高产胸苷磷酸化酶的大肠杆菌的诱变筛选

摘要

Objective:Escherichia coli can produce thymidine phosphorylase, which can promote the thymidine into the thymine, is the key enzyme in the synthesis of nucleoside drugs. Method: We used UV induced mutagenesis in Escherichia coli to produce a large number of thymidine phosphorylase, established a rapid and simple mutation screening methods. UV induced mutation was used two times, for the first time in 60 seconds, then stop for 10min later, continued exposure 5~30s. It could increase the positive mutation rate of Escherichia coli. Result: Finally we screened out the strains with high amount of thymidine phosphorylase , E17、E23、E32. Conclusion: After ultraviolet mutagenesis, the three strains of Escherichia coli producing ability of thymidine phosphorylase is stronger than that of the original strain, and it can be inherited stably.%目的：大肠杆菌（Escherichia coli）产生的胸苷磷酸化酶能够促进胸苷转化为胸腺嘧啶，是合成核苷类药物的关键酶。本研究利用紫外线诱导法诱变大肠杆菌使其高产胸苷磷酸化酶，并筛选出3株能够高产该酶的菌株。方法：紫外诱变采用二次照射法，第1次照射60s后停止照射10min后，各继续照射5～30s梯度时间，增加大肠杆菌的正突变率。结果：筛选出E17、E23、E32三株产胸苷磷酸化酶量较高的菌株。结论：经过紫外诱变筛选后，这3株大肠杆菌产胸苷磷酸化酶的能力比原始菌株强，并且可以稳定遗传。