富集培养的低温纤维素降解细菌菌群结构分析  

Analysis of Cold-adapted Cellulose-degrading Bacterial Community Structures Under Enrichment Culturing Conditions

作　　者：孟建宇[1] 樊兆阳[1] 李蘅[1] 陶羽[1] 王瑞刚[1] 冯福应[1] 

机构地区：[1]内蒙古农业大学生命科学学院应用与环境微生物研究所,呼和浩特010018

出　　处：《农业生物技术学报》2014年第5期607-614,共8页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金项目(No.30860015);内蒙古自治区自然科学基金项目(No.2009ZD03和No.2010BS0506);内蒙古自治区高等学校科学研究项目(No.NJ10054);内蒙古农业大学创新团队项目(No.NDPYTD2010-3);内蒙古自治区高等学校青年科技英才支持计划

摘　　要：内蒙古大兴安岭森林土壤中低温纤维素降解菌资源丰富,本研究旨在分析其菌群结构。在10℃下,用羧甲基纤维素钠(CMC-Na)培养基对样品中的低温纤维素降解菌进行富集传代培养,用3,5-二硝基水杨酸(DNS)法测定纤维素酶活性,以细菌16S rDNA通用引物(34lF和907R)进行V3～V5区域PCR扩增,以变性梯度凝胶电泳(DGGE)对PCR产物进行电泳,获得16S rDNA指纹图谱后分析细菌菌群结构。研究结果表明,低温富集传代过程中,相对酶活随着培养代数的增加而不断增加,第4代的相对酶活最高,为30.625 IU;每代都有菌种的消失和增加;富集培养液中包含属于α-变形菌纲(α-Proteobacteria)、β-变形菌纲(β-Proteobacteria)、γ-变形菌纲(γ-Proteobacteria)、硬壁菌门(Firmicutes)和拟杆菌门(Bacteroidetes)类群的细菌,其中硬壁菌门类群(65.5%)和β-变形菌纲类群(13.8%)的细菌为优势菌。本研究证实,内蒙古大兴安岭森林土壤有丰富的低温纤维素降解菌资源,有潜在的应用前景。There are abundant cold-adapted cellulose-degrading bacteria resources from soil in the Da Hinggan forest of Inner Mongolia. This study was aimed at analyzing the community structure of cold-adapted cellulose degrading bacteria. Cold-adapted cellulose-degrading bacteria were isolated by enrichment culture using sodium salt of caboxy methyl cellulose（CMC-Na） medium at 10 ℃. Cellulase activity was determined by 3, 5-dinitrosalicylic acid （DNS） method. The 16S rDNA genes （V3-V5 region） were amplified by using the universal primers （34lF and 907R）. The PCR products were electrophoresed by denaturing gradient gel electrophoresis （DGGE）. The 16S rDNA fingerprint was obtained and the bacterial community structures were analyzed. The results indicated that the relative enzyme activity increased constantly with the extension of incubation time during the process of enrichment, the 4th generation had the highest relative enzyme activity with 30.625 IU. Some bacteria died out or increased at each generation. Four DGGE profiles of microbial community structure of four generation enrichment cultures were obtained. There were five subgroups in enrichment culture, including α-Proteobacteria, β-Proteobacteria, γ-Proteobacteria, Firmicutes, and Bacteroidetes, respectively. The subgroups of Firmicutes （65.5%） and β-Proteobacteria （13.8%） were the dominant bacterial floras. This study confirmed that there were abundant resources of cold-adapted cellulose-degrading bacteria in the Da Hinggan forest of Inner Mongolia. These bacteria will have a potential value of application.

关 键 词：低温纤维素降解菌 富集培养 群落结构 变性梯度凝胶电泳(DGGE) 

分 类 号：Q503[生物学—生物化学]