A DNA cycle sequencing reaction that minimizes compressions on automated fluorescent sequencers.
We recently demonstrated that most band compressions (>70%) on DNA sequencing gels result from the presence of a single sequence motif, 5'-YGN1-2AR-3', where Y and R indicate base-pairing pyrimidine and purine residues, respectively. This finding raised the possibility that the use of 7-deaza-dATP instead of dATP in chain termination sequencing reactions would resolve most of the band compressions. Thus, we examined the effects of 7-deaza-dATP on DNA sequencing using thermostable DNA polymerases. The results indicate that the replacement of dATP with 7-deaza-dATP in conventional cycle sequencing reactions can successfully eliminate most band compressions without sacrificing sequencing performance.
Yamakawa
H
H
Laboratory of DNA Technology, Kazusa DNA Research Institute, 1532-3 Yana, Kisarazu, Chiba 292, Japan.
Ohara
O
O
D001331:Automation; D001482:Base Composition; D001483:Base Sequence; D004247:DNA / Q000737:chemistry*; D003838:Deoxyadenine Nucleotides; D007202:Indicators and Reagents; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry*; D011687:Purines; D011743:Pyrimidines; D013050:Spectrometry, Fluorescence / Q000379:methods
D003838:Deoxyadenine Nucleotides; D007202:Indicators and Reagents; D009838:Oligodeoxyribonucleotides; D011687:Purines; D011743:Pyrimidines; D004247:DNA; C030986:pyrimidine; C026600:2'-deoxyadenosine triphosphate