Genotyping Embryonic Stem (ES) Cell Colonies Prior to Picking.

MATERIALS Reagents AmpliTaq polymerase (Perkin Elmer) Bovine serum albumin (BSA) (nuclease-free; Boehringer Mannheim; stock 1.6 mg/ml) Dimethyl sulfoxide (DMSO) (Aldrich) DNA in ES PCR Lysis buffer (include positive and negative controls) dNTPs (Pharmacia; stock of the mix 10 mM) ES colonies in G418 selection (see Electroporating DNA into Embryonic Stem (ES) Cells and Selection) Light mineral oil (Sigma) ES PCR Lysis buffer Oligonucleotide primers: usually 20-mers, with a GC content of ~50% and a melting temperature between 55°C and 60°C PCR buffer (Gittschier) (10X stock) Phosphate buffered saline (PBS) Equipment Capillary tubes, finely drawn Colored markers Incubator, 5% CO2 at 37°C Microcentrifuge tube, 0.6-ml PCR equipment Stereomicroscope Genotyping Embryonic Stem (ES) Cell Colonies Prior to Picking -Soria... http://cshprotocols.cshlp.org/cgi/content/full/2006/21/pdb.prot4415?print...