤样本中制备的质粒富集 DNA 进行基因特异性 PCR，避免了对分离的苏云金芽孢杆菌菌株进行分析的需要。使用这种方法，我们克隆并鉴定了新型的 cry2Ah1 基因。该毒素基因在大肠杆菌 Rosetta (DE3)中过表达，表达的毒素在可溶性和不溶性部分都有积累。可溶性 Cry2Ah1 对玉米螟具有体重减轻活性，对 Cry1Ac 敏感和抗性棉铃虫种群均具有生长抑制活性。
