Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1994 Jul 11;22(13):2670–2677. doi: 10.1093/nar/22.13.2670

Multiplex strand displacement amplification (SDA) and detection of DNA sequences from Mycobacterium tuberculosis and other mycobacteria.

G T Walker 1, J G Nadeau 1, P A Spears 1, J L Schram 1, C M Nycz 1, D D Shank 1
PMCID: PMC308226  PMID: 8041630

Abstract

Strand Displacement Amplification (SDA) is an isothermal, in vitro method of amplifying a DNA target sequence prior to detection [Walker et al (1992) Nucleic Acids Res., 20, 1691-1693]. Here we describe a multiplex form of SDA that allows two target sequences and an internal amplification control to be co-amplified by a single pair of primers after common priming sequences are spontaneously appended to the ends of target fragments. Multiplex SDA operates at a single temperature, under the same simple protocol previously developed for single-target SDA. We applied multiplex SDA to co-amplification of a target sequence (IS6110) that is specific to members of the Mycobacterium tuberculosis-complex and a target (16S ribosomal gene) that is common to most clinically relevant species of mycobacteria. Both targets are amplified 10(8)-fold during a 2 hour, single temperature incubation. The relative sensitivity of the system was evaluated across a number of clinically relevant mycobacteria and checked for crossreactivity against organisms that are closely related to mycobacteria.

Full text

PDF
2670

Images in this article

2675Image
on p.2675
2675Image
on p.2675

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Baess I. Determination and re-examination of genome sizes and base ratios on deoxyribonucleic acid from mycobacteria. Acta Pathol Microbiol Immunol Scand B. 1984 Aug;92(4):209–211. doi: 10.1111/j.1699-0463.1984.tb02822.x. [DOI] [PubMed] [Google Scholar]
  2. Böddinghaus B., Rogall T., Flohr T., Blöcker H., Böttger E. C. Detection and identification of mycobacteria by amplification of rRNA. J Clin Microbiol. 1990 Aug;28(8):1751–1759. doi: 10.1128/jcm.28.8.1751-1759.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Fiss E. H., Chehab F. F., Brooks G. F. DNA amplification and reverse dot blot hybridization for detection and identification of mycobacteria to the species level in the clinical laboratory. J Clin Microbiol. 1992 May;30(5):1220–1224. doi: 10.1128/jcm.30.5.1220-1224.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Jones D. H., Winistorfer S. C. Sequence specific generation of a DNA panhandle permits PCR amplification of unknown flanking DNA. Nucleic Acids Res. 1992 Feb 11;20(3):595–600. doi: 10.1093/nar/20.3.595. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Mueller P. R., Wold B. In vivo footprinting of a muscle specific enhancer by ligation mediated PCR. Science. 1989 Nov 10;246(4931):780–786. doi: 10.1126/science.2814500. [DOI] [PubMed] [Google Scholar]
  6. Plikaytis B. B., Plikaytis B. D., Yakrus M. A., Butler W. R., Woodley C. L., Silcox V. A., Shinnick T. M. Differentiation of slowly growing Mycobacterium species, including Mycobacterium tuberculosis, by gene amplification and restriction fragment length polymorphism analysis. J Clin Microbiol. 1992 Jul;30(7):1815–1822. doi: 10.1128/jcm.30.7.1815-1822.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Rogall T., Flohr T., Böttger E. C. Differentiation of Mycobacterium species by direct sequencing of amplified DNA. J Gen Microbiol. 1990 Sep;136(9):1915–1920. doi: 10.1099/00221287-136-9-1915. [DOI] [PubMed] [Google Scholar]
  8. Rogall T., Wolters J., Flohr T., Böttger E. C. Towards a phylogeny and definition of species at the molecular level within the genus Mycobacterium. Int J Syst Bacteriol. 1990 Oct;40(4):323–330. doi: 10.1099/00207713-40-4-323. [DOI] [PubMed] [Google Scholar]
  9. Shyamala V., Ames G. F. Genome walking by single-specific-primer polymerase chain reaction: SSP-PCR. Gene. 1989 Dec 7;84(1):1–8. doi: 10.1016/0378-1119(89)90132-7. [DOI] [PubMed] [Google Scholar]
  10. Soini H., Skurnik M., Liippo K., Tala E., Viljanen M. K. Detection and identification of mycobacteria by amplification of a segment of the gene coding for the 32-kilodalton protein. J Clin Microbiol. 1992 Aug;30(8):2025–2028. doi: 10.1128/jcm.30.8.2025-2028.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Spargo C. A., Haaland P. D., Jurgensen S. R., Shank D. D., Walker G. T. Chemiluminescent detection of strand displacement amplified DNA from species comprising the Mycobacterium tuberculosis complex. Mol Cell Probes. 1993 Oct;7(5):395–404. doi: 10.1006/mcpr.1993.1058. [DOI] [PubMed] [Google Scholar]
  12. Takewaki S., Okuzumi K., Ishiko H., Nakahara K., Ohkubo A., Nagai R. Genus-specific polymerase chain reaction for the mycobacterial dnaJ gene and species-specific oligonucleotide probes. J Clin Microbiol. 1993 Feb;31(2):446–450. doi: 10.1128/jcm.31.2.446-450.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Thierry D., Cave M. D., Eisenach K. D., Crawford J. T., Bates J. H., Gicquel B., Guesdon J. L. IS6110, an IS-like element of Mycobacterium tuberculosis complex. Nucleic Acids Res. 1990 Jan 11;18(1):188–188. doi: 10.1093/nar/18.1.188. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Thornton C. G., Hartley J. L., Rashtchian A. Utilizing uracil DNA glycosylase to control carryover contamination in PCR: characterization of residual UDG activity following thermal cycling. Biotechniques. 1992 Aug;13(2):180–184. [PubMed] [Google Scholar]
  15. Walker G. T. Empirical aspects of strand displacement amplification. PCR Methods Appl. 1993 Aug;3(1):1–6. doi: 10.1101/gr.3.1.1. [DOI] [PubMed] [Google Scholar]
  16. Walker G. T., Fraiser M. S., Schram J. L., Little M. C., Nadeau J. G., Malinowski D. P. Strand displacement amplification--an isothermal, in vitro DNA amplification technique. Nucleic Acids Res. 1992 Apr 11;20(7):1691–1696. doi: 10.1093/nar/20.7.1691. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Walker G. T., Little M. C., Nadeau J. G., Shank D. D. Isothermal in vitro amplification of DNA by a restriction enzyme/DNA polymerase system. Proc Natl Acad Sci U S A. 1992 Jan 1;89(1):392–396. doi: 10.1073/pnas.89.1.392. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Wilton S., Cousins D. Detection and identification of multiple mycobacterial pathogens by DNA amplification in a single tube. PCR Methods Appl. 1992 May;1(4):269–273. doi: 10.1101/gr.1.4.269. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES