A DNA sequencing strategy that requires only five bases of known terminal sequence for priming

Journal Article · Tue Oct 24 04:00:00 EDT 1995 · Proceedings of the National Academy of Sciences of the United States of America

DOI:https://doi.org/10.1073/pnas.92.22.10162· OSTI ID:183560

Fu, D J ^[1]; Broude, N E; Smith, C L ^[2]

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1. Sequenom, Inc., Boston, MA (United States)
2. Boston Univ., MA (United States); and others

We have previously reported an enhanced version of sequencing by hybridization (SBH), termed positional SBH (PSBH). PSBH uses partially duplex probes containing single-stranded 3{prime} overhangs, instead of simple single-stranded probes. Stacking interactions between the duplex probe and a single-stranded target allow us to reduce the probe sizes required to 5-base single-stranded overhangs. Here we demonstrate the use of PSBH to capture relatively long single-stranded DNA targets and perform standard solid-state Sanger sequencing on these primer-template complexes without ligation. Our results indicate that only 5 bases of known terminal sequence are required for priming. In addition, the partially duplex probes have the ability to capture their specific target from a mixture of five single-stranded targets with different 3{prime}-terminal sequences. This indicates the potential utility of the PSBH approach to sequence mixtures of DNA targets without prior purification. 20 refs., 4 figs.

• Sponsoring Organization: USDOE
• DOE Contract Number: FG02-93ER61609
• OSTI ID: 183560
• Journal Information: Proceedings of the National Academy of Sciences of the United States of America, Journal Name: Proceedings of the National Academy of Sciences of the United States of America Journal Issue: 22 Vol. 92; ISSN PNASA6; ISSN 0027-8424
• Country of Publication: United States
• Language: English

Related Subjects

55 BIOLOGY AND MEDICINE
BASIC STUDIES
DNA SEQUENCING
EVALUATION
TECHNOLOGY ASSESSMENT