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Bacillus amyloliquefaciens DL-3의 carboxymethylcellulase를 재조합 균주 E. coli JM109/DL-3에서 생산하는 배지의 염 농도를 최적화하기 위한 통계학적 실험 방법의 비교

Comparison of Statistical Methods for Optimization of Salts in Medium for Production of Carboxymethylcellulase of Bacillus amyloliquefaciens DL-3 by a Recombinant E. coli JM109/DL-3

  • 이유정 (동아대학교 대학원 의생명과학과) ;
  • 김혜진 (동아대학교 대학원 의생명과학과) ;
  • 고와 (동아대학교 대학원 의생명과학과) ;
  • 정정한 (동아대학교 BK21 생물자원 실버바이오사업 인력양성단) ;
  • 이진우 (동아대학교 BK21 생물자원 실버바이오사업 인력양성단)
  • Lee, You-Jung (Department of Medical Bioscience, Graduate School of Dong-A University) ;
  • Kim, Hye-Jin (Department of Medical Bioscience, Graduate School of Dong-A University) ;
  • Gao, Wa (Department of Medical Bioscience, Graduate School of Dong-A University) ;
  • Chung, Chung-Han (BK21 Bio-Silver Program of Dong-A University) ;
  • Lee, Jin-Woo (BK21 Bio-Silver Program of Dong-A University)
  • 투고 : 2011.06.10
  • 심사 : 2011.09.05
  • 발행 : 2011.09.30

초록

재조합 균주인 E. coli JM109/DL-3를 사용하여 carboxymethylcellulase를 생산하기 위한 배지의 최적 염 농도를 orthogonal array method (OAM)과 response surface method (RSM) 등과 같은 통계학적인 방법으로 확립하고 그 결과를 비교하였다. OAM에 기초를 한 Qualitek-4 Software를 사용하여 실험을 계획하고, 그 결과를 분석한 결과는 K2HPO4가 균체의 생장 및 carboxymethylcellulase의 생산에 미치는 영향이 가장 크다는 사실을 확인하였다. 균체의 생육에 최적인 $K_2HPO_4$, NaCl, $MgSO_4{\cdot}7H_2O$$(NH_4)_2SO_4$의 농도는 10.0, 1.0, 0.2 및 0.6 g/l이었으나, carboxymethylcellulase의 생산에 최적인 각 염들의 농도는 각각 5.0, 1.0, 0.4 및 0.6 g/l이었다. RSM에 기초를 한 Design-Expert Software를 사용하여 실험을 계획하고, 그 결과를 분석한 결과는 $K_2HPO_4$가 균체의 생장 및 carboxymethylcellulase의 생산에 가장 중요한 인자라는 사실을 확인하였다. 균체의 생장에 최적인 $K_2HPO_4$, NaCl, $MgSO_4{\cdot}7H_2O$$(NH_4)_2SO_4$의 농도는 7.44, 1.08, 0.22 및 0.88 g/l이었으나, carboxymethylcellulase의 생산에 최적인 각 염들의 농도는 각각 5.84, 0.69, 0.28 및 0.54 g/l이었다. 기본적으로 OAM에 기초한 software를 사용하여 얻은 결과는 RSM에 기초한 software를 사용하여 얻은 결과와 유사하였다. 최적 조건에서 재조합 균주 E.coli JM109/DL-3이 생산하는 carboxymethylcellulase의 생산은 B. amyloliquifacience DL-에 비하여 1.92배 증가하였다.

The optimal concentrations of salts in medium for cell growth and the production of carboxymethylcellulase (CMCase) by a recombinant E. coli JM109/DL-3 were established using two statistical methods: orthogonal array method (OAM) and response surface method (RSM). The analysis of variance (ANOVA) of data based on OAM indicated that $K_2HPO_4$ gave maximum sum of square (S) and percentage contribution (P) for cell growth as well as production of CMCase. The optimal concentrations of $K_2HPO_4$, NaCl, $MgSO_4{\cdot}7H_2O$, and $(NH_4)_2SO_4$ in medium for cell growth extracted by Qualitek-4 (W32b) Software were 10.0, 1.0, 0.2, and 0.6 g/l, respectively, whereas those for the production of CMCase by E. coli JM109/DL-3 were 5.0, 1.0, 0.4, and 0.6 g/l. The analysis of variance (ANOVA) resulting from RSM indicated that a highly significant salt for cell growth was $K_2HPO_4$ ("probe>F" less than 0.0001), whereas $K_2HPO_4$ and $MgSO_4{\cdot}7H_2O$ were significant for the production of CMCase. The optimal concentrations of $K_2HPO_4$, NaCl, $MgSO_4{\cdot}7H_2O$, and $(NH_4)_2SO_4$ for cell growth extracted by Design Expert Software were 7.44, 1.08, 0.22, and 0.88 g/l, respectively, whereas those for production of CMCase were 5.84, 0.69, 0.28, and 0.54 g/l. The optimal concentrations of salts and their influences on cell growth and production of CMCase extracted by OAM were almost the same as those by RSM. Production of CMCase by a recombinant E. coli JM109/DL-3 under optimized concentration of salts was 1.93 times higher than that by Bacillus amyloliquifaciens DL-3.

키워드

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