가자미식해로부터 항균활성 유산균의 분리 및 동정
Isolation and Identification of Antimicrobial Lactic Acid Bacteria from Flounder Sikhae
- 주제(키워드) Lactic Acid Bacteria , Sikhae , Antimicrobial
- 발행기관 강릉원주대학교 일반대학원
- 지도교수 조순영
- 발행년도 2015
- 학위수여년월 2015. 8
- 학위명 석사
- 학과 및 전공 일반대학원 식품과학과
- 원문페이지 59
- 실제URI http://www.dcollection.net/handler/kangnung/000000007474
- 본문언어 한국어
초록/요약
This study has tested the antimicrobial activity in Gram(+) 2 strains(Bacillus cereus, Staphylococcus aureus) and Gram(-) 2 strains(Salmonella enterica, Escherichia coli) by separating unspecified number of lactic acid bacteria from flounder sikhae and creating a single colony state through subculture. Based on the result of the antimicrobial activity test by the O/D measurement using a microplate reader, the top 10 strains had an outstanding antimicrobial activity for each food borne pathogen were screened and then finally, the lactic acid bacteria had a high antimicrobial activity were selected based on the size measurement result of the inhibition zone by using the paper disc method. It was measured that the SL097 which is the finally selected lactic acid bacteria has 13 ㎜ clear zone in Bacillus cereus, 13 ㎜ clear zone in Staphylococcus aureus, 17㎜ clear zone in Salmonella enterica and 12㎜ clear zone in Escherichia coli. The identification result of the SL097 which has the biggest antimicrobial activity in tested lactic acid bacterias based on 16S rRNA gene nucleotide sequence has shown that it has 99% homology with Weissella halotolerans strain NRIC 1627 so that it was identified as Weissella halotolerans SL097. As a following experiment, the effect on the culture method, culture temperature, culture time, carbon source, nitrogen source, inorganic salts and pH was checked in order to review optimal growth conditions of W. halotolerans SL097. The test result of optimum growth conditions of W. halotolerans SL097 has shown that it’s considered as microaerophilic bacteria as the general culture in the incubator showed a better growth activity rather than the shaking culture and the highest growth activity were seen in 30℃ of the culture temperature and 36 hours of the incubation time. The highest growth activity was seen in maltose 2% of carbon source, enzymatic digest of animal tissue 1.0% of nitrogen source, beef extract 1.0%, yeast extract 0.5% and tryptone 1.0% and the growth was excellent in the medium where ammonium citrate 1.0% and NaCl 1.0% are added as inorganic salts. In terms of the growth activity by pH, it was excellent in pH 7. Also, it was confirmed that the growth is possible also in pH 2~3.
more목차
Ⅰ. 서론 1
Ⅱ. 재료 및 방법 3
1. 실험재료 3
2. 실험방법 3
1) 가자미식해 제조 3
2) 유산균 분리 및 선별 4
3) 항균활성 검정 4
(1) Microplate Reader를 이용한 O/D 측정법 6
(2) Paper disc법 6
4) 16S rRNA gene sequencing 7
(1) Chromosomal DNA의 추출 7
(2) 16S rRNA gene PCR 증폭 및 염기서열 분석 8
5) 최적생육조건 탐색 9
(1) 배양방법의 영향 9
(2) 배양온도의 영향 9
(3) 배양시간의 영향 9
(4) 탄소원의 영향 10
(5) 질소원의 영향 10
(6) 무기염류의 영향 11
(7) pH의 영향 11
6) 통계분석 12
Ⅲ. 결과 및 고찰 13
1. 유산균 분리 및 선별 13
2. 항균활성 검정 13
1) Microplate Reader를 이용한 O/D 측정 14
2) Paper disc법을 이용한 항균활성 26
3. 16S rRNA gene sequencing 35
4. 최적생육조건 38
1) 배양방법의 영향 38
2) 배양온도의 영향 40
3) 배양시간의 영향 42
4) 탄소원의 영향 44
5) 질소원의 영향 47
6) 무기염류의 영향 49
7) pH의 영향 52
Ⅳ. 요약 54
Ⅴ. 참고문헌 56
