CXIV. Total synthesis of the structural gene for an alanine transfer RNA from yeast. Enzymic joining of the chemically synthesized segments to form the DNA duplex corresponding to nucleotide sequence 46 to 77

Abstract

The DNA duplex corresponding to the nucleotide sequence 46 to 77 of the major yeast alanine transfer RNA (Fig. 1) has been synthesized. Enzymic joining was studied using reaction mixtures containing three, four, five and all of the six oligodeoxynucleotide segments. Several of the joining systems consisting of three or four segments either failed to show any joining or gave only moderate joining. During this work, the T4 ligase was found to promote the formation of bihelical structures when ATP was absent. The five-component system consisting of segments 11 to 15 gave rapid joinings to give the covalently linked duplex in the extent of 60 to 70%. The sixth component (segment 10) could be joined subsequently to give the duplex [C](Fig. 1]. Alternatively, [C] could also be prepared in a one-step reaction involving all the six components (Fig. 1) in a yield of 60%.