汽车尾气对小鼠骨髓嗜多染红细胞微核率影响的研究

为了探讨汽车尾气对人及动物的危害，对汽车尾气诱发小白鼠骨髓细胞的微核进行了研究，对远离车间的对照点和车间不同测试点的小白鼠骨髓红细胞的微核率（‰）进行了单因子方差和多重比较分析，测试点与厂区对照点小白鼠骨髓嗜多染色细胞的微核率（‰）差异极显著（P＜0.01）。汽车尾气对动物细胞具有明显的遗传毒理作用，且尾气的浓度越大细胞毒性越大。     

水溶性红曲色素研究

从优质红曲米中分离出一株红曲霉菌株,经亚硝基胍诱变得到一高产水溶性红曲色素变异株ＭＦ－０９８－１２２。对其产色素条件进行了初步探讨,确定以大米粉为碳源,硝酸钠或蛋白胨为氮源,ｐＨ６．０,培养温度３０℃,培养１２０ｈ,其水溶性色素产量最高。     

原生质体诱变选育透明质酸菌株及发酵条件的优化

采用N 甲基 N′ 硝基 N 亚硝基胍(NTG)诱变原生质体,选育了一株产透明质酸的兽疫链球菌(Streptococcuszooepidemicus)H 35.并经过发酵条件的优化,确定了溶菌酶作用的最佳条件是0.6mg/mL溶菌酶高渗溶液,30℃水浴下酶解20min;H 35的最佳发酵条件是葡萄糖4.0%、酵母膏2.0%、NaCl0.2%、KH2PO40.2%、MgSO4·7H2O0.06%、初始pH7.0、35℃,200r/min、培养18h.发酵液中透明质酸的含量从最初的0.47g/L提高到优化后的1.43g/L.     

定点突变技术在转氨酶改造中的应用

对定点突变技术进行简要介绍，并对此技术在改造转氨酶的活性、稳定性及底物特异性等方面的最新进展进行了综述。同时，也对定点突变技术的发展趋势进行了展望。以期能为该领域的研究人员提供有价值的参考。     

牛凝乳酶二硫键功能的研究：Cys45的定位突变

以ｐｈａｇｅｍｉｄ，ｐＢｌｕｅｓｃｒｉｐｔ为载体，把牛凝乳酶原基因的ＫｐｎＩ－ＫｐｎＩ片段插入，将牛凝乳酶４５位Ｃｙｓ用Ａｌａ取代，根据遗传密码子的兼并性，在其附近引入限制性内切酶ＣｌａＩ位点．利用寡聚核苷酸介导的定位突变技术，以获得凝乳酶Ｃｙｓ４５Ａｌａ的突变基因．为了提高突变效率，采用Ｋｕｎｋｅｌ等创立的含尿嘧啶单链ＤＮＡ模板法，经酶切鉴定及双脱氧末端终止法测序，证明已获得预期的突变。     

The role of β18-β19 loop structure in insecticidal activity of Cry1Ac toxin from Bacillus thuringiensis

The β18-β19 loop in domain III of Cry1Ac toxin is unique among Bacillus thuringiensis Cry proteins. In this study, the role of the loop structure in insecticidal activity of Cry1Ac toxin was investigated. Alanine scanning mutations within the loop were initially generated and most mutants were over-expressed and reduced toxicity at different degrees, except mutant N546A that showed almost 2 times enhanced toxicity against Helicoverpa armigera larvae. Further mutagenic analysis of N546 revealed that a charged amino acid in this position would cause very unfavorable influence on insecticidal activity. In addition, the deletion of N546 led to protein instability because of destruction of the loop integrity. Besides, mutant W544F was much more toxic than W544Y, indicating that hydrophobic nature of the position was important for maintaining the stability and activity of Cry1Ac protein. These findings are the first biological evidence for a structural function of β18-β19 loop in insecticidal activity of the Cry1Ac toxin.     

组氨酸标签木聚糖酶的构建表达和酶特性鉴定

通过改变终止密码子位点,在一个耐热木聚糖酶突变株DSB的C末端添加了6聚组氨酸标签(His-tag),并完成了表达和酶特性鉴定.通过PCR将嗜热真菌DSM 10635来源的木聚糖酶突变体基因dsb的终止密码子序列定点突变为谷氨酸密码子序列,连接到表达载体pET-22b(+),转化Escherichia coli.BL21(DE3),诱导表达的耐热木聚糖酶DSB在C端含有6×His-tag.表达产物经硫酸铵分级沉淀和Ni Sepharose层析纯化,获得了电泳纯重组酶DSB.结果表明:添加组氨酸标签的酶与原始酶相比,酶学特性无变化.SDS-PAGE电泳结果显示该酶分子量约为23 kDa,重组酶最适pH为6.5,最适温度为75℃,在pH 5¹0具有良好的稳定性,在pH 6.5,70℃条件下处理30min相对酶活力仍保留80%以上.