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Author Dnas, Structured ♦ Lindstroèm, Ulf M. ♦ Kool, Eric T.
Source CiteSeerX
Content type Text
File Format PDF
Subject Domain (in DDC) Computer science, information & general works ♦ Data processing & computer science
Subject Keyword Group Strategy ♦ Orthogonal Oligonucleotide ♦ Secondary Structure ♦ General Need ♦ Intact Dma Group ♦ Synthesized Unit ♦ Synthetic Dna ♦ Unwanted Hybridization ♦ Ultra Mild Deprotection ♦ Potassium Carbonate ♦ Dna Base ♦ Rna Fragment ♦ Dna Structure ♦ Undesired Site ♦ Group Dimethylacetamidine ♦ Exocyclic Amine ♦ Enzymatic Ligation ♦ General Problem ♦ Pro-tecting Group ♦ Successful Ligation ♦ Conventional Method ♦ Common Amine ♦ Protecting Group ♦ Prevent Hybridization ♦ Adenine Base ♦ Dma Group
Abstract A general problem that exists in the assembly of large and organized DNA structures from smaller fragments is secondary structure that blocks or pre-vents it. For example, it is common to assemble longer synthetic DNA and RNA fragments by liga-tion of smaller synthesized units, but blocking secondary structure can prevent the formation of the intended complex before enzymatic ligation can occur. In addition, there is a general need for pro-tecting groups that would block reactivity of some DNA bases in a sequence, leaving others free to react or hybridize. Here we describe such a strategy. The approach involves the protecting group dimethylacetamidine (Dma), which we show to remain intact on exocyclic amines of adenine bases while other bases carrying commercially available `ultra mild deprotection ' protecting groups are removed by potassium carbonate in methanol. The intact Dma groups prevent unwanted hybridization at undesired sites, thus encouraging it to occur where intended, and allowing for successful ligations. The Dma group is then deprotected by treatment with ammonia in methanol. Other common amine protecting groups such as benzoyl and allyloxycarbonyl were not successful in such a strategy, at least in part because they did not prevent hybridization. We demonstrate the method in the synthesis of a circular 54mer oligonucleotide composed of nine human telomere repeats, which was not possible to assemble by conventional methods.
Educational Role Student ♦ Teacher
Age Range above 22 year
Educational Use Research
Education Level UG and PG ♦ Career/Technical Study
Publisher Date 2002-01-01