Imaging of DNA sequences with chemiluminescence - PubMed (original) (raw)

Imaging of DNA sequences with chemiluminescence

R Tizard et al. Proc Natl Acad Sci U S A. 1990 Jun.

Abstract

We have coupled a chemiluminescent detection method that uses an alkaline phosphatase label to the genomic DNA sequencing protocol of Church and Gilbert [Church, G. M. & Gilbert, W. (1984) Proc. Natl. Acad. Sci. USA 81, 1991-1995]. Images of sequence ladders are obtained on x-ray film with exposure times of less than 30 min, as compared to 40 h required for a similar exposure with a 32P-labeled oligomer. Chemically cleaved DNA from a sequencing gel is transferred to a nylon membrane, and specific sequence ladders are selected by hybridization to DNA oligonucleotides labeled with alkaline phosphatase or with biotin, leading directly or indirectly to deposition of enzyme. If a biotinylated probe is used, an incubation with avidin-alkaline phosphatase conjugate follows. The membrane is soaked in the chemiluminescent substrate (AMPPD) and is exposed to film. Dephosphorylation of AMPPD leads in a two-step pathway to a highly localized emission of visible light. The demonstrated shorter exposure times may improve the efficiency of a serial reprobing strategy such as the multiplex sequencing approach of Church and Kieffer-Higgins [Church, G. M. & Kieffer-Higgins, S. (1988) Science 240, 185-188].

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References

1. 1. Biochemistry. 1969 Dec;8(12):4752-6 - PubMed
2. 1. Biochem Biophys Res Commun. 1968 May 10;31(3):421-6 - PubMed
3. 1. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463-7 - PubMed
4. 1. Clin Chem. 1979 Sep;25(9):1531-46 - PubMed
5. 1. Methods Enzymol. 1980;65(1):499-560 - PubMed

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