Construction and Preliminary Characterization of Full length Enriched cDNA Libraries for Nonhuman Primates

Abstract
Comparative sequence analysis of primate genes is a powerful tool for studying human evolution. However, current public databases have little entries of nonhuman primate mRNA sequences and expressed sequence tags (ESTs). In an attempt to establish a database of nonhuman primate ESTs, we have constructed full length-enriched cDNA libraries for two species, cynomolgus monkey (Macaca fascicularis) and chimpanzee (Pan troglodytes), using the Oligo-Capping method. In the cynomolgus monkey, cDNA libraries were constructed from brain, liver, heart and kidney. Those from brain consisted of eight region-specific cDNA libraries (frontal lobe, temporal lobe, medulla oblongata, cerebellum cortex, hypophysis, mesencephalon, parietal lobe and brain stem). The ESTs were obtained by single-pass sequencing from the 5'-ends of randomly selected cDNA clones. A total of, 13, 738 ESTs were obtained from these cDNA libraries. In the chimpanzee, a skin cDNA library was constructed and 650 SSTs were obtained. Sequence similarity test against GenBank (database: nr and dbEST) indicated that 92.2% of ESTs in the cynomolgus monkey and 86.8% of ESTs in the chimpanzee matched with registered sequences in the public databases. In order to investigate the degree of identity in the 5'-end regions of mRNAs that include both 5'-untranslated regions (UTRs) and partial coding sequences (CDSs), 68 cynomolgus monkey ESTs and 49 chimpanzee ESTs were aligned with human homologous sequences. Sequence identities between cynomolgus monkey and human were 94.3% in the 5'-UTRs and 97.8% in the 5'-CDSs. Those between chimpanzee and human were 97.2% in 5'UTRs and 99.3% in the 5'-CDSs. These results show that the 5'-end regions of the mRNAs are highly conserved in primate species. There were, however, several exceptional genes that showed significant sequence differences in the 5'-end regions.