一株高产琼胶酶海洋细菌的筛选与鉴定  

Screening and Identification of an Effective Agarase-producing Marine Bacterium

作　　者：周钦茂 谢燕纯 陈彦梅 张扬 柯德森[1] 陈琼华[1] ZHOU Qin-mao;XIE Yan-chun;CHEN Yan-mei;ZHANG Yang;KE De-sen;CHEN Qiong-hua(College of Life Science,Guangzhou University,Guangzhou 510000)

机构地区：[1]广州大学生命科学学院,广州510000

出　　处：《生物技术通报》2018年第12期140-146,共7页Biotechnology Bulletin

基　　金：广州市科技计划项目(201510010108);2017国家级大学生创新训练项目(201711078003);第十六届全国"挑战杯"竞赛立项(N-J-17-02)

摘　　要：从广东省南澳岛采集龙须菜(Gracilaria lemaneiformis),分离海洋来源的琼胶酶产生菌,并对其进行分类鉴定,为琼胶酶的开发利用奠定基础。利用4种不同的筛选培养基分离产琼胶酶的菌株,通过形态、生理生化特征和16S rRNA基因序列分析对菌株进行鉴定并构建系统发育树,通过DNS法测定琼胶酶活力,研究菌株所产琼胶酶的类型,对菌株的生长曲线及发酵产酶曲线进行初步测定。结果显示,分离得到一株高产琼胶酶的菌株ZQM2017,该菌株为革兰氏阴性短杆菌,16S rRNA基因序列分析显示该菌株属于弧菌属(Vibrio sp.),结合形态特征和生理生化实验结果鉴定为溶藻弧菌(Vibrio alginolyticus);可同时产α-琼胶酶与β-琼胶酶;该菌株在液体培养基中28℃,180 r/min振荡培养时,其对数期出现在3-9 h,发酵5 h即有明显产酶,当发酵至46 h,所产琼胶酶活力达到最高109.87 U/mL发酵液。从南澳岛龙须菜上自主分离筛选得到的海洋细菌ZQM2017,经鉴定命名为Vibrio alginolyticus ZQM2017,可同时分泌α-琼胶酶和β-琼胶酶,所产琼胶酶初始活力高达109.87 U/mL。This work is to isolate,screen and identify the effective agarase-producing strain from Gracilaria lemaneiformis collected from Nan'ao Island,Guangdong province,for building a foundation for the utilization of agarase.Four different screening culture-mediums were used to screen the agarase-producing bacterium,the morphology,physiological and biochemical properties were employed to identify the isolated strain,and phylogenetic tree was constructed.DNS was adapted to determine the enzyme activity of the agarase,to study the type of the agarase from the strain,and to draw its growth curve and fermentation curve.As results,an effective agarase-producing strain ZQM2017was isolated from the samples of G.lemaneiformis,which was a gram-negative bacterium,its16S rRNA sequence was99%homologous to52strains of Vibrio species in the database of NCBI.Combining with the morphological and physiological-biochemical characteristics,it was identified the strain ZQM2017as Vibrio alginolyticus.The type of agarase by V.alginolyticus ZQM2017was bothα-agarase andβ-agarase.Under the condition of180r/min and28℃,the logarithmic phase of the strain was between3h to9h and the enzyme obviously produced at5h,and the enzyme activity of agarase from V.alginolyticus ZQM2017reached109.87U/mL after fermenting for46h.In conclusion,the marine bacterium ZQM2017isolated and screened independently from Nan'ao Island was identified as V.alginolyticus ZQM2017,and can secretα-agarase as well asβ-agarase with the initial enzyme activity of109.87U/mL.

关 键 词：琼胶酶 溶藻弧菌 筛选与鉴定 16SrRNA 

分 类 号：Q93[生物学—微生物学]