酿酒酵母的连续复合诱变及其突变株的快速筛选

为了获得高产的工业用酿酒酵母(Saccharomyces cerevisiae)菌株,利用己烯雌酚(DES)和UV对酿酒酵母菌QD进行连续复合诱变,采用改进的发酵小管集气法结合乙醇定量测定来进行突变株的筛选,并与常规复合诱变及筛选方法进行比较.结果表明:连续复合诱变的正向突变率最大值为18.9%,与常规诱变方法的最大正向突变率15.6%相比提高了21.2%,说明运用该诱变方法能获得较高的正向突变率;改进的筛选方法可以快速筛选出目的突变株,同时,获得了一株高产乙醇的突变株,在甘蔗汁发酵培养基中其乙醇产量达到9 720 mg/100 mL,比常规诱变方法筛选到的突变株乙醇产量(9 170 mg/100 mL)高出6.0%;与常规诱变方法相比,连续复合诱变方法不仅能够得到较高的正向突变率,而且能够获得更加高产的突变株.     

屎肠球菌hyl基因突变株的构建

目的：构建屎肠球菌类透明质酸酶（hyaluronidase,hyl）基因突变株,研究hyl基因的功能。方法：用pETX4577质粒构建屎肠球菌hyl基因重组自杀质粒,通过体内同源重组,筛选获得hyl基因的突变株,体外研究hyl基因缺失对突变株生长能力的影响。结果：经同源重组,利用卡那霉素抗性筛选,PCR、脉冲场电泳和southern blot进行鉴定获得基因突变株＊hyl。突变株在体外的生长能力低于野生株。结论：hyl基因突变株构建成功,hyl基因在生长中起着重要的作用,可能是屎肠球菌的毒力因子之一。     

两种拟南芥coi1突变体的鉴定与初步分析

COI1(coronatine-insensitive 1)蛋白是茉莉酸信号途径的信号开关,coi1突变体是茉莉酸信号调控机制研究中的重要实验材料。对从拟南芥生物资源中心购买的两种COI1(At2g39940)的T-DNA插入突变体SALK-095916和SALK-045434,通过基因型分析筛选获得了纯合突变体。进一步的基因表达检测和茉莉酸敏感性试验表明,两种纯合突变体中COI1基因的表达量分别约为野生型植株的40%和50%,突变体幼苗对茉莉酸的敏感性显著降低。获得的SALK-095916(命名为coi1-22)和SALK-045434(命名为coi1-23)纯合突变体可以作为实验材料在进一步的茉莉酸信号途径机制研究中使用。     

大肠杆菌holo-ACP突变体的表达纯化及相应acyl-ACP的性质初探

酰基-酰基载体蛋白（acyl-acyl carrier protein,acyl-ACP）作为酰基供体,在脂肪酸、聚酮等天然产物合成途径中具有重要作用.目前,常以酰基-辅酶A（acyl-CoA）来代替尚未商品化的acyl-ACP进行相关酶的体外活性研究.由于底物蛋白acyl-ACP的ACP部分与相关酶发生相互作用,影响酶的催化特性,这种替代无法真实认识相关酶的酰基转移特性.本文以大肠杆菌pET-28a（+）-ACP为模板,构建12株单点突变体,表达纯化出相应holo-ACP,并进一步合成C16:0-ACP和C18:1-ACP.高效液相色谱的结果表明,ACP单点突变会影响acyl-ACP整体的性质,其中T40残基的改变对acyl-ACP的疏水性、紫外吸收响应和稳定性均产生了显著影响.ACP突变体的性质表征为acyl-ACP与相关酶的互作机制研究奠定了基础.      

Enhanced H+ transport activity of tonoplast vesicles isolated from roots of salt-tolerant mutant of wheat under NaCl stress

Both enhanced were H⁺ transport activities of tonoplast vesicles isolated from roots of the salt-tolerant mutant and wild type of wheat with treatment of NaCl, but the activity of the mutant was significantly higher than that of wild type. H⁺ transport activity was indicated as the stable value of fluorescence quenching per mg membrane proteins. The H⁺ transport activities dependent on ATP of the mutant and wild type were 1099 and 558 respectively and their activities dependent on PPi were 358 and 228 separately.     

Discovery of a male-biased mutant family and identification of a male-specific SCAR marker in gynogenetic gibel carp Carassius auratus gibelio

Gibel carp (Carassius auratus gibelio) is a uniquely gynogenetic species with a minor ratio of males in natural habitats, but its male origin and sex determination mechanisms have been unknown. In this study, a male-biased mutant family was discovered from the gynogenetic gibel carp, and a male-specific SCAR marker was identified from the mutant family. Normal spermatogenesis was observed in the male testes by immunofluorescence histochemistry. Nearly identical AFLP profiles were observed between males and females, but a male-specific 86 bp AFLP fragment was screened by sex-pool bulked segregant analysis and individual screening. Based on the male-specific AFLP fragment, a total of 579 bp sequences were cloned by genome walking. Subsequently, a male-specific SCAR marker was designed, and the male-specific DNA fragment was confirmed to be steadily transmitted to the next generation and consistently detected only in males.     

茄假单胞菌寄主花生特异性毒性基因的定位

通过对从茄假单胞菌中克隆的pGX1252进行亚克隆分析,发现含pGX1252右边4.5kbKpnI/EcoRI片段的亚克隆pGX1404仍象pGX1252一样能使对花生不致病菌株T2014在花生上致病.用转座子Tn5-loc对pGX1404进行了饱和插入诱变,一共分离得到6个在pGX1404上不同位置的独立插入突变,其中离pGX1404右末端约1.4kb、2.2kg的两个Tn5-loc插入使pGX1404丧失了扩大T2014寄主范围的能力,证实hsv基因位于pGX1404的右边.     

莱茵衣藻类胡萝卜素生物合成及其遗传控制的研究进展

随着对类胡萝卜素药用、食品、化妆等方面认识的提高,人类对类胡萝卜素的种类和产量需求也越来越高。单细胞真核微藻-莱茵衣藻是国际上研究光合作用遗传控制的最佳模式生物,也是天然类胡萝卜素主要来源之一。本文主要介绍了类胡萝卜素生物合成途径及用莱茵衣藻研究类胡萝卜素光合遗传控制等方面的最新研究进展。     

一株产谷氨酸菌株的复合诱变选育及突变株的生物学特性

对产谷氨酸的谷氨酸棒杆菌(Corynebacterium glutamicum)进行以硫酸二乙酯(DES)和紫外线(UV)相结合的复合诱变,经选育得到一株高产突变株LG - 01.将其接种在含葡萄糖初始浓度为150.0 g/L,初始pH值为7.2～7.6的液体培养基中,于30℃下培养48 h后,其谷氨酸产量达到106.10 g/L,比原始菌株提高了25.74％(同等条件下,原始菌株的谷氨酸产量为84.38 g/L).对该突变株的产谷氨酸代谢特性进行研究,结果表明:其最适初始pH值为7.2,最佳培养温度为30 ℃,以葡萄糖为最佳碳源,其谷氨酸产量最高时相应的葡萄糖浓度为150.0 g/L,其某些生物学特性发生了改变,如延迟期变长,对数期的细胞生长能力较强,对底物的利用率比原始菌株约提高5％等.