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1st strand cDNA was primed with a Not I - oligo(dT)15 primer [5'pGACTAGTTCTAGATCGCGAGCGGCCGCCCTTTTTTTTTTTTTTT3']; double-stranded cDNA was ligated to Sal I adaptors (BRL), digested with Not I and cloned into the Not I and Sal I sites of the pSPORT1 vector (BRL). Library was constructed by Matthew Clark (Lehrach lab; ICRF, London and Max Planck Institut fuer Molekulare Genetik,Berlin). cDNAs for EST analysis were selected following oligonucleotide hybridization fingerprinting of arrayed clones from zebrafish late somitogenesis (26 ss), adult liver or embryonic shield stage (5.6 h) libraries. Fingerprint data were used to computationally cluster cDNAs, and a single cDNA from each cluster was chosen for sequencing. In some cases multiple members of the same cluster were sequenced to assess clustering parameters or single clones were sequenced additional times to assess quality control.
Nucleotide
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