Direct DNA sequencing of PCR products.
PCR products can be sequenced using either the dideoxy (Sanger) or chemical (Maxam-Gilbert) approaches. In the dideoxy methods presented here, the target sequence is amplified and an excess of one strand of the target sequence (relative to its complement) is then generated by "asymmetric PCR," where one primer is present in vast excess over the other. This single-stranded product serves as the template for conventional dideoxy sequencing methods. Another procedure prepares PCR products for use as templates fes for characterizing unlabeled product by genomic sequencing and chemical sequencing of end-labeled products are also presented.
Dorit
R L
RL
Yale University, New Haven, Connecticut, USA.
Ohara
O
O
Hwang
C B
CB
Kim
J B
JB
Blackshaw
S
S
D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D004277:DNA, Single-Stranded / Q000235:genetics* / Q000302:isolation & purification; D054306:Dideoxynucleotides; D005092:Exonucleases; D007202:Indicators and Reagents; D016133:Polymerase Chain Reaction / Q000379:methods*; D017422:Sequence Analysis, DNA / Q000379:methods*; D013698:Templates, Genetic
D004277:DNA, Single-Stranded; D054306:Dideoxynucleotides; D007202:Indicators and Reagents; D004247:DNA; D005092:Exonucleases