本文選題：益生菌　切入點：16S　出處：《第三軍醫(yī)大學》2006年碩士論文　論文類型：學位論文

【摘要】： 益生菌制劑產品菌種與活菌數含量是其產品質量的關鍵。國外益生菌產品的微生物分析表明,部分產品的微生物種屬和活菌數含量與產品標簽上標注的種屬和數量不符。然而,國內還沒有這方面的研究報道。本課題對國內主要的五種醫(yī)用益生菌制劑產品進行菌種分析�？疾炝宋宸N醫(yī)用益生菌制劑多個批次產品的活菌數含量,有統(tǒng)計意義的反映產品在有效期內的活菌數含量情況。通過結合表型特征分析法與16S rDNA序列分析法對從產品中分離的10株菌進行種屬鑒定。同時將PCR-DGGE分析方法應用到鑒定益生菌制劑產品菌株組成,為益生菌制劑產品的質量控制提供快速、可靠、重復性好的分析方法。 方法與結果:(1)通過篩選獲得到適合的選擇培養(yǎng)基,并用于5種醫(yī)用益生菌制劑25批產品的活菌計數,計數結果顯示:產品Ⅰ中的三種菌的活菌數變化范圍分別為10~5～10~(10)CFU/粒、10~5～10~8CFU/粒、10~6～10~8 CFU/粒,產品Ⅱ的活菌數變化范圍為10~8～10~9 CFU/粒,產品Ⅲ的活菌數變化范圍為10~8～10~9 CFU/粒,產品Ⅳ中的兩種菌的活菌數變化范圍分別為10~7～10~8 CFU/粒和10~8～10~(10) CFU/粒,產品Ⅴ中的三種菌的活菌數分別為10~7 CFU/粒、10~6 CFU/粒、10~6 CFU/粒。(2)分離純化10株產品菌株,提取單菌株基因組DNA,PCR擴增16S rRNA基因,純化并測定PCR產物序列,將測得序列與GenBank數據庫的序列進行同源性分析,并建立系統(tǒng)進化樹。在GenBank數據庫中登陸其10株菌的16S rDNA序列,獲得如下序列號:PFK1(DQ295034),PFK2(DQ295035),PFK3(DQ295036),LZCL(DQ295040),ZCS1(DQ295037),MMA1(DQ295041),MMA2(DQ295038),JSQ1(DQ295042),JSQ2(DQ295039),JSQ3(DQ295043)�；诰�16S rDNA序列的Blast和系統(tǒng)進化樹分析結果表明產品Ⅰ中含有的三種菌分別為長雙歧桿菌、植物乳桿菌和糞腸球菌;產品Ⅱ中含有的菌是動物雙歧桿菌;產品Ⅲ中含有的菌是地衣芽孢桿菌;產品Ⅳ中含有的兩種菌是屎腸球菌和枯草芽孢桿菌;產品Ⅴ中含有的三種菌分別為動物雙歧桿菌、保加利亞乳桿菌、嗜熱鏈球菌。(3)將分離菌株進行形態(tài)、生理和糖發(fā)酵實驗,結果表明這些表型鑒定的結論與16 S rRNA基因序列同源性分析結論一致,分離純化5種產品的10株菌中仍有相同的3株菌與標簽標注的不一致,即其中標簽上分別標注為青春雙歧桿菌和長雙歧桿菌的兩株菌鑒定結果均為動物雙歧桿菌,
[Abstract]:The microbiological analysis of probiotic products in foreign countries shows that the microbial species and the number of live bacteria in some products are not consistent with the species and quantity labeled on the product label. There is no such research report in China. In this paper, the main five kinds of medical probiotics products in China were analyzed, and the number of live bacteria in five kinds of medical probiotics products were investigated. It is statistically significant to reflect the content of live bacteria in the valid period of product. By combining phenotypic characteristic analysis and 16s rDNA sequence analysis, the species of 10 strains isolated from the product were identified. Meanwhile, the method of PCR-DGGE analysis was applied to identify the species of 10 strains isolated from the product. Used to identify the composition of probiotic product strains, To provide a rapid, reliable and reproducible analytical method for the quality control of probiotics. Methods and results: a suitable medium was obtained by screening and used to count live bacteria in 25 batches of 5 kinds of medical probiotics. The counting results showed that the number of live bacteria in product I ranged from 10 ~ 5 ~ 10 ~ (10) to 10 ~ (10) ~ 10 ~ (10) ~ (10) ~ (10) ~ (10) ~ (10) C ~ (10) C ~ (8) ~ (10) C ~ (10) C ~ (10) ~ (8) CFU/, to 10 ~ (8) ~ 10 ~ (9) CFU/ for product 鈪，

本文編號：1607690