For gene specific library, gene specific primer (GB1-Cter-1st-4, 5’-TCCCAGAGGTATGAG-3’) was optimized for reverse transcription at 42°C using SMARTer PCR cDNA Synthesis Kit (Clontech, Mountain View, CA). Using a template switching method (18), double strand cDNAs (dscDNAs) were generated from the commercial prefrontal cortex RNAs. For the amplification step, SMARTer2A-22mer (5’-AAGCAGTGGTATCAACGCAGAG-3’) was designed based on a provided primer, SMARTerIIA Oligonucleotide, which was bound at the 5’ end of transcripts during the template switching step. After amplification with SMARTer2A-22mer and another gene specific primer (GB1-Cter-2nd-1, 5’- CTACTGGCCTGTCCTCCCTCA-3’), dscDNA generation was confirmed. Gel extraction removed unbound primers and small amplification products. Following the SOLiD™ 3 System Library Preparation Guide (ABI, Carlsbad, CA), gene specific library was prepared. For libraries of alcoholic and control samples, we used previously described primer sets (19).
