Subcloning into large plasmid
Posted 19 October 2004 - 12:04 PM
what are optimal ligation conditions (especially temp. and time) for subcloning c. 4 kb insert into c. 16 kb plasmid (both cut with Not I & BstE II enzymes)?
Thank you for any suggestions or links.
Posted 19 October 2004 - 05:38 PM
Posted 26 October 2004 - 01:54 PM
Posted 27 October 2004 - 02:53 AM
This can for instance be achieved in a standard thermo cycler. Alternatively one can fill a thermo flask with water, put your reaction tube in it (with an appropriate floater) and put the flask o/n in a fridge (4 °C). Due to the isolation the temperature will reduce slowly.