Enhancing DNA electrotransformation efficiency in escherichia coli DH10B electrocompetent cells
Electron. j. biotechnol
; 13(5): 21-22, Sept. 2010. ilus, tab
Artigo
em Inglês
| LILACS
| ID: lil-591903
Biblioteca responsável:
CL1.1
ABSTRACT
Electrotransformation also known as electroporation is the most reliable and efficient tool for plasmid DNA uptake. Electrotransformation efficiency is function of many factors which include (1) number of cell washes prior to electroporation, (2) electroporation cell number, (3) electroporation DNA amount, and (4) cell growth phase. Those factors have limitedly been concomitantly investigated in E. coli DH10B strain. This study is aimed to explore above key factors to define the optimal conditions for high electrotransformation efficiency. The results showed that electrotransformation efficiency of E. coli DH10B was enhanced to 1.5 x 10(9) cfu/ug by washing cells three times with 15 ml of 10 percent glycerol. This washed off extra salts from cell suspension and enhanced electrotransformation by preventing arcing and enhancing cell resistance while ensuring minimal level of conductivity. Early exponential phase at 0.15 OD600 was the best growth phase for enhancing electrotransformation of E. coli DH10B. The results also showed that higher electrotransformation efficiency was similarly achieved when 0.5 x 10(10) and 0.6 x 10(10) cell numbers were electroporated with DNA amount ranging from 10 to 40 pg. This study confirmed the optimal conditions for electro competent cell preparation and plasmid DNA electrotransformation, which can result highest transformation efficiency.
Texto completo:
Disponível
Coleções:
Bases de dados internacionais
Contexto em Saúde:
Doenças Negligenciadas
Problema de saúde:
Doenças Negligenciadas
/
Zoonoses
Base de dados:
LILACS
Assunto principal:
Transformação Bacteriana
/
DNA Bacteriano
/
Eletroporação
/
Escherichia coli
Idioma:
Inglês
Revista:
Electron. j. biotechnol
Assunto da revista:
Biotecnologia
Ano de publicação:
2010
Tipo de documento:
Artigo
País de afiliação:
Estados Unidos
Instituição/País de afiliação:
Langston University/US