Microbiology and Biotechnology Letters (한국미생물·생명공학회지)

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Evaluation of the Colonization of Lactobacillus plantarum in Mouse Gut by Terminal Restriction Fragment Length Polymorphism Analysis

Terminal Restriction Fragment Length Polymorphism 분석을 이용한 Lactobacillus plantarum의 생쥐 장관 정착 평가

  • Jung, Gwangsick (Department of Food Science and Biotechnology, Kyonggi University) ;
  • Lee, Jong-Hoon (Department of Food Science and Biotechnology, Kyonggi University)
  • 정광식 (경기대학교 식품생물공학과) ;
  • 이종훈 (경기대학교 식품생물공학과)
  • Received : 2012.08.20
  • Accepted : 2012.10.07
  • Published : 2012.12.28
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Abstract

T-RFLP (terminal restriction fragment length polymorphism) analysis, one of the most highly adopted culture-independent microbial community analysis methods, was applied to evaluate the colonization of probiotics in experimental animal gut. Lactic acid bacteria that exhibited cinnamoyl esterase activity were isolated from Korean fermented vegetables and identified by 16S ribosomal RNA sequence analysis. Lactobacillus plantarum KK3, which demonstrated high chlorogenic acid hydrolysis by cinnamoyl esterase activity, and acid/bile salt resistances, was cultured, freeze-dried, and fed to mice and the microbiota in their feces were monitored by T-RFLP analysis. The T-RF of L. plantarum was detected in the feces of mice after the start of administration and lasted at least 31 days after the initial 7 day feeding. T-RFLP analysis was considered a useful tool to evaluate the gut colonization of probiotic L. plantarum. In order to prove that L. plantarum was from viable cells, we reisolated L. plantarum in the feces using cinnamoyl esterase activity media as the screening step. The colonization of L. plantarum KK3 in the mouse gut was confirmed by this research.

배양 비의존적 미생물 군집분석법의 하나로 활발하게 이용되고 있는 T-RFLP (terminal restriction fragment length polymorphism) 분석과 실험동물을 이용한 생균제의 장관 정착 평가를 시도해 보았다. 장관 정착 평가를 위한 생균제는 cinnamoyl esterase 활성 보유 유산균을 발효 침채류로부터 분리하였고, 16S ribosomal RNA 유전자 염기서열 분석을 통해 동정하였다. 분리 균주 중, cinnamoyl esterase에 의한 chlorogenic acid 분해활성이 높고, 내산성 및 담즙산 내성의 평가 결과가 우수한 Lactobacillus plantarum KK3 균주를 생균제로 선정하였다. 배양 후, 냉동건조하여 제조한 생균제를 생쥐에게 투여한 다음, T-RFLP 분석을 이용하여 생쥐 분변의 미생물상을 모니터링하였다. 생균제 투여에 따라 검출되기 시작한 L. plantarum의 T-RF가 투여 중지 후 24일까지 지속됨을 확인하여 T-RFLP 분석법이 생균제로 사용한 L. plantarum의 장관 정착 평가에 유용한 것으로 나타났다. 생균제 투여 전 생쥐의 분변에서 검출되지 않았던 cinnamoyl esterase 활성 보유 L. plantarum이 투여 중지 후 24일차 분변에서 배양법으로 검출되어 L. plantarum KK3 균주의 생쥐 장관 정착이 확인되었다.

Keywords

References

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