- Digestion, Vector Preparation and Ligation (Lykke-Andersen Lab, Department of MCD Biology, University of Colorado)
Procedures for cloning including digestion, dephosphorylation of vector, preparation of insert and ligation
Added: Thu Feb 26 2009, Hits: 6312, Reviews: 0
Cloning from Low Melt Agarose
(Nonet Lab - Washington University School of Medicine)
A quick method for cloning fragments with the following ends: 1) a double cut fragment being cloned into a double cut vector if the two restriction sites are non-cohesive. 2) a single cut fragment being cloned into a single cut vector if the vector has been properly dephosphorylated. 3) A small blunt fragment, such as PCR product, being cloned into a dephosphorylated blunt vector.
Added: Sat Nov 01 2003, Hits: 1549, Reviews: 0
Ligations in Low Melting Temperature Agarose
(Donis Keller Lab)
Ligation of plasmid vector and insert DNAs is carried out directly in melted agarose gel slices. This method is used for subcloning well defined restriction fragments and is probably not suitable for genomic library construction. The method works for both blunt-end and cohesive termini ligations.
Added: Tue May 14 2002, Hits: 1105, Reviews: 0
- Subcloning Tips - very important heating step (Schimmelpenninck)
Put your insert and vector in an Eppendorff tube and then heat it for 5 minutes, then add ligation buffer and ligase
Added: Wed Sep 25 2002, Reviews: 0