Modification of the rib operon derived from Bacillus subtilis and its expression in Escherichia coli

A riboflavin operon （rib operon） derived from Bacillus subtilis 368 was modified on structure and the resulting operons were expressed in various strains of Escherichia coli. The results showed that the optimization of the rib operon and the host strain used for expression are two main factors affecting the riboflavin production. Replacing the promoterl and rfn box of the rib operon with a strong constructive promoter spol drastically increased the expression of the rib genes. When E. coli JM109 was used as the host strain, the highest riboflavin production reached 95.3μg/mL （about eight times higher than that of the unmodified r/b operon）. In addition, when tetracycline （20 μg/mL） was used as the selective pressure, compared with the ampicillin resistant transformants, a higher riboflavin yield was obtained in tetracycline resistant host strain.     

甘露醇和AgNO_3对小麦细胞再分化的影响

比较了甘露醇和AgNO3 对小麦 (TriticumaestivumL.)品种保7059幼穗细胞再分化和IAA氧化酶、IAA过氧化物酶活性的影响 .结果表明 ,AgNO3 能够明显提高小麦愈伤组织再生率 ,但甘露醇使其再生率降低 ,AgNO3 对IAA氧化酶和IAA过氧化物酶活性无明显影响,但甘露醇使2种酶活性明显升高 .     

Red/ET重组AHBA生物合成基因簇打靶载体的构建与鉴定

为确定3-氨基-5-羟基-苯甲酸(AHBA)生物合成基因簇在链霉菌中与次生代谢产物的关系,运用PCR技术,从33株AHBA合酶基因阳性菌株扩增与AHBA生物合成基因簇中编码AHBA合酶(A)、氧化还原酶(O)、磷酸化酶(P)基因,获得24株AOP基因阳性菌株.根据靶基因A基因下游和P基因上游同源序列设计50 bp引物,中间插入卡那霉素抗性基因的DNA片段,进行PCR,获得外源DNA片段.经过电转化,将外源DNA片段和pKC1139-AOP重组质粒共转入含重组酶质粒大肠杆菌HS996/ pSC101-BAD-gba-(Tet).在Red重组酶的作用下,外源DNA片段与重组质粒pKC1139-AOP上的AHBA基因簇的同源区域重组,构建了AHBA基因簇打靶载体.研究显示了Red/ET重组工作效率高、操作简单、精确的优点,可大大缩短构建打靶载体的时间.