【摘要】 L 乙内酰脲酶产生菌节杆菌BT80 1的染色体DNA经Sau3AI部分酶切后分离 30kb左右的片段 ,与经HpaI和PstI酶切的黏粒载体pKC5 0 5进行连接 ,将连接产物用包装蛋白包装 ,转染大肠杆菌DH5α得到 10 0 0 0多个转化子 ,构建成节杆菌BT80 1的基因组文库。通过薄层层析等方法筛选得到了 1个阳性克隆 ,通过亚克隆得到了乙内酰脲酶的完整基因 ,该基因能分别利用自身的启动子和T5启动子在大肠杆菌中进行表达产生有活性的蛋白更多还原

【Abstract】 Hydantoinase can be widely used in enzymic production of various amino acids. In order to obtain the hydantoinase genes in Arthrobacter BT801, its chromatosomal DNA is isolated and partialy digested with Sau 3A I to collect fragments of about 30kb. Then, this fragment is inserted into the Hpa I and Pst I site of cosmid pKC505. The genomic library was thus constructed by packing in vitro with λ phage package protein and transfecting E.coli DH5α. A positive transformant was selected from the library using thin layer chromatography and other methods. A DNA fragment containing complete hydantoinase genes was sequenced by sub cloning into pUC18. The gene can express active protein under control of its own promoter and T5 promoter in E. coli. The isolation of the gene established foundition for research and application of the hydantoinase.更多还原