Bacterial Diversity And The Metagenome Library Construction Of The Rumen From Bactrian Camel(camelus Bactrianus)

Rumen microorganisms played an important role in ruminant digestion and absorption of nutrients,had great potential applications in the field of rumen adjusting,food fermentation,biomass utilization etc.In order to investigate the composition of microorganisms in the rumen of bactrian camel(Camelus Bactrianus),this paper studied the microbial diversity by culture-independent approach.Metagenomic library construction and function of valuable genes selected provided the scientific basis and technology platform for the further development and utilization of rumen microorganisms.This study achieved the following results:The method of extracted high-quality total DNA from rumen microbes was established. This research had adapted the direct extraction method with freeze thawing-enzyme-SDS combination,which can extract more complete rumen microbial DNA.The total DNA fragment was larger than 23 kb.Rumen bacteria 16S rDNA gene clone library was constructed.The library's coverage was up to 57% and Shannon-Wiener index (H ') was 4.57.ARDRA technique was used to analyze the 331 clones in this library,and 130 operational taxonomic units(OTUs) were obtained.Firmicutes,Bacteroidetes,Proteobacteria, Verrucomicrobia,TM7 and unclassified bacteria were existed in the rumen.Firmicutes was the dominant bacteria,which was up to 63.8%.There were Proteobacteria,Firmicutes, Actinobacteria,Bacteroidetes,Tenericutes, Chloroflexi and Planctomycetes in the Bactrian camel rumen by using T-RFLP technical analysis.Above results were showed that combining with these two methods,we can detect more abundant types of bacterias.Analysis results showed that the similarity of Bactrian camel rumen bacteria to known sequence in Genebank was between 85% -99%,sequences with similarity above 97% was only 35.4%,unknown sequences account for 64.6%,indicating that only a small number of species were known in Bactrian camel rumen bacteria,majority were unknown new genus,species or potentially other new taxon;Among the known species,cultivable bacterial account for only 5.4%,the rest 94.6% were uncultivable.Plasmid metagenomic library was constructed with the total rumen microbial DNA fragments and pBluescriptSK(+) vector.424 clones in the library were screened with CMCase andβ-glucosidase activity,and 30 clones were showedβ-glucosidase activity and 6 clones showed CMCase activity,these clones have to be further analyzed and identified.