低温地区纤维素降解菌的筛选鉴定与纤维素酶活力测定

Screening and Identification of Cellulose Degrading Bacteria and Determination of Cellulase Activity in Low-temperature Areas

【目的】筛选低温地区纤维素降解菌,为畜禽粪便无害化处理提供理论依据。【方法】通过适温性测定与刚果红染色法,对吉林低温地区冻土和牛粪中初步分离并筛选低温、纤维素降解菌;通过16S rDNA测序对菌株进行分子生物学鉴定;通过葡萄糖标准曲线的绘制测定纤维素酶的活性。【结果】在牛粪与冻土中通过适温性测定筛选出3株低温菌株(D-3、D-7和D-16),通过刚果红染色法与酶活性测定筛选出5株纤维素酶活性较高的菌株(X-1、X-5、X-9、X-11和X-18),均属于细菌,经过16S rDNA测序鉴定,将测序结果利用BLAST软件进行同源性比对,并建立基因系统发育树得知,3株低温细菌分别为弗雷德里克斯堡假单胞菌、醋酸钙不动杆菌和产黄假单胞菌,5株纤维素降解菌株分别为蜡样芽孢杆菌、枯草芽孢杆菌、解淀粉芽孢杆菌、短小芽孢杆菌和沙福芽孢杆菌。【结论】吉林低温地区可培养纤维素降解菌的类群组成及纤维素降解能力。筛选出的5株活性较高的纤维素降解菌株(X-1、X-5、X-9、X-11和X-18)可作为有效降解纤维素的潜力菌株应用于粪污堆肥发酵剂的研制。

【Objective】The cellulose degrading bacteria in low-temperature areas were screened to provide the theoretical basis for the harmless treatment of livestock manure.【Method】The low-temperature and cellulose degrading bacteria were preliminary isolated and screened from the frozen soil and cow dung in Jilin low-temperature areas by temperature suitability test and Congo red staining method;The molecular biological identification of the strain was carried out through 16S rDNA sequencing;The cellulase activity was determined by plotting the glucose standard curve.【Result】Three low-temperature strains(D-3,D-7 and D-16)were screened from cow dung and frozen soil by temperature suitability test,and five strains with high cellulase activity(X-1,X-5,X-9,X-11 and X-18)were screened by Congo red staining and enzyme activity test,which all belonged to bacteria.After 16S rDNA sequencing and identification,the homology alignment of sequencing results was carried out with BLAST software,then based on the established gene phylogenetic tree,the three low-temperature bacteria were identified as Pseudomonas frederiksbergensis,Acinetobacter calcoaceticus and Pseudomonas synxantha,and five cellulose degrading strains were identified as Bacillus cereus,Bacillus subtilis,Bacillus amyloliquefaciens,Bacillus pumilus and Bacillus safensis.【Conclusion】The group composition and cellulose degradation ability of cellulose degrading bacteria can be cultivated in Jilin low-temperature areas.The obtained five cellulose degrading strains with high activity(X-1,X-5,X-9,X-11 and X-18)can be used as potential strains for effectively degrading cellulose,which applied in the development of manure compost fermentation agent.