Termination of DNA synthesis by N6 -alkylated, not 3′- O -alkylated, photocleavable 2′-deoxyadenosine triphosphates
| العنوان: | Termination of DNA synthesis by N6 -alkylated, not 3′- O -alkylated, photocleavable 2′-deoxyadenosine triphosphates |
|---|---|
| المؤلفون: | Demetra Farley, Sherry Metzker, Brian P. Stupi, Michael L. Metzker, Sidney E. Morris, Weidong Wu, Vladislav A. Litosh, Dena L. Mansouri |
| المصدر: | Nucleic Acids Research |
| بيانات النشر: | Oxford University Press (OUP), 2007. |
| سنة النشر: | 2007 |
| مصطلحات موضوعية: | Alkylation, Base Pair Mismatch, Photochemistry, Ultraviolet Rays, DNA polymerase, Sequence analysis, DNA-Directed DNA Polymerase, 01 natural sciences, 03 medical and health sciences, chemistry.chemical_compound, symbols.namesake, Deoxyadenine Nucleotides, Deoxyadenosine, Genetics, 030304 developmental biology, Sanger sequencing, 0303 health sciences, Deoxyadenosines, biology, 010405 organic chemistry, 2'-deoxyadenosine, DNA, Sequence Analysis, DNA, Combinatorial chemistry, 0104 chemical sciences, Chemistry, Terminator (genetics), chemistry, Biochemistry, biology.protein, symbols, Human genome |
| الوصف: | The Human Genome Project has facilitated the sequencing of many species, yet the current Sanger method is too expensive, labor intensive and time consuming to accomplish medical resequencing of human genomes en masse. Of the 'next-generation' technologies, cyclic reversible termination (CRT) is a promising method with the goal of producing accurate sequence information at a fraction of the cost and effort. The foundation of this approach is the reversible terminator (RT), its chemical and biological properties of which directly impact the performance of the sequencing technology. Here, we have discovered a novel paradigm in RT chemistry, the attachment of a photocleavable, 2-nitrobenzyl group to the N(6)-position of 2'-deoxyadenosine triphosphate (dATP), which, upon incorporation, terminates DNA synthesis. The 3'-OH group of the N(6)-(2-nitrobenzyl)-dATP remains unblocked, providing favorable incorporation and termination properties for several commercially available DNA polymerases while maintaining good discrimination against mismatch incorporations. Upon removal of the 2-nitrobenzyl group with UV light, the natural nucleotide is restored without molecular scarring. A five-base experiment, illustrating the exquisite, stepwise addition through a homopolymer repeat, demonstrates the applicability of the N(6)-(2-nitrobenzyl)-dATP as an ideal RT for CRT sequencing. |
| تدمد: | 1362-4962 0305-1048 |
| الوصول الحر: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::329b08ea9142648ff7d6fc86ef42efb9Test https://doi.org/10.1093/nar/gkm689Test |
| حقوق: | OPEN |
| رقم الانضمام: | edsair.doi.dedup.....329b08ea9142648ff7d6fc86ef42efb9 |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 13624962 03051048 |
|---|