Analysis Of Metagenome From Communities Of Microorganisms In The Cow Rumen

In ecosphere, 60% of total life-form are microorganisms which contain an extensive suite of gene resources. Thus, there has been an increasing worldwide interest in all kinds of environmental microbial communities. Such as: soil,sea,lake and so on. Ruminal microbial communities are composed of various distinct groups mostly specialized in polysaccharide hydrolysis and fermentations of straws, which supply ruminants their nutrient needs for energy, protein and vitamin. Several decads years ago ,the study of rumen had began .The knowledge of rumen was still limited because of cultivation techniques. To circumvent difficultes and limitations associated with cultivation techniques, the culture independented technology that bypass the cultivation of microorganisms make it possible to take the good advantage of the gene resource. Firstly, the mixed genomic DNA were extracted and purified from rumen environmental samples directly in this study. The DNA yields that have been got ranged from 0.1～0.3μg per ml of liquid environmental sample. Secondly,culture independent method was used to study the diversity of rumen bacteria. The total DNA directly extracted from rumen fluid was used as templet.Molecular diversity of rumen bacteria was analyzed by PCR amplification and sequencing of 16S rDNA clone libraries prepared from the DNA of rumen content. Random 45 clones, containing almost full size 16S rDNA sequences (1.5 kb), were completely sequenced and subjected to an on line similarity search. The bacterial community structure was also revealed by phylogenetic tree of known sequences. Finally, the mixed genomic DNA library was constructed by insert in restriction fragments of the purified DNA in to plasmids pSK(+)and then transferring into DH10B after the genomic DNA were digested with the restrictions enzyme EcoRⅠpartly .The library contained 1.9×105 positive clones and the average foreign fragment DNA was about 4.2 kb. And the efficiency of mixed genomic DNA was the approximately 1000 inserting –containing clones which contained the fragment arranged from 3 kb to 10kb were obtained from 1ml samples.Clone libraries were analyzed by DNA sequencing and comparing with gene bank .Among 750 randomly-selected clones contained inserting fragments which may confer some enzymes and protein. Such as: lactase,catalase ,beta-glucosidase,ATP-dependent protease,beta-glucosidase and so on. Keywords:rumen microbe, the extraction of mixed genomic DNA, diversity analyse plasmid...