Cycle sequencing using degenerate primers containing inosines

Z. Shen, J. Liu, R. L. Wells, M. M. Elkind

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

A modified protocol for cycle sequencing DNA amplified by polymerase chain reaction (PCR) is described. The method involves two sequential linear PCR amplifications using a small amount of double-stranded DNA as a template and a stringent annealing temperature: 1) α-35S-dATP labeling of specific primers initially in degenerative primer mixture and 2) dideoxyribonucleotide termination of the extended and α-35S-dATP-labeled specific primers. The method does not require end labeling and is useful in sequencing PCR-amplified DNA sequences from highly degenerate primers when the sequences of the regions flanking those to be primed are unknown.

Original languageEnglish (US)
Pages (from-to)83+85+88+90
JournalBioTechniques
Volume15
Issue number1
StatePublished - Jan 1 1993
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Biochemistry, Genetics and Molecular Biology(all)

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