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  • Title: Joining of 3'-modified oligonucleotides by T4 RNA ligase. Synthesis of a heptadecanucleotide corresponding to the bases 61--77 from Escherichia coli tRNAfMet.
    Author: Ohtsuka E, Nishikawa S, Markham AF, Tanaka S, Miyake T, Wakabayashi T, Ikehara M, Sugiura M.
    Journal: Biochemistry; 1978 Nov 14; 17(23):4894-9. PubMed ID: 363145.
    Abstract:
    Chemically synthesized fragments corresponding to the 3' end of tRNAfMet from Escherichia coli were joined by T4-induced RNA ligase to yield a heptadecanucleotide (bases 61--77). The 3' terminus of C-C-A was modified by introduction of the ethoxymethylidene group to prevent intra- and intermolecular self-joining reactions at the 3' end. The terminal trimer was phosphorylated using polynucleotide kinase and joined to C-A-A with RNA ligase. The hexamer [C-A-A-C-C-A(ethoxymethylidene)] corresponding to bases 72--77 was obtained in a yield of 60%. An undecanucleotide (bases 61--71) which had been synthesized in a yield of 34% by similar enzymatic joining of U-C-C-G-G to pC-C-C-C-C-G was allowed to react with the 5'-phosphorylated hexamer (bases 72--77) using an excess of RNA ligase to yield the heptadecanucleotide U-C-C-G-G-C-C-C-C-C-G-C-A-A-C-C-A (bases 61--77). The product was identified by homochromatography and nearest neighbor analysis.
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