Foc4 2个谷胱甘肽S-转移酶基因的克隆、鉴定及其在外源氧化胁迫下的表达

为鉴定香蕉枯萎病菌(尖孢镰刀菌古巴专化型4号生理小种,Fusarium oxysporum f.sp.cubense race 4,Foc4)中的2个假想谷胱甘肽S转移酶(GSTs),采用RT-PCR方法克隆了这2个GSTs基因cDNA编码序列,随后分别将2个基因定名为Fogst1和Fogst2。其中,Fogst1的开放阅读框长609 bp,编码202个氨基酸残基,Fogst2的开放阅读框长693 bp,编码230个氨基酸残基。进化树分析表明:Fogst1属于GSTs超家族的sigma(σ)亚型成员,Fogst2属于GSTs超家族中目前未知的亚家族成员。为了验证Fogst1和Fogst2的表达,分别构建了Fogst1和Fogst2的原核表达重组载体pET28a-Fogst1和pET28a-Fogst2,并将pET28a-Fogst1和pET28a-Fogst2转化到大肠杆菌表达菌株BL21,经IPTG诱导后获得以可溶形式表达的重组蛋白Fogst1和Fogst2。GSTs活性分析表明,以CDNB为底物检测,2个重组蛋白均具有GSTs酶活性。分别取外源氧化胁迫处理后1、5、12、24 h菌丝样品进行相对荧光定量PCR分析,结果表明:Fogst1和Fogst2在前5 h表达量均大幅上调,表达量随后下调并恢复正常水平。这些结果均暗示Fogst1和Fogst2可能参与了Foc4抗外源氧化胁迫过程。     

Chloroplast glyceraldehyde-3-phosphate dehydrogenase contains a single disulfide bond located in the C-terminal extension to the B subunit.

Mass mapping analysis based on cyanylation and CN-induced cleavage indicates that the two cysteine residues in the C-terminal extension of the B subunit of the light-activated pea leaf chloroplast glyceraldehyde-3-phosphate dehydrogenase form a disulfide bond. No evidence was found for a disulfide bond in the A subunit, nor was there any indication of a second disulfide bond in the B subunit. The availability of the structure of the extended glyceraldehyde-3-phosphate dehydrogenase from the archaeon Sulfolobus solfataricus allows modeling of the B subunit. As modeled, the two cysteine residues in the extension are positioned to form an interdomain disulfide cross-link.     

新城疫病毒非编码序列的遗传演化趋势

【目的】探讨新城疫病毒全基因序列中非编码序列的分子演化规律。【方法】结合本研究室2012年自产蛋下降鸭群中分离测序的一株鸭源新城疫病毒全序列,从GenBank下载35株不同基因型新城疫病毒全长cDNA序列,获取非编码序列,分别绘制引导序列、尾随序列、F-HN及HN-L基因间隔序列(IGS)的遗传进化树,比较编码基因内5’及3’UTR序列核苷酸序列替代特点。【结果】非编码序列的长度及位置高度保守,而其核苷酸基因序列在不断发生变异,且变异趋势与编码基因序列相一致。【结论】新城疫病毒在整个基因组上编码和非编码序列同步发生变异。     

Biodegradation of volatile organic compounds by five fungal species

Five fungal species, Cladosporium resinae (ATCC 34066), Cladosporium sphaerospermum (ATCC 200384), Exophiala lecanii-corni (CBS 102400), Mucor rouxii (ATCC 44260), and Phanerochaete chrysosporium (ATCC 24725), were tested for their ability to degrade nine compounds commonly found in industrial off-gas emissions. Fungal cultures inoculated on ceramic support media were provided with volatile organic compounds (VOCs) via the vapor phase as their sole carbon and energy sources. Compounds tested included aromatic hydrocarbons (benzene, ethylbenzene, toluene, and styrene), ketones (methyl ethyl ketone, methyl isobutyl ketone, and methyl propyl ketone), and organic acids ( n-butyl acetate, ethyl 3-ethoxypropionate). Experiments were conducted using three pH values ranging from 3.5 to 6.5. Fungal ability to degrade each VOC was determined by observing the presence or absence of visible growth on the ceramic support medium during a 30-day test period. Results indicate that E. lecanii-corni and C. sphaerospermum can readily utilize each of the nine VOCs as a sole carbon and energy source. P. chrysosporium was able to degrade all VOCs tested except for styrene under the conditions imposed. C. resinae was able to degrade both organic acids, all of the ketones, and some of the aromatic compounds (ethylbenzene and toluene); however, it was not able to grow utilizing benzene or styrene under the conditions tested. With the VOCs tested, M. rouxiiproduced visible growth only when supplied with n-butyl acetate or ethyl 3-ethoxypropionate. Maximum growth for most fungi was observed at a pH of approximately 5.0. The experimental protocol utilized in these studies is a useful tool for assessing the ability of different fungal species to degrade gas-phase VOCs under conditions expected in a biofilter application.     

生物破乳剂产生菌的筛选及其方法研究

针对生物破乳剂产生菌筛选难的问题, 采用显色法、溶血细胞测试法、表面张力测定法和排油圈法从6种不同菌源对生物破乳菌产生菌进行了筛选。通过试验筛选得到了17株生物破乳剂产生菌, 其中24h内破乳率高于70%的破乳菌有5株; 油田含油污泥、采油废水生物处理污泥和污水处理厂剩余污泥是筛选破乳菌的较好的菌源; 显色法、溶血圈法存在检测范围的局限性; 表面张力测定法和排油圈法是最为简易和准确的生物表面活性剂产生菌的筛选方法, 采用模型乳状液对生物破乳剂产生菌进行筛选最为直接和准确, 但工作量大、所需时间长, 因此在筛选高效破乳菌时, 建议采用表面张力、排油圈法进行初筛, 而后通过模型乳状液破乳进行验证。     

植酸酶phyAm基因结构延伸突变改善酶的热稳定性

将来源于黑曲霉N25的植酸酶基因phyA^m重组于大肠杆菌表达载体pET-30b（＋）,以重组表达载体pET30b-FphyA^e为模板经PCR扩增获得结构延伸突变植酸酶基因phyA^m（在植酸酶基因C端增加了来源于pET-30b-FphyA^m载体上13氨基酸残基）。含突变基因的重组表达载体pPIC9k-phyA^e在GS115酵母中表达。纯化的突变酶pp-NP^e与野生型酶PP-NP^m-8相比：PP-NPA^e的最适反应温度上升了3气,75℃处理10min,热稳定性提高21％,比活力略有提高。最适反应pH为5．6,有效pH范围pH4,6到pH6．6。比未突变酶扩大了0.4单位。     

滇产回心草及回心康抗缺血心肌脂质过氧化作用及对前列环素／血栓素的影响

采用结扎大鼠冠脉造成急性心肌缺血模型,动态观测Ⅱ导联心电图ST段的变化,以S-T段上移为指标反映缺血程度,观察了滇产回心草及回心康对急性心肌缺血损伤的保护作用,同时检测血清SOD、MDA、PGI2和TXA2.结果表明,回心草及回心康均能显著减少S-T段上移,均以2 g/kg组为显著;其1 g/kg及2 g/kg均使心肌梗塞范围缩小(p＜0.05,p＜0.01);均使心肌缺血大鼠血清SOD显著提高(p＜0.05及p＜0.01);MDA显著降低(p＜0.05,p＜0.01).回心草及回心康还能提高大鼠血清PGI2水平,降低TXA2含量,以回心草2 g/kg及回心康2 g/kg为显著(p＜0.05),呈剂量依赖性.实验结果提示,回心草及回心康均具有抗心肌缺血及抗脂质过氧化作用,其提高PGI2/TXA2可能与其抗心肌缺血作用机制有关.     

我国新分离虫媒病毒的初步鉴定

1990－1994年,从新疆地区的蚊、蜱和病人血清分离了多株病毒,为了明确这些病毒的分类地位,对其中的20株病毒进行了组织培养细胞感染实验和血清学检验,对部分毒株做了动物接种实验和理化性质鉴定。结果显示：20株病毒均可使BHK－21细胞病变（1－3天）,主要表现为细胞圆宿、聚集,融合,破碎,脱落等;致Vero细胞病变为2－4天;15株病毒致C6／36细胞病变（2－4天）,5株病毒对C6／36细胞连续观察7天未见细胞病变。11株病毒对乳鼠2－4天致死,对成年鼠2－5天致死。选取6株病毒进行理化性质鉴定,4株病毒（90260、91002、91004和91028）对5－氟脱氧尿苷耐受,对乙醚和酸敏感,提示为有膜RNA病毒;一株病毒（90265）对5－氟脱氧尿苷、乙醚和酸均敏感,提示为有膜DNA病毒;另一株病毒（9059）对5－氟脱氧尿苷耐受,对乙醚和酸也耐受,提示可能为无膜RNA肠道病毒。20株病毒中,17株病毒与甲病毒、乙型脑炎病毒和布尼亚病毒的特异性免疫腹水不反应,提示这些病毒中可能不存在甲病毒、黄病毒和布尼亚病毒;3株病毒（90260、91002和91004）只与甲病毒的特异性免疫腹水反应,与乙型脑炎病毒和布尼亚病毒的不反应,提示这三株病毒为甲病毒。