Construction of a Full-Length Enriched and a 5′-End Enriched cDNA Library Using the Oligo-Capping Method

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Abstract

With the completion of the draft sequence of the human genome (1,2), it is now essential to extract biological information from the large volumes of human genomic sequence data. A number of attempts, which can be comprehensively termed “functional genomics,” are being carried out to decipher which parts of the human genome are transcribed, how the transcripts are spliced and translated, and what functions the eventual protein products conduct. For these purposes, a full-length complementary DNA (cDNA) that contains the entire sequence of the mRNA from the cap structure to the poly(A) tail is a unique resource because a variety of information about the gene functions is contained in a full-length cDNA sequence. The intensive analysis of a full-length cDNA would enable us to identify the following:

  1. The exact position of the mRNA transcriptional start site, which is indispensable for the identification of the adjacent promoter.

  2. The sequence of the complete 5′ untranslated region (5′ UTR), which is related to the translation efficiency and the cellular localization of mRNA.

  3. The continuous protein coding region (CDS), which is required for producing a recombinant protein.

  4. The sequence of the complete 3′ untranslated region (3′ UTR), which is related to the translation efficiency, the cellular localization, and the stability of mRNA.