Researching Of The Diversity Of Intestinal Flora Of Adult Ailuropoda Based On 16S RDNA-RFLP

As a endangered mammal, ailuropoda belongs to ursidae. Ailuropoda is on the verge of extinction because of disease; especially intestinal disease. Normal floras in the intestinal canal are benefical to organism. Their quantity and proportion always holds the dynamic balance in usual. They play a part in the physiological activity such as nutrition digestion, immunity, antitoxin, anticancer. It is dangerous to the health of ailuropoda, if there are intestinal diseases caused by the changes of quantity and construction of the normal floras.For exploring the intestinal flora construction of adult ailuropoda in autumn, this experiment researched autumnal intestinal floras diversity of 3 adult ai luropoda in Sichuan Yaan Bifengxia base, China Conservation and Research Centre for the ailuropoda (total 5 feces:TT9, TT10, TT11, XM10, YM10) by 16S rDNA-RFLP. At first, amplify the total 16S rDNA by general primer, construct 16S rDNA library. Then, cluster analysis the RFLP by MspⅠand HinfⅠ, analyse the clustering results by measure of a diversity.At last, analyse the select dominated flora by sequencing,construct cladogram. Finding indicated that:(1) There are 57 OTUs in TT9,48 OTUs in TT10,30 OTUs in TT11,35 OTUs in XM10,52 OTUs in YM10, after constructing clone library and RFLP clustering. The coverages of the library are 58.33%,67.31%,80%,71.88%, 68.52%. This indicated the good representation of the library.(2) By analyzing the RFLP results according to the measure of a diversity, the indexs,dMa,R2,JSW,H',Hmax,Dg,Schaol) show that the diversity of intestinal bacterium of ailuropoda on September is higher than November. In the adult ailuropoda, the diversity of intestinal bacterium in the youth is higher than older. There are some impact to the diversity of intestinal flora by living environment of ailuropoda.(3) After sequencing of select dominated flora, blasting on the NCBI, and constructing the cladogram by DNAstar, the results showed that dominated floras included Proteobacteria,Firmicutes and Bacteroidetes. The main of Proteobacteria are Escherichia,Pseudomonas,Shigella;the main of Firmicutes is Streptococcus and the Bacteroidetes'is Sphingobacterium.(4) Pseudomonas (36.11% of total clones) and Escherichia(7.4% of total clones) are the main flora in TT9. Escherichia (12.5% of total clones) is the main flora in TT10, Escherichia (21.0% of total clones),Shigella (21.0% of total clones) and Streptococcus (12.4% of total clones) are the main flora in TT11. Shigella (45.8% of total clones) is the main flora in XM10. Escherichia (13.9% of total clones), Pseudomonas (8.3% of total clones) s the main flora in YM10.