摘要
目的优化重组葡萄糖脱氢酶(GDH)基因表达条件,研究其稳定性。方法在摸索培养基性质、抗生素浓度、培养温度、表达时间等条件的的基础上确定最佳表达条件;同时在表达酶液中添加甘油、咪唑等以期提高其稳定性。结果当氨卞青霉素浓度为100mg/ml,LB培养基中,37℃培养条件下,细菌生长接近饱和后更换等体积新鲜LB培养基诱导过夜,可获得较高的蛋白表达量;同时在表达酶液中添加20%甘油或20%甘油+0.05M咪唑可使酶蛋白基本稳定。结论载体的拷贝数、稳定性和宿主菌的生长状态是影响GDH表达的因素;一定浓度的甘油可提高葡萄糖脱氢酶的稳定性。
Objective To optimize the conditions of expressing recombinated glucose dehydrogenase（GDH） gene and improve its stability. Methods On the bases of testing the conditions of expressing GDH gene, such as culture medium, concentration of antibiotics, tempreture of bacterium growing and the length of shaking, the ideal expression conditions were concluded. Glycerol or mizol was added into the culture medium to see whether its stability could be inproved. Results The higher level of GDH expression was obtained when the concentration of ammonia penicillin （AMP） was upto 100 μg/ml, temperature was 37℃, and the culture medium was changed with the same volum of LB medium at the time of saturating growing for overnight. The stability of GDH was remarkably increased by adding 20% glycerol or 20% glycerol combined with 0. 05 M mizol. Conclusion The factors of influencing GDH gene expression are the number and stability of vector copies and grOwing status of host. A certain amount of glycerol can increase the stability of GDH expression.
出处
《江苏医药》
CAS
CSCD
北大核心
2008年第7期712-714,共3页
Jiangsu Medical Journal
基金
镇江市科技计划(社会发展)资助项目(SH2005028)
