瘤胃细菌GH48家族糖苷水解酶基因多样性

Gene diversity of the bacterial 48 family glycoside hydrolase (GH48) in rumen environment

【目的】了解瘤胃细菌第48家族糖苷水解酶基因(GH48)多样性,为木质纤维素高效降解提供新的基因资源。【方法】通过基因序列比对,设计gh48的简并引物;同时提取两个瘤胃样品的总DNA和总RNA,并将总RNA逆转录成cDNA。以总DNA和cDNA为模板,通过PCR扩增分别建立克隆文库并对克隆文库进行测序;对所得序列进行out种类划分和聚类分析。【结果】本研究共得到了455条编码GH48家族蛋白的基因序列,核苷酸序列之间的相似性为58.65%-100%。对序列的进一步分析表明,89%可以作为区分其种类的界定标准。以此为依据确定所得到的基因序列分别编码66种不同的GH48家族蛋白,分别聚为5个相对独立的类群,其中新类群C中OTU65所代表的序列是cDNA克隆文库中的优势序列,分别占两个cDNA克隆文库的36.4%和19.5%。我们的结果揭示瘤胃细菌gh48基因具有丰富的多样性,同时,其中也存在优势表达的GH48家族蛋白。

[Objective]The gene diversity of the bacterial 48 family glycoside hydrolase（GH48） in rumen environment was studied and new gene resources for efficient cellulose degradation were provided.[Methods]A pair of gh48 degenerate primers was designed through sequences alignment of the gh48 gene sequences from ruminal Ruminococcus.The total DNA and RNA were extracted from two rumen samples and cDNA was synthetized through reverse transcription from total RNA. Four gh48 gene clone libraries were constructed and analyzed.[Results]In total 455 gh48 gene sequences were obtained from the 4clone libraries. Sequence similarity among the 455 gene sequences varies between 58.65% and 100%.They fell into 66 species with the sequence similarity ≥89%,and divided into 5 different clusters.OTU65 in cluster Crepresents an abundant gh48 gene which in both DNA and cDNA clones libraries,accounting for 36.4% and 19.5% respectively.Our studyrevealsrich gene diversity of the 48 family glycoside hydrolase and provided new gene resources for cellulose degradation.