Effect of DNA concentration on recombinant plasmid recovery after blunt-end ligation.
Biotechniques
; 12(2): 190, 192-3, 1992 Feb.
Article
em En
| MEDLINE
| ID: mdl-1616707
We describe conditions for optimal recovery of recombinant plasmids after blunt-end ligation. It was found that one of the most critical parameters of the blunt-end ligation reaction is total DNA concentration (vector plus incoming DNA). This concentration was optimal in the range of 1-5 micrograms/ml of reaction mixture. Concentrations larger than 10 micrograms/ml result in strong inhibition. The optimal molar relationship between incoming DNA and vector was found to be 1 or less. Under these conditions, using dephosphorylated vector, recombinants are generated at a frequency of 10(6) colonies per microgram of insert, provided that transforming efficiency is about 5 x 10(7) colonies per microgram of plasmid DNA.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Plasmídeos
/
DNA Ligases
/
DNA Recombinante
/
Clonagem Molecular
Idioma:
En
Revista:
Biotechniques
Ano de publicação:
1992
Tipo de documento:
Article
País de afiliação:
Argentina
País de publicação:
Reino Unido