本文選題：大腸桿菌Nissle + 分離鑒定��； 參考：《微生物學通報》2017年04期

【摘要】：【目的】證實大腸桿菌Nissle 1917作為自然菌株存在于動物豬體內(nèi),并能從豬糞便中分離。建立大腸桿菌Nissle 1917的原位雜交鑒定方法。【方法】采集135份健康斷奶仔豬的新鮮糞便制備DNA模板,以人源大腸桿菌Nissle 1917為陽性對照菌株,分別針對Nissle 1917的I型菌毛亞單位Fim A、F1C菌毛亞單位Foc A及兩個質(zhì)粒pMUT1和pMUT2的相關基因序列設計5對特異性引物進行PCR擴增;并將其中427 bp大小的質(zhì)粒片段pMUT2(a)作為目的片段回收純化,用地高辛隨機引物標記法制成DNA探針�！窘Y(jié)果】從其中的2份DNA模板中擴增出上述5對特異性引物PCR預期大小相符的片段,初步認為大腸桿菌Nissle 1917可能存在于豬體內(nèi)。應用制備的探針通過菌落原位雜交的方法從2份陽性糞便樣品中篩選出2株陽性菌落,通過血清學檢驗、PCR擴增和測序進一步鑒定為陽性Nissle 1917菌株�！窘Y(jié)論】動物源益生菌Nissle 1917的分離鑒定,為優(yōu)良動物源益生菌研究和應用奠定了基礎。
[Abstract]:[objective] to confirm that Escherichia coli Nissle 1917 exists as a natural strain in animal pigs and can be isolated from pig feces. An in situ hybridization method for identification of Escherichia coli Nissle 1917 was established. [methods] DNA template was prepared from fresh feces of 135 healthy weaned piglets. Nissle 1917 was used as positive control strain. Five pairs of specific primers were designed for PCR amplification of Nissle 1917 type I pili subunit Foc A and two plasmids pMUT1 and pMUT2 related genes. The plasmid fragment of 427 BP in size was recovered and purified as the target fragment. Digoxigenin random primer labeling method was used to make DNA probe. [results] five pairs of specific primer PCR fragments were amplified from the two DNA templates. It was suggested that E. coli Nissle 1917 might exist in pigs. Two positive colonies were screened from 2 fecal samples by colony in situ hybridization. The positive strain of Nissle 1917 was further identified by serological analysis and sequencing. [conclusion] the isolation and identification of probiotic strain Nissle 1917 from animal sources laid a foundation for the study and application of excellent probiotics from animals.
【作者單位】： 揚州大學獸醫(yī)學院;江蘇省動物重要疫病與人獸共患病防控協(xié)同創(chuàng)新中心;國藥集團揚州威克生物工程有限公司;
【基金】：國家自然科學基金項目(No.31672579,30571374,30771603,31072136,31270171) 江蘇高校優(yōu)勢學科建設工程資助項目 科技部轉(zhuǎn)基因生物新品種培育重大專項(No.2014ZX08006-001B)~~
【分類號】：S852.61
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本文編號：1810366