Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:6.5.1.2 (DNA ligase)
 2,749 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We established a novel way to clone 5' ends of unknown length and sequence of individual cDNAs. T4 DNA ligase is employed to ligate an annealed duplex of complementary primers, one of them with a 4-nucleotide-long randomized overlap, to first-strand cDNA, generating a new 5' end. Subsequent PCR with a down-stream primer and a primer with specificity for this new 5' end leads to products that can easily be cloned and sequenced. Considerations for the choice of primers for ligation and amplification are given. We have used this method to determine the 5' sequences of three independent mRNAs: the human collagen type-X gene, the chicken anchorin CII gene, and the human cytidine deaminase gene. We will discuss this method in comparison with other methods published for the amplification of unknown 5' ends of mRNA species.
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PMID:Determination of 5' ends of specific mRNAs by DNA ligase-dependent amplification. 826 92