Solid-phase ligation of synthetic DNA fragments

S. A. Filippov, S. V. Kalinichenko, D. S. Esipov, E. V. Shibanova, L. N. Shingarova, V. G. Korobko, V. N. Dobrynin

M. M. Shemyakin Institute of Bioorganic Chemistry, Academy of Sciences of the USSR, Moscow

Abstract: A synthetic oligodeoxyribonucleotide (oligo) covalently bound by an internucleotide linkage to the succinylated Sephacryl S-500 support through 1,9-diaminononanc spacer was used as starting compound to assemble the E. coli rec A promoter DNA fragment from synthetic oligos by means of T4 DNA ligase. The solid-phase assembly of the designed DNA was performed by two ways: stepwise ligation of two pairs of oligos (2 diads) or simultaneous ligation of four oligos (tetrade). Both ways gave equal results with some preference in the tetrade case. The reliability of E. coli promoter DNA fragment assembly was demonstrated by cloning it in a plasmid vector and sequencing the cloned DNA by the solid-phase Maxam Gilbert technique.

Russian Journal of Bioorganic Chemistry 1990, 16 (8):1045-1051

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