Methylated CpG Island Amplification (MCA)

Methylated CpG island Amplification Protocol written by Minoru Toyota2. Materials2.1. MCARestriction enzymes SmaI, XmaIT4 DNA ligaseTaq DNA polymerase10X PCR reaction buffer:670mM Tris-HCl, pH 8.840mM MgCl2160 mM NH4(SO4)2100 mM b -Mercaptoethanol1 mg/ml bovine serum albumin. Tris-EDTA (TE) pH 8.0DNA precipitation reagents:Phenol/Chloroform pH 8-9 3M NaOAc (for general precipitation)5M NH4Oac (for precipitation and quantitation when dNTPs are present)100% ETOHAgarose gel electrophoresis reagentsFilter hybridization reagents:96 pin replicator system (Nunc)Nylon membranesDNA hybridizati
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