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Proc Natl Acad Sci U S A. Nov 1990; 87(22): 8923–8927.

Automated DNA diagnostics using an ELISA-based oligonucleotide ligation assay.


DNA diagnostics, the detection of specific DNA sequences, will play an increasingly important role in medicine as the molecular basis of human disease is defined. Here, we demonstrate an automated, nonisotopic strategy for DNA diagnostics using amplification of target DNA segments by the polymerase chain reaction (PCR) and the discrimination of allelic sequence variants by a colorimetric oligonucleotide ligation assay (OLA). We have applied the automated PCR/OLA procedure to diagnosis of common genetic diseases, such as sickle cell anemia and cystic fibrosis (delta F508 mutation), and to genetic linkage mapping of gene segments in the human T-cell receptor beta-chain locus. The automated PCR/OLA strategy provides a rapid system for diagnosis of genetic, malignant, and infectious diseases as well as a powerful approach to genetic linkage mapping of chromosomes and forensic DNA typing.

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  • Landegren U, Kaiser R, Caskey CT, Hood L. DNA diagnostics--molecular techniques and automation. Science. 1988 Oct 14;242(4876):229–237. [PubMed]
  • Antonarakis SE. Diagnosis of genetic disorders at the DNA level. N Engl J Med. 1989 Jan 19;320(3):153–163. [PubMed]
  • Botstein D, White RL, Skolnick M, Davis RW. Construction of a genetic linkage map in man using restriction fragment length polymorphisms. Am J Hum Genet. 1980 May;32(3):314–331. [PMC free article] [PubMed]
  • Donis-Keller H, Green P, Helms C, Cartinhour S, Weiffenbach B, Stephens K, Keith TP, Bowden DW, Smith DR, Lander ES, et al. A genetic linkage map of the human genome. Cell. 1987 Oct 23;51(2):319–337. [PubMed]
  • Lander ES. DNA fingerprinting on trial. Nature. 1989 Jun 15;339(6225):501–505. [PubMed]
  • Nakamura Y, Leppert M, O'Connell P, Wolff R, Holm T, Culver M, Martin C, Fujimoto E, Hoff M, Kumlin E, et al. Variable number of tandem repeat (VNTR) markers for human gene mapping. Science. 1987 Mar 27;235(4796):1616–1622. [PubMed]
  • Southern EM. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. [PubMed]
  • Conner BJ, Reyes AA, Morin C, Itakura K, Teplitz RL, Wallace RB. Detection of sickle cell beta S-globin allele by hybridization with synthetic oligonucleotides. Proc Natl Acad Sci U S A. 1983 Jan;80(1):278–282. [PMC free article] [PubMed]
  • Myers RM, Maniatis T, Lerman LS. Detection and localization of single base changes by denaturing gradient gel electrophoresis. Methods Enzymol. 1987;155:501–527. [PubMed]
  • Cotton RG, Rodrigues NR, Campbell RD. Reactivity of cytosine and thymine in single-base-pair mismatches with hydroxylamine and osmium tetroxide and its application to the study of mutations. Proc Natl Acad Sci U S A. 1988 Jun;85(12):4397–4401. [PMC free article] [PubMed]
  • 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 Natl Acad Sci U S A. 1989 Apr;86(8):2766–2770. [PMC free article] [PubMed]
  • Chehab FF, Kan YW. Detection of specific DNA sequences by fluorescence amplification: a color complementation assay. Proc Natl Acad Sci U S A. 1989 Dec;86(23):9178–9182. [PMC free article] [PubMed]
  • Newton CR, Graham A, Heptinstall LE, Powell SJ, Summers C, Kalsheker N, Smith JC, Markham AF. Analysis of any point mutation in DNA. The amplification refractory mutation system (ARMS). Nucleic Acids Res. 1989 Apr 11;17(7):2503–2516. [PMC free article] [PubMed]
  • Wu DY, Ugozzoli L, Pal BK, Wallace RB. Allele-specific enzymatic amplification of beta-globin genomic DNA for diagnosis of sickle cell anemia. Proc Natl Acad Sci U S A. 1989 Apr;86(8):2757–2760. [PMC free article] [PubMed]
  • Saiki RK, Scharf S, Faloona F, Mullis KB, Horn GT, Erlich HA, Arnheim N. Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science. 1985 Dec 20;230(4732):1350–1354. [PubMed]
  • Mullis KB, Faloona FA. Specific synthesis of DNA in vitro via a polymerase-catalyzed chain reaction. Methods Enzymol. 1987;155:335–350. [PubMed]
  • Saiki RK, Gelfand DH, Stoffel S, Scharf SJ, Higuchi R, Horn GT, Mullis KB, Erlich HA. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science. 1988 Jan 29;239(4839):487–491. [PubMed]
  • Wu DY, Wallace RB. The ligation amplification reaction (LAR)--amplification of specific DNA sequences using sequential rounds of template-dependent ligation. Genomics. 1989 May;4(4):560–569. [PubMed]
  • Pinto-Cisternas J, Castro de Guerra D. Utilidad de los apellidos en estudios de biología humana. Usos y perspectivas en poblaciones Latinoamericanas. Rev Med Chil. 1988 Nov;116(11):1191–1197. [PubMed]
  • Guatelli JC, Whitfield KM, Kwoh DY, Barringer KJ, Richman DD, Gingeras TR. Isothermal, in vitro amplification of nucleic acids by a multienzyme reaction modeled after retroviral replication. Proc Natl Acad Sci U S A. 1990 Mar;87(5):1874–1878. [PMC free article] [PubMed]
  • Kwoh DY, Davis GR, Whitfield KM, Chappelle HL, DiMichele LJ, Gingeras TR. Transcription-based amplification system and detection of amplified human immunodeficiency virus type 1 with a bead-based sandwich hybridization format. Proc Natl Acad Sci U S A. 1989 Feb;86(4):1173–1177. [PMC free article] [PubMed]
  • Landegren U, Kaiser R, Sanders J, Hood L. A ligase-mediated gene detection technique. Science. 1988 Aug 26;241(4869):1077–1080. [PubMed]
  • Alves AM, Carr FJ. Dot blot detection of point mutations with adjacently hybridising synthetic oligonucleotide probes. Nucleic Acids Res. 1988 Sep 12;16(17):8723–8723. [PMC free article] [PubMed]
  • Wu DY, Wallace RB. Specificity of the nick-closing activity of bacteriophage T4 DNA ligase. Gene. 1989;76(2):245–254. [PubMed]
  • MacLean CJ, Morton NE. Estimation of myriad haplotype frequencies. Genet Epidemiol. 1985;2(3):263–272. [PubMed]
  • Riordan JR, Rommens JM, Kerem B, Alon N, Rozmahel R, Grzelczak Z, Zielenski J, Lok S, Plavsic N, Chou JL, et al. Identification of the cystic fibrosis gene: cloning and characterization of complementary DNA. Science. 1989 Sep 8;245(4922):1066–1073. [PubMed]
  • Crystal RG. The alpha 1-antitrypsin gene and its deficiency states. Trends Genet. 1989 Dec;5(12):411–417. [PubMed]
  • Robinson MA. Allelic sequence variations in the hypervariable region of a T-cell receptor beta chain: correlation with restriction fragment length polymorphism in human families and populations. Proc Natl Acad Sci U S A. 1989 Dec;86(23):9422–9426. [PMC free article] [PubMed]
  • Li Y, Szabo P, Robinson MA, Dong B, Posnett DN. Allelic variations in the human T cell receptor V beta 6.7 gene products. J Exp Med. 1990 Jan 1;171(1):221–230. [PMC free article] [PubMed]
  • Wilson RK, Lai E, Concannon P, Barth RK, Hood LE. Structure, organization and polymorphism of murine and human T-cell receptor alpha and beta chain gene families. Immunol Rev. 1988 Jan;101:149–172. [PubMed]
  • Lai E, Concannon P, Hood L. Conserved organization of the human and murine T-cell receptor beta-gene families. Nature. 1988 Feb 11;331(6156):543–546. [PubMed]
  • Charmley P, Chao A, Concannon P, Hood L, Gatti RA. Haplotyping the human T-cell receptor beta-chain gene complex by use of restriction fragment length polymorphisms. Proc Natl Acad Sci U S A. 1990 Jun;87(12):4823–4827. [PMC free article] [PubMed]
  • Gibbs RA, Nguyen PN, McBride LJ, Koepf SM, Caskey CT. Identification of mutations leading to the Lesch-Nyhan syndrome by automated direct DNA sequencing of in vitro amplified cDNA. Proc Natl Acad Sci U S A. 1989 Mar;86(6):1919–1923. [PMC free article] [PubMed]
  • Grompe M, Muzny DM, Caskey CT. Scanning detection of mutations in human ornithine transcarbamoylase by chemical mismatch cleavage. Proc Natl Acad Sci U S A. 1989 Aug;86(15):5888–5892. [PMC free article] [PubMed]
  • Odum N, Hyldig-Nielsen JJ, Morling N, Sandberg-Wollheim M, Platz P, Svejgaard A. HLA-DP antigens are involved in the susceptibility to multiple sclerosis. Tissue Antigens. 1988 May;31(5):235–237. [PubMed]
  • Beall SS, Concannon P, Charmley P, McFarland HF, Gatti RA, Hood LE, McFarlin DE, Biddison WE. The germline repertoire of T cell receptor beta-chain genes in patients with chronic progressive multiple sclerosis. J Neuroimmunol. 1989 Jan;21(1):59–66. [PubMed]
  • Seboun E, Robinson MA, Doolittle TH, Ciulla TA, Kindt TJ, Hauser SL. A susceptibility locus for multiple sclerosis is linked to the T cell receptor beta chain complex. Cell. 1989 Jun 30;57(7):1095–1100. [PubMed]
  • Urban JL, Kumar V, Kono DH, Gomez C, Horvath SJ, Clayton J, Ando DG, Sercarz EE, Hood L. Restricted use of T cell receptor V genes in murine autoimmune encephalomyelitis raises possibilities for antibody therapy. Cell. 1988 Aug 12;54(4):577–592. [PubMed]
  • Acha-Orbea H, Mitchell DJ, Timmermann L, Wraith DC, Tausch GS, Waldor MK, Zamvil SS, McDevitt HO, Steinman L. Limited heterogeneity of T cell receptors from lymphocytes mediating autoimmune encephalomyelitis allows specific immune intervention. Cell. 1988 Jul 15;54(2):263–273. [PubMed]
  • Zaller DM, Osman G, Kanagawa O, Hood L. Prevention and treatment of murine experimental allergic encephalomyelitis with T cell receptor V beta-specific antibodies. J Exp Med. 1990 Jun 1;171(6):1943–1955. [PMC free article] [PubMed]
  • Olson M, Hood L, Cantor C, Botstein D. A common language for physical mapping of the human genome. Science. 1989 Sep 29;245(4925):1434–1435. [PubMed]
  • Li HH, Gyllensten UB, Cui XF, Saiki RK, Erlich HA, Arnheim N. Amplification and analysis of DNA sequences in single human sperm and diploid cells. Nature. 1988 Sep 29;335(6189):414–417. [PubMed]
  • Cui XF, Li HH, Goradia TM, Lange K, Kazazian HH, Jr, Galas D, Arnheim N. Single-sperm typing: determination of genetic distance between the G gamma-globin and parathyroid hormone loci by using the polymerase chain reaction and allele-specific oligomers. Proc Natl Acad Sci U S A. 1989 Dec;86(23):9389–9393. [PMC free article] [PubMed]

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