Gene knockout and knockin on the Escherichia coli lac operon loci using pBR322-red system / 生物工程学报
Chinese Journal of Biotechnology
; (12): 192-197, 2005.
Article
em Zh
| WPRIM
| ID: wpr-249926
Biblioteca responsável:
WPRO
ABSTRACT
pBR322-Red is a newly constructed recombineering plasmid, which contains a part of the pBR322 vector, a series of regulatory elements of lambda-prophage and Red recombination genes. In the beginning, we studied the best working conditions of pBR322-Red, and then modified lac operon in E. coli W3110 chromosome using the plasmid as follow: Firstly, we knockout the lacI gene using Red-mediated recombineering with overlapping single stranded DNA oligonucleotides. Secondly, we substituded the lacA and lacY genes with lacZ, a report gene, by Red-mediated linearized double strands DNA homologous recombination. Finally, we detected the expression of lacZ on these loci for the first time. The results suggested that pBR322-Red system is suitable for modifying W3110 chromosome with various recombination strategies.
Texto completo:
1
Índice:
WPRIM
Assunto principal:
Plasmídeos
/
Recombinação Genética
/
Cromossomos Bacterianos
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Bacteriófago lambda
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Escherichia coli
/
Técnicas de Introdução de Genes
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Técnicas de Inativação de Genes
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Genética
/
Óperon Lac
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Métodos
Limite:
Humans
Idioma:
Zh
Revista:
Chinese Journal of Biotechnology
Ano de publicação:
2005
Tipo de documento:
Article