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. 1990 Jun;46(6):1149–1155.

Theoretical underpinning of the single-molecule-dilution (SMD) method of direct haplotype resolution.

J C Stephens 1, J Rogers 1, G Ruano 1
PMCID: PMC1683843  PMID: 2339707

Abstract

In a recent paper we have shown that DNA haplotypes of multiply heterozygous individuals can be resolved directly by polymerase-chain-reaction (PCR) amplification of a single molecule of genomic template. Our method (the single-molecule-dilution [SMD] method) relies on the stochastic separation of maternal and paternal alleles at high dilution. The stochasticity of separation and the potential for DNA shearing (which could separate the loci of interest) are two factors that can compromise the results of the experiment. This paper explores the consequences of these two factors and shows that the SMD method can be expected to work very reliably even in the presence of a moderate amount of DNA shearing.

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Selected References

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

  1. Aquadro C. F., Desse S. F., Bland M. M., Langley C. H., Laurie-Ahlberg C. C. Molecular population genetics of the alcohol dehydrogenase gene region of Drosophila melanogaster. Genetics. 1986 Dec;114(4):1165–1190. doi: 10.1093/genetics/114.4.1165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Boehnke M., Arnheim N., Li H., Collins F. S. Fine-structure genetic mapping of human chromosomes using the polymerase chain reaction on single sperm: experimental design considerations. Am J Hum Genet. 1989 Jul;45(1):21–32. [PMC free article] [PubMed] [Google Scholar]
  3. Botstein D., White R. L., Skolnick M., Davis R. W. 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] [Google Scholar]
  4. Bowcock A. M., Bucci C., Hebert J. M., Kidd J. R., Kidd K. K., Friedlaender J. S., Cavalli-Sforza L. L. Study of 47 DNA markers in five populations from four continents. Gene Geogr. 1987 Apr;1(1):47–64. [PubMed] [Google Scholar]
  5. Clark A. G. Inference of haplotypes from PCR-amplified samples of diploid populations. Mol Biol Evol. 1990 Mar;7(2):111–122. doi: 10.1093/oxfordjournals.molbev.a040591. [DOI] [PubMed] [Google Scholar]
  6. Cui X. F., Li H. H., Goradia T. M., Lange K., Kazazian H. H., 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. doi: 10.1073/pnas.86.23.9389. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Kreitman M., Aguadé M. Genetic uniformity in two populations of Drosophila melanogaster as revealed by filter hybridization of four-nucleotide-recognizing restriction enzyme digests. Proc Natl Acad Sci U S A. 1986 May;83(10):3562–3566. doi: 10.1073/pnas.83.10.3562. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Kwok S., Higuchi R. Avoiding false positives with PCR. Nature. 1989 May 18;339(6221):237–238. doi: 10.1038/339237a0. [DOI] [PubMed] [Google Scholar]
  9. Li H. H., Gyllensten U. B., Cui X. F., Saiki R. K., Erlich H. A., Arnheim N. Amplification and analysis of DNA sequences in single human sperm and diploid cells. Nature. 1988 Sep 29;335(6189):414–417. doi: 10.1038/335414a0. [DOI] [PubMed] [Google Scholar]
  10. Maeda N., Bliska J. B., Smithies O. Recombination and balanced chromosome polymorphism suggested by DNA sequences 5' to the human delta-globin gene. Proc Natl Acad Sci U S A. 1983 Aug;80(16):5012–5016. doi: 10.1073/pnas.80.16.5012. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ruano G., Fenton W., Kidd K. K. Biphasic amplification of very dilute DNA samples via 'booster' PCR. Nucleic Acids Res. 1989 Jul 11;17(13):5407–5407. doi: 10.1093/nar/17.13.5407. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Saiki R. K., Gelfand D. H., Stoffel S., Scharf S. J., Higuchi R., Horn G. T., Mullis K. B., Erlich H. A. Primer-directed enzymatic amplification of DNA with a thermostable DNA polymerase. Science. 1988 Jan 29;239(4839):487–491. doi: 10.1126/science.2448875. [DOI] [PubMed] [Google Scholar]
  13. Schaeffer S. W., Aquadro C. F., Anderson W. W. Restriction-map variation in the alcohol dehydrogenase region of Drosophila pseudoobscura. Mol Biol Evol. 1987 May;4(3):254–265. doi: 10.1093/oxfordjournals.molbev.a040438. [DOI] [PubMed] [Google Scholar]
  14. Wainscoat J. S., Hill A. V., Boyce A. L., Flint J., Hernandez M., Thein S. L., Old J. M., Lynch J. R., Falusi A. G., Weatherall D. J. Evolutionary relationships of human populations from an analysis of nuclear DNA polymorphisms. Nature. 1986 Feb 6;319(6053):491–493. doi: 10.1038/319491a0. [DOI] [PubMed] [Google Scholar]

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