comprehensive 454 transcriptome analysis of Triticum aestivum cv. kukri
Size-selected cDNA fragments were blunt-end repaired and a mixture of two partially double-stranded oligonucleotide adapters were ligated to the ends of the cDNA fragments.  PCR amplification was performed using different combinations of primers to specifically amplify cDNA fragments corresponding to the 3', 5' and internal regions of the mRNA transcripts.  The three populations of cDNA fragments were pooled in equimolar amounts and directly used for emulsion PCR to prepare immobilised template for GSFLX Titanium sequencing according to standard Roche protocols.