Analyze The Intestinal Flora Of Adult And Elderly Panda By16S RDNA-RFLP Technology

The giant Panda is rare and endangered animal sendemic to China,and it regarded as "National Treasure" and a "living fossil", now the giant Panda has been listed as national grade I protected wild animals and also has been listed as the species of 《Convention on International Trade in Endangered Species of Wild Fauna and Flora)) Appendix I.There are many reasons endangered giant panda, but diseases, especially intestinal diseases occupied a large proportion.The giant Panda’sintestinal flora in normal circumstances to keep the balance in the number and proportion, they play an important role inmaintaining the balance and stability of the intestinal micro ecology systems.Intestinal florahave a significant impact to digestion, immune and other physiological activities of giant Panda.The research about giant Panda’s intestinal flora is still less, in order to explore the similarities and differences in intestinal flora of the adult giant Panda and elderly giant Panda,also in order to provide the basis for comprehensive analysis and comparison the similarities and differences in intestinal flora of all ages giant Panda.In this paper, using16S rDNA-RFLP technology to study the diversity of intestinal flora of the adult and elderly giant Pandawhich be in captivity.Construction of16S rDNA clone library of intestinal flora of the adult and elderly giant Panda;usingMsp I and HinfI this two restriction endonuclease to clustering analysis RFLP (Restriction Fragment Length Polymorphisma);then,measure and analyse the diversity of the clustering results;finally, analyse the resulting flora’ sequence.The results show that:.1. Construction of16S rDNA clone library of intestinal flora of the adult and elderly giant Panda,after RFLP clustering, getting48OTUs from adultgiant Panda and41OTUs from elderly giant Panda.The coverage (Coverage C) of the two clone libraries, are85.4%and87.3%. The capacity values reached more than50%,explain that the picked positive clones had a very good representative, the clone library can truly reflect the overall distribution of the intestinal floraof adult and elderly giant Panda.2. Diversity measurements and analysis results, Shannon Index, H’, Simpson dominance index (D), species richness index Margalef, diversity index of Chaol (S) have showned that the diversity of intestinal flora of the adult and elderly giant Panda were relatively high. But the diversity and abundance of intestinal flora of adult giant Panda is higher than elderly giant Panda.This indicates that the age of the giant Panda have effects on the diversity of the intestinal flora.3. Determination of intestinal flora’ sequences of the adult and elderly giant Panda, learned after BLAST on NCBI, the intestinal flora of adult and elderly giant Panda are both mainly attributable to two bacteria:Proteobacteria and Firmicutes.Among them, the mainly in Proteobacteria of adult giant Panda are Escherichia coli (30.71%), followed by Pseudomonas sp.(8.57%), Shigella (5.00%), Aggregatibacter (3.57%), Oceanisphaera (1.43%), Aranicola sp.(0.71%), Comamonas sp.(0.71%); the mainly in Firmicutes are Streptococcus (32.14%), followed by Clostridium (1.43%), Turicibacter (1.43%), Weissella (1.43%); of which there are12.81%of bacteria is uncultured bacteria. The mainly in Proteobacteria of intestinal flora of elderly giant Panda were Escherichia coli (37.82%), followed by Pseudomonas sp.(7.69%), Shigella (1.28%), Oceanisphaera (0.64%); the mainly in Firmicutes were Streptococcus (23.72%), followed by Weissella (16.67%), Clostridium (0.64%); of which there are11.54%of bacteria is uncultured bacteria.4. The dominant flora in the intestinal flora of adult and elderlygiant Pandais obvious.The major dominant flora both are Escherichia coliand Streptococcus,But the proportion of Escherichia coliin the intestinal flora of adult giant Panda is lower than Streptococcus, then the proportion of Escherichia coliin the intestinal flora of elderly giant Pandais higher than the Streptococcus, Escherichia coliis the most dominant bacteria in the intestinal flora of the elderly giant Panda. TheWeissella also accounted for a large proportion,and becomed the third dominant florain the intestinal flora of the elderlygiant Panda.