Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Review Article
  • Published:

The rapid detection of unknown mutations in nucleic acids

Abstract

The task of identifying mutations in nucleic acid sequences is a vital component of research in mammalian genetics. With the advent of the polymerase chain reaction, several useful mutation detection techniques have evolved in recent years. The different methods have complementing strengths and a suitable procedure for virtually any experimental situation is now available.

This is a preview of subscription content, access via your institution

Access options

Buy this article

Prices may be subject to local taxes which are calculated during checkout

Similar content being viewed by others

References

  1. Chehab, F.F. & Wall, J. Detection of multiple cystic fibrosis mutations by reverse dot blot hybridization: a technology for carrier screening. Hum. Genet. 89, 163–168 (1992).

    Article  CAS  PubMed  Google Scholar 

  2. Ng, I.S. et al. Methods for analysis of multiple cystic fibrosis mutations. Hum. Genet. 87, 613–617 (1991).

    Article  CAS  PubMed  Google Scholar 

  3. Ballabio, A. et al. Molecular heterogeneity of steroid sulfatase deficiency: a multicenter study on 57 unrelated patients, at DNA and protein levels. Genomics 4, 36–40 (1989).

    Article  CAS  PubMed  Google Scholar 

  4. Tsui, L.C. The spectrum of cystic fibrosis mutations. Trends Genet. 8, 392–398 (1992).

    Article  CAS  PubMed  Google Scholar 

  5. Koenig, M., Monaco, A.P. & Kunkel, L.M. The complete sequence of dystrophin predicts a rod-shaped cytoskeletal protein. Cell 53, 219–226 (1988).

    Article  CAS  PubMed  Google Scholar 

  6. Sinclair, A.H. et al. A gene from the human sex-determining region encodes a protein with homology to a conserved DNA-binding motif. Nature 346, 240–244 (1990).

    Article  CAS  PubMed  Google Scholar 

  7. Rooney, D.E. & Czepulkowski, B.H. Human cytogenetics: a practical approach (IRL Press, Oxford, New York, 1992).

    Google Scholar 

  8. Zhang, F.R., Heilig, R., Thomas, G. & Aurias, A. A one-step efficient and specific non-radioactive non-fluorescent method for in situ hybridization of banded chromosomes. Chromosoma 99, 436–439 (1990).

    Article  CAS  PubMed  Google Scholar 

  9. Baurmann, H., Cherif, D. & Berger, R. Interphase cytogenetics by fluorescent in situ hybridization (FISH) for characterization of monosomy-7-associated myeloid disorders. Leukemia 7, 384–391 (1993).

    CAS  PubMed  Google Scholar 

  10. Taylor, C. et al. Diagnosis of Ewing's sarcoma and peripheral neuroectodermal tumour based on the detection of t(11;22) using fluorescence in situ hybridization. Br. J. Cancer 67, 128–133 (1993).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Kuwano, A., Ledbetter, S.A., Dobyns, W.B., Emanuel, B.S. & Ledbetter, D.H. Detection of deletions and cryptic translocations in Miller-Dieker syndrome by In situ hybridization. Am. J. hum. Genet. 49, 707–714 (1991).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Lupski, J.R. et al. DNA duplication associated with Charcot-Marie-Tooth disease type 1A. Cell. 66, 219–232 (1991).

    Article  CAS  PubMed  Google Scholar 

  13. Southern, E.M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J. molec. Blol. 98, 503–517 (1975).

    Article  CAS  Google Scholar 

  14. Cooper, D.N. & Youssoufian, H. The CpG dinucleotide and human genetic disease. Hum. Genet. 78, 151–155 (1988).

    Article  CAS  PubMed  Google Scholar 

  15. Mullis, K.B. & Faloona, F.A. Specific synthesis of DNA in vitro via a polymerase-catalyzed chain reaction. Meth. Enzymol. 155, 335–350 (1987).

    Article  CAS  Google Scholar 

  16. Sanger, F., Nicklen, S. & Coulson, A.R. DNA sequencing with chain-terminating inhibitors. Proc. natn. Acad. Sci. U.S.A. 74, 5463–5467 (1977).

    Article  CAS  Google Scholar 

  17. Maxam, A.M. & Gilbert, W. A new method for sequencing DNA. Proc. natn. Acad. Sci. U.S.A. 74, 560–564 (1977).

    Article  CAS  Google Scholar 

  18. Chamberlain, J.S., Gibbs, R.A., Ranier, J.E., Nguyen, P.N. & Caskey, C.T. Deletion screening of the Duchenne muscular dystrophy locus via multiplex DNA amplification. Nucleic Acids Res. 16, 11141–11156 (1988).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Niemann, S.S. et al. Molecular genetic analysis of 67 patients with Duchenne/Becker muscular dystrophy. Hum. Genet. 90, 65–70 (1992).

    Google Scholar 

  20. Gibbs, R.A., Nguyen, P.N., Edwards, A., Civitello, A.B. & Caskey, C.T. Multiplex DNA deletion detection and exon sequencing of the hypoxanthine phosphoribosyltransferase gene in Lesch-Nyhan families. Genomics 7, 235–244 (1990).

    Article  CAS  PubMed  Google Scholar 

  21. Ballabio, A., Ranier, J.E., Chamberlain, J.S., Zollo, M. & Caskey, C.T. Screening for steroid sulfatase (STS) gene deletions by multiplex DNA amplification. Hum. Genet. 84, 571–573 (1990).

    Article  CAS  PubMed  Google Scholar 

  22. Abbs, S. & Bobrow, M. Analysis of quantitative PCR for the diagnosis of deletion and duplication carriers in the dystrophin gene. J. med. Genet. 29, 191–196 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Orita, M., Iwahana, H., Kanazawa, H., Hayashi, K. & Sekiya, T. Detection of polymorphisms of human DNA by gel electrophoresis as single-strand conformation polymorphisms. Proc. natn. Acad. Sci. U.S.A. 86, 2766–2770 (1989).

    Article  CAS  Google Scholar 

  24. Ainsworth, P.J., Surh, L.C. & Coulter, M.M.B. Diagnostic single strand conformational polymorphism, (SSCP): a simplified non-radioisotopic method as applied to a Tay-Sachs B1 variant. Nucl. Acids Res. 19, 405–406 (1991).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Yap, E.P. & McGee, J.O. Nonisotopic SSCP detection in PCR products by ethidium bromide staining. Trends Genet. 8, 49 (1992).

    Article  CAS  PubMed  Google Scholar 

  26. Michaud, J. et al. Strand-separating conformational polymorphism analysis: efficacy of detection of point mutations in the human ornithine delta-aminotransferase gene. Genomics 13, 389–394 (1992).

    Article  CAS  PubMed  Google Scholar 

  27. Sheffield, V.C., Beck, J.S. & Kwitek, A.E. Analysis of the efficiency of single base substitution detection by SSCP 1–149 (Cold Spring Harbor Laboratory, New York, 1992).

    Google Scholar 

  28. Iwahana, H., Yoshimoto, K. & Itakara, M. Detection of point mutations by SSCP of PCR-amplified DNA after endonuclease digestion. Biotechniques 12, 64 (1992).

    CAS  PubMed  Google Scholar 

  29. Sarkar, G., Yoon, H.S. & Sommer, S.S. Screening for mutations by RNA single-strand conformation polymorphism (rSSCP): comparison with DNA-SSCP. Nucl. Acids Res. 20, 871–878 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Myers, R.M., Maniatis, T. & Lerman, L.S. Detection and localization of single base changes by denaturing gradient gel electrophoresis. Meth. Enzymol. 155, 501–527 (1987).

    Article  CAS  Google Scholar 

  31. Sheffield, V.C., Cox, D.R., Lerman, L.S. & Myers, R.M. Attachment of a 40-base-pair G + C-rich sequence (GC-clamp) to genomic DNA fragments by the polymerase chain reaction results in improved detection of single-base changes. Proc. natn. Acad. Sci. U.S.A. 86, 232–236 (1989).

    Article  CAS  Google Scholar 

  32. Lerman, L.S. & Silverstein, K. Computational simulation of DNA melting and its application to denaturing gel electrophoresis. Meth. Enzymol. 155, 482–501 (1987).

    Article  CAS  Google Scholar 

  33. Myers, R.M., Fischer, S.G., Lerman, L.S. & Maniatis, T. Nearly all single base substitutions in DNA fragments joined to a GC-clamp can be detected by denaturing gradient gel electrophoresis. Nucl. Acids Res. 13, 3131–3145 (1985).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Pellegata, N.S. et al. Detection of K-ras mutations by denaturing gradient gel electrophoresis (DGGE): a study on pancreatic cancer. Anticancer Res. 12, 1731–1735 (1992).

    CAS  PubMed  Google Scholar 

  35. Fodde, R. et al. Eight novel inactivating germ line mutations at the APC gene identified by denaturing gradient gel electrophoresis. Genomics 13, 1162–1168 (1992).

    Article  CAS  PubMed  Google Scholar 

  36. Rosatelli, M.C. et al. Molecular characterization of beta-thalassemia in the Sardinian population. Am. J. hum. Genet. 50, 422–426 (1992).

    CAS  PubMed  PubMed Central  Google Scholar 

  37. Tee, M.K., Moran, C. & Nicholas, F.W. Temperature gradient gel electrophoresis: detection of a single base substitution in the cattle beta-lactoglobulin gene. Anim. Genet. 23, 431–435 (1992).

    Article  CAS  PubMed  Google Scholar 

  38. Nagamine, C.M., Chan, K. & Lau, Y.F.C. A PCR artifact: Generation of heteroduplexes. Am. J. hum. Genet. 45, 337–339 (1989).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Perry, D.J. & Carrell, R.W. Hydrolink gels: A rapid and simple approach to the detection of DNA mutations in thromboembolic disease. J. clin. Pathol. 45, 158–160 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. White, M.B., Carvalho, M., Derse, D., O'Brien, S.J. & Dean, M. Detecting single base substitutions as heteroduplex polymorphisms. Genomics 12, 301–306 (1992).

    Article  CAS  PubMed  Google Scholar 

  41. Tassabehi, M. et al. Waardenburg's syndrome patients have mutations in the human homologue of the pax-3 paired box gene. Nature 355, 635–636 (1992).

    Article  Google Scholar 

  42. Keen, J., Lester, D., Ingleheam, C., Curtis, A. & Bhattacharya, S. Rapid detection of single base mismatches as heteroduplexes on HydroLink gels. Trends Genet. 7, 5 (1991).

    Article  CAS  PubMed  Google Scholar 

  43. Artlich, A., Horn, M., Lorenz, B., Bhattacharya, S. & Gal, S. Recurrent 3-bp deletion at codon 255/256 of the rhodopsin gene in a German pedigree with autosomal dominant retinitis pigmentosa. Am. J. hum. Genet. 50, 876–878 (1992).

    CAS  PubMed  PubMed Central  Google Scholar 

  44. Veres, G., Gibbs, R.A., Scherer, S.E. & Caskey, C.T. The molecular basis of the sparse fur mouse mutation. Science 237, 415–417 (1987).

    Article  CAS  PubMed  Google Scholar 

  45. Gibbs, R.A. & Caskey, C.T. Identification and localization of mutations at the Lesch-Nyhan locus by ribonuclease A cleavage. Science 236, 303–305 (1987).

    Article  CAS  PubMed  Google Scholar 

  46. Myers, R.M., Larin, Z. & Maniatis, T. Detection of single base substitutions by ribonuclease cleavage at mismatches in RNA:DNA duplexes. Science 230, 1242–1246 (1985).

    Article  CAS  PubMed  Google Scholar 

  47. Cotton, R.G., Rodrigues, N.R. & Campbell, R.D. Reactivity of cytosine and thymine in single-base-pair mismatches with hydroxylamine and osmium tetroxide and its application to the study of mutations. Proc. natn. Acad. Sci. U.S.A. 85, 4397–4401 (1988).

    Article  CAS  Google Scholar 

  48. Forrest, S.M., Dahl, H.H., Howells, D.W., Dianzani, I. & Cotton, R.G. Mutation detection in phenylketonuria by using chemical cleavage of mismatch: importance of using probes from both normal and patient samples. Am. J. hum. Genet. 49, 175–183 (1991).

    CAS  PubMed  PubMed Central  Google Scholar 

  49. Zheng, H. et al. Fidelity of targeted recombination in human fibroblasts and murine embryonic stem cells. Proc. natn. Acad. Sci. U.S.A. 88, 8067–8071 (1991).

    Article  CAS  Google Scholar 

  50. Saleeba, J.A., Ramus, S.J. & Cotton, R.G.H. Complete mutation detection using unlabeled chemical cleavage. Hum. Mut. 1, 63–69 (1992).

    Article  CAS  PubMed  Google Scholar 

  51. Kilimann, M.W., Pizzuti, A., Grompe, M. & Caskey, C.T. Point mutations and polymorphisms in the human dystrophin gene identified in genomic DNA sequences amplified by multiplex PCR. Hum. Genet. 89, 253–258 (1992).

    Article  CAS  PubMed  Google Scholar 

  52. Ganguly, A. & Prockop, D.J. Detection of single–base mutations by reaction of DNA heteroduplexes with a water–soluble carbodiimide followed by primer extension: application to products from the polymerase chain reaction. Nucl. Acids Res. 18, 3933–3939 (1990).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Gyllensten, U.B. & Erlich, H.A. Generation of single-stranded DNA by the polymerase chain reaction and its application to direct sequencing of the HLA-DQA locus. Proc. natn. Acad. Sci. U.S.A. 85, 7652–7656 (1988).

    Article  CAS  Google Scholar 

  54. Syvanen, A.C., Aalto, S.K., Kontula, K. & Soderlund, H. Direct sequencing of affinity-captured amplified human DNA application to the detection of apolipoprotein E polymorphism. FEBS Lett. 258, 71–74 (1989).

    Article  CAS  PubMed  Google Scholar 

  55. Stoflet, E.S., Koeberl, D.D., Sarkar, G. & Sommer, S.S. Genomic amplification with transcript sequencing. Science 239, 491–494 (1988).

    Article  CAS  PubMed  Google Scholar 

  56. Ruano, G. & Kidd, K.K. Coupled amplification and sequencing of genomic DNA. Proc. natn. Acad. Sci. U.S.A. 88, 2815–2819 (1991).

    Article  CAS  Google Scholar 

  57. Craxton, M. Linear amplification sequencing, a powerful method for sequencing DNA. Methods 3, (1991).

    Article  CAS  Google Scholar 

  58. Rosenthal, A. & Charnock, J.D.S. New protocols for DNA sequencing with dye terminators. DNA Seq. 3, 61–64 (1992).

    Article  CAS  PubMed  Google Scholar 

  59. Sarkar, G., Yoon, H.S. & Sommer, S.S. Dideoxy fingerprinting (ddE): a rapid and efficient screen for the presence of mutations. Genomics 13, 441–443 (1992).

    Article  CAS  PubMed  Google Scholar 

  60. Hattori, M., Yoshioka, K. & Sakaki, Y. High–sensitive fluorescent DNA sequencing and its application for detection and mass-screening of point mutations. Electrophoresis 13, 560–565 (1992).

    Article  CAS  PubMed  Google Scholar 

  61. Lu, A.L. & Hsu, I.C. Detection of single DNA base mutations with mismatch repair enzymes. Genomics 14, 249–255 (1992).

    Article  CAS  PubMed  Google Scholar 

  62. Chelly, J., Concordet, J.P., Kaplan, J.C. & Kahn, A. Illegitimate transcription: transcription of any gene in any cell type. Proc. natn. Acad. Sci. U.S.A. 86, 2617–2621 (1989).

    Article  CAS  Google Scholar 

  63. Chelly, J. et al. Illegitimate transcription. Application to the analysisof truncated transcripts of the dystrophin gene in nonmuscle cultured cells from Duchenne and Becker patients. J. clin. Invest. 88, 1161–1166 (1991).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. Jackson, D.P., Hayden, J.D. & Quirke, P. in PCR: A practical approach. (eds. McPherson, M.J., Quirke, P. & Taylor, G.R.) 41–42 (IRL press, Oxford, 1991).

    Google Scholar 

  65. Lin, B., Cotton, R.G., Trent, D.W. & Wright, P.J. Geographical clusters of dengue virus type 2 isolates based on analysis of infected cell RNA by the chemical cleavage at mismatch method. J. virol. Methods. 40, 205–218 (1992).

    Article  CAS  PubMed  Google Scholar 

  66. Grompe, M., Versalovic, J., Koeuth, T. & Lupski, J.R. Mutations in the Escherichia coli dnaG gene suggest coupling between DNA replication and chromosome partitioning. J. Bacteriol. 173, 1268–1278 (1991).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  67. Sheffield, V.C., Fishman, G.A., Beck, J.S., Kimura, A.E. & Stone, E.M. Identification of novel rhodopsin mutations associated with retinitis pigmentosa by GC-clamped denaturing gradient gel electrophoresis. Am. J. hum. Genet. 49, 699–706 (1991).

    CAS  PubMed  PubMed Central  Google Scholar 

  68. Rossiter, J.F.R., Grompe, M. & Caskey, C.T. in PCR: A practical approach. (eds. McPherson, M.J., Quirke, P. & Taylor, G.R.) 75–83 (IRL press, Oxford, 1991).

    Google Scholar 

  69. Bevan, I.S., Rapley, R. & Walker, M.R. Sequencing of PCR-amplified DNA. PCR Meth. Appl. 1, 222–228 (1992).

    Article  CAS  Google Scholar 

  70. Grompe, M., Muzny, D.M. & Caskey, C.T. Scanning detection of mutations in human ornithine transcarbamoylase by chemical mismatch cleavage. Proc. natn. Acad. Sci. U.S.A. 86, 5888–5892 (1989).

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Grompe, M. The rapid detection of unknown mutations in nucleic acids. Nat Genet 5, 111–117 (1993). https://doi.org/10.1038/ng1093-111

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1038/ng1093-111

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing