Rapid Amplification of cDNA Ends

Both 5' and 3' RACE were performed on 12 human poly(A)⁺ RNAs (brain, heart, kidney, spleen, liver, colon, small intestine, muscle, lung, stomach, testis, placenta) using the BD SMART™ RACE cDNA amplification kit (BD BioScience-Clontech Catalogue No.634914, Mountain View, CA 95043, USA). Double-stranded cDNA synthesis and adaptor ligations to the synthesized cDNA were done according to the manufacturer's instructions. RACE fragments were separated on agarose gels and one or two strong single bands per gene purified and sequenced directly. Thus, successful RACE reactions appearing as a smear on the agarose gel would be discarded, therefore producing an approximate 54% success rate.