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Resources research on thermophilic protease-producing bacteria in normal temperature environment
常温环境中产嗜热蛋白酶菌资源

WANG Guo-hui,
王国惠

生态学杂志 , 2007,
Abstract: 通过计数、分离与筛选,对常温环境嗜热菌和产嗜热蛋白酶菌的分布及资源状况进行了研究。结果表明,常温环境中存在着一定数量的嗜热菌和产嗜热蛋白酶菌。土壤与水体相比,其嗜热菌资源相对丰富,且耕作肥沃的土壤中产嗜热蛋白酶菌多于贫瘠土壤;在水环境中,无论湖水、江水还是处理中的废水,在常温条件下均有一定比例的嗜热菌和产嗜热蛋白酶菌。在啤酒废水曝气阶段,产嗜热蛋白酶菌占嗜热菌的比例较大,达45%。本研究筛选的1株嗜热菌其产嗜热蛋白酶活性较高,该菌株在pH7.6、温度68℃条件下其蛋白酶活力达到642U·ml^-1。该项研究为开发产嗜热蛋白酶菌资源,在工业和环境治理等方面的应用提供重要科学依据。
Isolation and characterization of Bacillus thermophilic bacteria AT07-1 and its application in sludge solubilization and bio-hydrogen production by anaerobic fermentation
嗜热菌株AT07-1的分离鉴定及其在污泥溶解预处理厌氧发酵产氢中的应用

汤迎,李小明,杨永林,杨麒,曾光明,郭亮,朱小峰,李焕利,潘维
环境科学学报 , 2009,
Abstract: 从湖南大学花园土壤中采集样品,用稀释涂布法分离和纯化适于污泥溶解的典型嗜热菌菌株,从中选出一株高效菌进行形态观察,并对其进行生理生化鉴定.结果表明,该菌株的革兰氏染色为阳性,呈细杆状,产芽孢,为好氧或兼性需氧细菌,最适生长温度为65℃,最适生长pH值为6.8~7.5.通过16SrDNA碱基测序和对比证实,该菌株是目前尚未报道过的一株嗜热菌,GenBank中注册命名为Bacillus thermophilic bacteria AT071(注册号:FJ231108).同时,将其纯种菌用于嗜热酶溶解(solubilization by thermophilic enzyme,S-TE)污泥稳定化处理,接种该嗜热菌可促进污泥中悬浮固体的溶解,2.5d时接种试验挥发性悬浮固体 (volatile suspended solid,VSS)溶解率达到59.41%,比不接种试验同期提高了25.58%,达到了EPA规定的污泥稳定化要求.进一步对经AT071预处理的污泥厌氧产氢效果进行研究,试验结果显示,接种处理后污泥发酵产氢效果良好,高出未接种处理(65℃热处理)污泥26.4%,发酵气体中只含有H2和CO2.
Process Characteristics of Bioleaching Meizhou Chalcopyrite by Thermophilic Acidianus brierleyi
嗜热布氏酸菌对梅州黄铜矿的生物浸出过程特性

SHI Xian-ai,LI Cong-ying,LIN Hui,MENG Chun,GUO Yang-hao,
石贤爱
,李聪颖,林晖,孟春,郭养浩

过程工程学报 , 2005,
Abstract: 研究了嗜热布氏酸菌对梅州黄铜矿的浸出机理和浸出过程特性.嗜热布氏酸菌(简称A.B菌)在梅州黄铜矿表面的吸附过程符合朗格缪尔等温线,最大吸附量XAm5.00×108cell/g,吸附平衡常数KA5.88×10?7mL/cell.梅州黄铜矿的浸出主要是通过A.B菌的直接氧化起作用.单独A.B菌处理时,铜浸出速率为0.0137g/(L?h),是Fe3+化学氧化速率的6倍.A.B菌(65℃)对梅州黄铜矿的浸出速率是常温氧化亚铁硫杆菌(简称T.F)的16倍(31℃).A.B菌生长和浸矿的最适温度均为65℃,微生物生长最佳pH为2.0,而浸矿最适pH为1.5.A.B菌处理10d可使铜浸出率达91.3%,具有潜在的工业应用价值.
Identification and Characterization of a Novel Thermophilic Geobacillus Degrading Phenol
一株嗜热菌的分离鉴定及其苯酚降解特性

TANG Yun,LIU Mu-Zhi,LIANG Feng-Lai,FENG Lu,LIU Ru-Lin,
唐赟
,刘沐之,梁凤来,冯露,刘如林

微生物学通报 , 2006,
Abstract: 从油田地层水中分离到一株嗜热并高效降解苯酚的BF80菌株,其最适生长和降解苯酚的温度为60℃65℃。利用API50CHB/E系统和16SrDNA序列分析对菌株BF80进行了分类鉴定,该菌株的形态和生理生化特性与Geobacillusthermoglucosidasius基本相同,其16SrDNA序列与GeobacillusthermoglucosidasiusBGSCW95A1(=ATCC43742)的相似性为99·22%。在接种量为1%的条件下,该菌在20h内能完全降解3mmol/L的苯酚;在pH值5·59·0范围内能保持对苯酚良好的降解能力,并在12mmol/L苯酚的无机盐培养基中也能生长和降解苯酚,表明该菌能耐受高浓度苯酚并可用于高温含酚废水的生物处理。
PURIFICATION AND PROPERTIES OF β -GLUCOSIDASE FROM THERMOPHILIC CHEATOMIUM THERMOPHILE
一株嗜热毛壳菌β-葡萄糖苷酶的分离纯化及特性

CHU Chun-Xue,LI Duo-Chuan,GUO Run-Fang,
楚春雪
,李多川,郭润芳

菌物学报 , 2004,
Abstract: 研究了液体发酵嗜热毛壳菌Chaetomium thermophile产生的β-葡萄糖苷酶的分离纯化及特性。粗酶液经硫酸铵沉淀、DEAE-Sepharose Fast Flow阴离子层析、Phenyl-Sepharose 疏水层析、Sephacryl S-100分子筛层析等步骤后获得凝胶电泳均一的β-葡萄糖苷酶。经12.5%SDS-PAGE和凝胶过滤层析方法分别测得该酶的分子量大小约为78.4kDa和81kDa。该酶反应的最适温度和pH值分别为60℃和4.5-5.0。有较好的酸稳定性和热稳定性。金属离子对β-葡萄糖苷酶的活性影响较大, 其中Ca2+对酶有激活作用, 而Ag+、Cu2+ 、Hg2+对酶有显著的抑制作用。该酶对水杨苷具有很强的底物特异性。
Solubilization of excess sludge by thermophilic enzymes: Effect of metal ions
金属离子对剩余污泥水解嗜热菌酶促效果影响研究

ZHOU Jun,ZHENG Wei,LI Xiaoming,YANG Qi,CHEN Wei,LUO Kun,
周俊
,郑伟,李小明,杨麒,陈伟,罗琨

环境科学学报 , 2011,
Abstract: 利用从土壤中采集分离出来的嗜热菌进行嗜热酶溶解(S-TE)污泥稳定化处理,同时添加四种金属离子,考察各离子对剩余污泥的水解效果的影响。实验结果表明,在一定浓度范围内,四种金属离子都能强化污泥酶促反应,反应5 h左右时蛋白酶和淀粉酶活力达到最大值。反应过程中,200 μmol L-1 K+对蛋白酶活性的促进作用最为明显,5 h时蛋白酶活力最高可达2.20 U mL-1,相比空白样提高了48.6%,Ca2+、Mg2+和Fe2+的最佳投加浓度分别为200、100和100 μmol L-1,相应浓度下蛋白酶活力分别可达到1.91、1.92和2.14 U mL-1。K+对淀粉酶活的促进作用也最显著,且实验范围内离子浓度越高作用越明显,400 μmol L-1 K+情况下淀粉酶活力最大可达3.64 U mL-1,Ca2+在所取浓度范围内均促进淀粉酶活性,最大可提高35.0%,添加100 μmol L-1 Mg2+后淀粉酶活最大值可达2.02 U mL-1,200 μmol L-1 Fe2+能使淀粉酶活从2.42 U mL-1提高到3.22 U mL-1。SCOD在酶促反应前期增加明显,后期增长趋于平缓,添加200 μmol L-1 Mg2+后SCOD最高可达到4590.73 mg L-1,相比空白样提高了12.6%。氨氮和溶解性磷酸盐受金属离子的促进作用不明显。
Identification of a thermophilic bacterium and preliminary characterization of the secreted xylanase
一株产木聚糖酶嗜热菌的鉴定及酶学性质

CHEN Xue-Min,LIU Pei-Pei,ZHANG Bo,
陈学敏
,刘培培,张波

微生物学通报 , 2011,
Abstract: The bacterium isolated from a water sample of Yunnan tengchong hot spring can secrete a kind of thermophilic xylanase.It was identified and named as Geobacillus sp.PZH1 by morphologic observation,physio-biochemical characteristics and 16S rDNA sequence alignment.Subsequently,its secreted xylanase and the xylanase's characteristics were researched preliminarily.SDS-PAGE electrophoresis and zymogram analysis suggested that the xylanase's molecular mass was 69 kD;the optimum pH and temperature of the partially...
Isolation, identification and enzyme characterization of a thermophilic cellulolytic anaerobic bacterium
嗜热厌氧纤维素分解菌的分离、鉴定及其酶学特性

Yinping Zhao,Shichun M,Yingjie Sun,Yan Huang,Yu Deng,
赵银瓶
,马诗淳,孙颖杰,黄艳,邓宇

微生物学报 , 2012,
Abstract: Objective] To identify a thermophilic bacterium from horse manure to degrade cellulose efficiently,and to enrich microbial resources producing cellulolytic ethanol by co-culturing with thermophilic ethanol producing bacterium.Methods] We used Hungate anaerobic technique to isolate a strain named as HCp from horse manure mixed culture;its phylogeny was identified through 16S rDNA sequencing.Enzymatic assays were determined using DNS method.Results] The isolated HCp cells were straight with rods size of(0.35-0.50)μm ×(2.42-6.40)μm,in the form of single or paring.This strain belongs to a strictly anaerobic Gram-negative bacterium,it is able to form spores,shows motile ability and resistance to neomycin.The strain could degrade filter paper cellulose,cellulose powder,microcrystalline cellulose,cotton wool,rice straw and gelatin,and it was also able to utilize abundant saccharides as substrates such as cellobiose,glucose,xylose,xylan,raffinose,maltose,sorbose,fructose and galactose.The growth pH ranges from 6.5 to 8.5,temperature from 35 to 70℃ and concentration of NaCl on cellulose from 0% to 1.0%,while the optima of pH6.85,60℃ and 0.2% NaCl.Under the optimal growth conditions,the filter paper cellulose degradation rate was up to 90.40% after 10 days.The optimum temperatures for FPA,CMCase,β-glucosidase and xylanase were 70℃,70℃,70℃,and 60℃ respectively.CMCase activity was found with high thermal stability.The phylogenetic analysis based on partial 16S rDNA revealed that HCp was close to Acetivibrio cellulolyticus and A.cellulosolvens with 97.5% sequence similarities.Conclusion] Strain HCp is thermophilic,efficiently cellulolytic anaerobe.It is able to utilize vast substrates and produce highly thermostable enzymes.It is a potential bacterium that can be used for cellulolytic ethanol production.
Characterization of Thermophilic Strain SY-14 with Capability to Lyse Bacterial Cells
嗜热溶胞菌SY-14的基本特性研究

SONG Yu-dong,HU Hong-ying,XI Jin-ying,
宋玉栋
,胡洪营,席劲瑛

环境科学 , 2007,
Abstract: One spore-forming thermophilic bacterial strain SY-14, isolated from sewage sludge compost, showed significant capability to lyse bacterial cells. The strain was identified as Geobacillus sp. based on morphological characteristics and homology identification of 16S rDNA sequence. The optimal temperature and pH for growth were about 60 degrees C and pH 6.0-7.0 respectively. The culture supernatant of SY-14 showed lytic activity against both intact and thermal inactivated bacterial cells, and the cell lysis percentages at 6 hours were 70% and 85% respectively. The lytic activity of the culture supernatant decreased significantly after heat treatment, which inferred the lytic activity mainly derived from extracellular lytic enzymes of SY-14. The lytic activity of the culture supernatants of SY-14 increased significantly during the log phase in the batch culture process, and then decreased quickly after the maximum activity was reached. The culture supernatant of SY-14 showed lytic activity against all the five tested Gram-negative strains and some tested Gram-positive strains.
Agrobacterium tumefaciens-mediated transformation as a tool for insertional mutagenesis in thermophilic fungus Thermomyces lanuginosus
农杆菌介导的获取疏绵状嗜热丝孢菌插入突变体体系的建立

Hua Han,Xiaoxue Xu,Yanjie Peng,Dehui Kong,Duochuan Li,
韩华
,徐晓雪,彭延杰,孔德慧,李多川

微生物学报 , 2012,
Abstract: 摘要:【目的】建立疏绵状嗜热丝孢菌的稳定遗传转化体系并获得插入突变体。【方法】利用农杆菌介导的方法建立疏绵状嗜热丝孢菌的遗传转化体系;分别通过Southern杂交、克隆转移DNA(T-DNA)侧翼序列来确定T-DNA 在疏绵状嗜热丝孢菌基因组中的拷贝数和插入位点。【结果】成功建立了可靠的疏绵状嗜热丝孢菌的遗传转化体系。共培养过程中使用萌发孢子是成功建立疏绵状嗜热丝孢菌遗传转化体系的必要条件。疏绵状嗜热丝孢菌萌发的孢子与农杆菌在28℃共培养48h时,转化效率最高。乙酰丁香酮(AS)在农杆菌预培养及疏绵状嗜热丝孢菌萌发的孢子与农杆菌的共培养阶段都是必需的,且在共培养阶段当AS浓度为500 μM时转化效率最高。Southern 杂交验证表明,79.2%的转化子为T-DNA单拷贝插入,且通过热不对称PCR(TAIL-PCR)分析得出T-DNA在该菌基因组中的插入位点是随机的。通过该转化系统筛选到部分表型突变体。【结论】我们首次报道了利用ATMT技术成功转化嗜热真菌-疏绵状嗜热丝孢菌,证明了该方法是一种简单有效的获得插入突变体的方法,并为该嗜热真菌进行基因定位提供了工具。
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