Isolation And Identification Of Poephogus Grunniens Lactic Acid Bacteria And Analysis Of Its 16S RDNA-SSCP

In this research,71 strains lactic acid bacteria were isolated and identified which from poephogus grunniens stool and intestinal tract content (93 samples from 7 areas). They are Lactobacillus brevis(5 strains), Lactobacillus coprophilus(3 strains), intestines Bacterium lacticum (8 strains), Lactobacillus acidophilus (8 strains), Lactobbacillus bulgaricus(3 strains), Lactobacillus curvatus (2 strains), Lactobacillus fermenti (11 strains) , Lactobacillus casei(4 strains), Lactobacillus lactis (6 strains), Lactobacillus plantarum(9 strains),Streptococcus thermophilus (3 strains), lactic acid galactococcus (2 strains), Ent-rococcus faecalis (3 strains), urina enterococcus(2 strains), hai enterococcus (2 strains), which based on biochemical test and growing test in different T, pH,and NaCl (%) .they are 59 Bacillus acidi lactici,12 coccobacteria,15 species totally.We choose two strains from every species and made it into thalline suspension after 36h cultivation.Thalline suspension were ajusted to 2.0, 3.0, 4.0 and put into 37℃ for 5h then colony count were done after being cultivated in 37'C for 24h.As a result Q₁, Q₆, Q₃, Q₄, Q₇, Q₈, ZH₁,70, 69, 59,23, 54, N₃, N₄, 10, Z₅, 1, 3, 19 can survive more than 10% in pH 2,40% in pH 3 and more than 85% in pH 4. Q₁, Q₆, Q₃, Q₄,Q₉, Q₁₀, ZH₁, 70, 69, 59, N₁,N₅, N₃, N₄,10, Z₅,1, 3,19 can all survive at 0.6% Chleolate peptone water atter its 36h cultivation were detected in OD₆₆₀ by spectro photometer.Bacteriostasis collar about 13mm-30mm mostly at 20mm-27mm,were show in inhibition to pathogens of lactobacillus culture fluid test, inhibition to pathogens of lactobacillus matebolite test, inhibition activity of matebolite after treatment with heat test, inhibition activity of matebolite after treatment with acid or alkali test, inhibition activity of matebolite after enzyme treating test. All the inhibition test was tested by monolayer agar plate diffusion method.470bp, which from variable region 371 to 838 of 16SrDNA of 10 species were amplified by using general primer of lactobacillus.This primer was design by using primer software after comparing with different 16SrDNA in GenBank. Ten different strap were detected in this gene by using SSCP.We sequenced 5 species from this 10 and processed it by using DNAStar. High homology was Exist between these five species from 87.3% to 95.6%. Very high homology was found after comparing with standard sequence from GeneBank ,that is :Q₁, 98.51%; Q₇,98.94%; ZH₁, 98.3%; 19,98.94%; N₃,98.73%(more than 98 %) which shows this 5 species are Streptococcus thermophilus, Enterococcus faecalis, Lactobacillus brevis,...