Effect of DNA concentration on recombinant plasmid recovery after blunt-end ligation
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/...
| Veröffentlicht in: | BioTechniques. - 1988. - 12(1992), 2 vom: 01. Feb., Seite 190, 192-3 |
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| Weitere Verfasser: | , |
| Format: | Aufsatz |
| Sprache: | English |
| Veröffentlicht: |
1992
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| Zugriff auf das übergeordnete Werk: | BioTechniques |
| Schlagworte: | Comparative Study Journal Article DNA, Recombinant DNA Ligases EC 6.5.1.- |
| Zusammenfassung: | 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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| Beschreibung: | Date Completed 05.08.1992 Date Revised 23.11.2016 published: Print Citation Status MEDLINE |
| ISSN: | 0736-6205 |