Anchor-ligated cDNA libraries: a technique for generating a cDNA library for the immediate cloning of the 5' ends of mRNAs.
A new, efficient and simple method based on single-stranded anchor ligation has been developed that allows for the generation of an anchor-ligated library from which the 5' ends in many mRNAs can be cloned. Random hexamers are used to prime poly A+ RNA for the first strand synthesis of a cDNA library, and a modified single-stranded anchor is then ligated to the 3' end of the cDNA using T4 RNA ligase. The library can then be used to amplify the 5' end of any mRNA expressed in the tissue. The efficiency and utility of this method has been demonstrated by amplification of three human genes: transferrin receptor, tissue-type plasminogen activator and beta actin.
Apte
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Clontech Laboratories, Palo Alto, CA 94303-4607.
Siebert
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D000199:Actins / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers; D004262:DNA Restriction Enzymes; D018076:DNA, Complementary / Q000235:genetics*; D015723:Gene Library; D006801:Humans; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000138:chemical synthesis; D010920:Placenta / Q000737:chemistry; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics*; D011990:Receptors, Transferrin / Q000235:genetics; D010959:Tissue Plasminogen Activator / Q000235:genetics
D000199:Actins; D017931:DNA Primers; D018076:DNA, Complementary; D009841:Oligonucleotides; D012333:RNA, Messenger; D011990:Receptors, Transferrin; D004262:DNA Restriction Enzymes; D010959:Tissue Plasminogen Activator