Skip to main content
SpringerLink
Log in
Book cover

Lipoprotein Protocols pp 1–34Cite as

PCR Methodology Applied to Genetic Studies of Lipoprotein Metabolism and Atherosclerosis

  • Protocol

  • 1010 Accesses

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 110))

Abstract

During the 1980s molecular genetics became widely applied to hpoprotein research. The cloning of candrdate genes for atherosclerosis (apohpoprotems, lipid-processing enzymes, lipid transfer proteins, and hpoprotem receptors) allowed the identrfication of mutations responstble for rare dysitpoproteinemtas, as well as more common genetic variants. The cloned candidate genes were used as probes on Southern blots to find polymorphic nucleottde substitutions at restriction enzyme sates (restriction fragment-length polymorphisms or RFLPs) that could be used as genetic markers in linkage and assoclatron studtes. Such molecular-genetic studies have shown that genetic variation is a component in individual variation in lipoprotein metabolism and rusk of atherosclerosis.

This is a preview of subscription content, log in via an institution.

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   109.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Saikl, R. K., Scharf, S., Faloona, F., Mullis, K., Horn, G T., Erltch, H A., and Arnhelm, N. (1985) Enzymatic amplification of β-globin genomlc sequences and restriction site analysis for diagnosis of sickle cell anemia. Science 230, 1350–1354.

    Article  Google Scholar 

  2. Innis, M A, Gelfand, D H, Smnsky, J. J., and White, T J (eds.) (1990) PCR Protocols. A Guide to Methods and Appltcations Academic, San Diego, CA

    Google Scholar 

  3. McPherson, M J, Quirke, P., and Taylor, G. R., (eds.) (1991) PCR A Practzcal Approach Oxford Uruverstty Press, New York.

    Google Scholar 

  4. Hallassos, A., Chomel, J. C, Tesson, L., Baudis, M, Kruh, J, Kaplan, J. C, and Knzis, A (1989) Modification of enzymatically amplified DNA for the detection of point mutations. Nucleic Acids Res 17, 3606.

    Article  Google Scholar 

  5. Htxson, J. E. and Vernier, D T (1990) Restriction isotyping of human apolipoprotein E by gene amplification and cleavage with HhaI J. Lipid Res 31, 545–548.

    Google Scholar 

  6. Davtgnon, J, Gregg, R. E., and Sing, C F. (1988) Apohpoprotein E polymorphism and atherosclerosis Arteriosclerosis 8, 1–21.

    Google Scholar 

  7. Tenkanen, H and Ehnholm, C (1993) Molecular biology of apohpoprotein A-IV. Curr. Oprn Lipidol 4, 95–99.

    Article  CAS  Google Scholar 

  8. McCombs, R. J, Marcadts, D. E., Ellis, J, and Wemberg, R. B. (1994) Attenuated hypercholesterolemic response to a high-cholesterol diet in subjects heterozygous for the apolipoprotein A-IV-2 allele N Engl J Med 331, 70 & 710

    Article  Google Scholar 

  9. Mata, P., Ordovas, J. M., Lopez-Miranda, J., Lichtenstem, A H, Clevidence, B, Judd, J T, and Schaefer, E J. (1994) ApoA-IV phenotype affects diet-induced plasma LDL cholesterol lowering. Arterioscler Thromb 14, 884–891

    PubMed  CAS  Google Scholar 

  10. Hixson, J E and Powers, P. K. (1991) Restriction isotyping of human apohpoprotem A-IV rapid typing of known tsoforms and detection of a new tsoform that deletes a conserved repeat. J Lipid Res 32, 1529–1535

    PubMed  CAS  Google Scholar 

  11. Sambrook, J, Fritsch, E F., and Mamatts, T (1989) Molecular Clonzng A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY

    Google Scholar 

  12. Hobbs, H H., Esser, V, and Russell, D W (1987) AvaII polymorphism in the human LDL receptor gene. Nucleic Acids Res 15, 379

    Article  PubMed  CAS  Google Scholar 

  13. Yamamoto, T, Davis, C G, Brown, M S., Schnetder, W. J., Casey, M L, Goldstein, J L., and Russell, D. W (1984) The human LDL receptor acystemerich protein with multiple Alu sequences in its mRNA Cell 39, 27–38.

    Article  PubMed  CAS  Google Scholar 

  14. D’Aqmla, R T., Bechtel, L J, Videler, J A, Eron, J J, Gorczyca, P, and Kaplan, J C (1991) Maximizing sensitivity and specificity of PCR by preamplification heating. Nucleic Acids Res 19, 3749.

    Article  Google Scholar 

  15. Mulhs, K B (1991) The polymerase chain reaction in an anemic mode. how to avoid cold ohgodeoxyribonuclear fusion PCR Methods Appl 1, 1–4.

    Google Scholar 

  16. Don, R. H., Cox, P. T., Wamwrtght, B J, Baker, K, and Mattick, J S (1991) Touchdown PCR to circumvent spurious priming during gene amplification Nucleic Acids Res 19, 4008.

    Article  PubMed  CAS  Google Scholar 

  17. Hecker, K H. and Roux, K H (1996) High and low annealing temperatures increase both specificity and yield in touchdown and stepdown PCR Biotechniques 20, 478–485

    PubMed  CAS  Google Scholar 

  18. Fischer, S G. and Lerman, L S (1983) DNA fragments differing by single basepair substitutions are separated in denaturing gradient gels: correspondence with melting theory Proc Natl Acad Sci USA 80, 1579–1583.

    Article  PubMed  CAS  Google Scholar 

  19. Wartell, R. M., Hosseini, S. H., and Moran, Jr, C P (1990) Detecting base pair substitutions in DNA fragments by temperature-gradient gel electrophoresis Nucleic Acids Res 18, 2699–2705

    Article  PubMed  CAS  Google Scholar 

  20. Ortta, M., Suzukt, Y., Sektya, T., and Hayasht, K (1989) Rapid and sensitive detection of point mutations and DNA polymorphtsms using the polymerase chain reaction. Genomics 5, 874–879.

    Article  Google Scholar 

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

    Article  PubMed  CAS  Google Scholar 

  22. Hence, K, Harders, J., Wiese, U, and Riesner, D. (1994) Temperature gradient gel electrophoresis (TGGE) for the detection of polymorphic DNA and RNA, Mol Biol in Protocols for Gene Analysis (Harwood, A. J., ed), Humana, Totowa, NJ, pp. 211–228

    Chapter  Google Scholar 

  23. Thibonnier, M. and Schork, N J. (1995) The genetics of hypertension. Curr Opm Genet Dev 5, 362–370.

    Article  CAS  Google Scholar 

  24. Jeunemattre, X., Soubrter, F, Kotelevtsev, Y V., Lifton, R. P., Williams, C S., Charm, A., Hunt, S., Hopkins, P. N., Williams, R R., Lalouel, J.-M., and Corvol, P (1992) Molecular basis of human hypertension role of angtotensmogen Cell 71, 169–180.

    Article  Google Scholar 

  25. Hixson, J. E. and Powers, P K (1995) Detection and charactertzation of new mutations in the human angiotensmogen gene (AGT) Human Genet 96, 110–112.

    Article  CAS  Google Scholar 

  26. Collins, T, Williams, A., Johnston, G. I., Kim, J., Eddy, R, Shows, T, Gtmbrone, Jr., M. A., and Bevilacqua, M. P (1991) Structure and chromosomal location of the gene for endothehal-leukocyte adhesion molecule 1 J Biol Chem 266, 2466–2473.

    PubMed  CAS  Google Scholar 

  27. Powers, P. K. and Hixson, J. E. (1993) BanI and PvuII polymorphisms im intion2 of selectin E (SELE). Human Mol Genet 2, 1082.

    Article  CAS  Google Scholar 

  28. Lerman, L. S. and Silverstem, K (1987) Computational simulation of DNA melting and its application to denaturing gradient gel electrophoresis. Methods Enzymol. 155, 482–501.

    Article  PubMed  CAS  Google Scholar 

  29. Steger, G. (1994) Thermal denaturation of double-stranded nucleic acids: prediction of temperatures critical for gradtent gel electrophoresis and polymerase cham reaction. Nucleic Acids Res 22, 2760–2768.

    Article  PubMed  CAS  Google Scholar 

  30. Fischer, S G and Lerman, L. S. (1979) Length-independent separation of DNA restriction fragments in two-dimensional gel electrophoresis. Cell 16, 19l–200.

    Article  Google Scholar 

  31. Parker, S., Angelico, M. C, Laffel, L., and Krolewski, A. S. (1993) Application of denaturing gradient gel electrophoresis to detect DNA sequence differences encoding apohpoprotein E isoforms. Genomics 16, 245–247

    Article  PubMed  CAS  Google Scholar 

  32. Hayashi, K. (1991) PCR-SSCP: a simple and sensitive method for detection of mutations in the genomic DNA. PCR Methods Appl 1, 34–38.

    PubMed  CAS  Google Scholar 

  33. Utermann, G (1989) The mysteries of lipoprotein(a). Science 246, 904–910

    Article  PubMed  CAS  Google Scholar 

  34. Cohen, J., Chiesa, G., and Hobbs, H. H. (1993) Sequence polymorphisms in the apolipoprotein(a) gene J Clin Invest. 91, 163 & 1636

    Google Scholar 

  35. Hongyo, T., Buzard, G. S., Calvert, R. J., and Weghorst, M. (1993) Cold SSCP. a simple, rapid and non-radtoactive method for optimized singlestrand conformation polymorphism analyses Nucleic Acids Res. 21, 3637–3642.

    Article  PubMed  CAS  Google Scholar 

  36. Vidal-Puig, A and Moller, D. E (1994) Comparative sensmvity of alternative single-strand conformation polymorphism (SSCP) methods Biotechniques 17, 490–496.

    PubMed  CAS  Google Scholar 

  37. Iwahana, H, Adzuma, K, Takahashi, Y., Katashima, R., Yoshimoto, K., and Itakura, M. (1995) Multiple fluorescence-based PCR-SSCP analysis with postlabeling PCR Methods Appl. 4, 275–282

    PubMed  CAS  Google Scholar 

  38. Wallace, R B., Shaffer, J., Murphy, R F., Bonner, J., Hirose, T, and Itakura, K (1979) Hybridization of synthetic oligdeoxyrtbonucleotides to ØDNA the effect of single base pair mismatch Nucleic Acids Res 6, 3543–3557

    Article  PubMed  CAS  Google Scholar 

  39. Saiki, R. K., Bugawan, T L, Horn, G T, Mulhs, K B, and Erlich, H. A. (1986) Analysis of enzymatically amplified β-globin and HLA-DQα DNA with allelespecific oligonucleotide probes. Nature 324, 163–166

    Article  PubMed  CAS  Google Scholar 

  40. Weisgraber, K H., Newhouse, Y. M., and Mahley, R W. (1988) Apolipoprotein E genotyping using the polymerase chain reaction and allelespecific oligonucleotide probes, Biochem Biophys Res Commun 157, 1212–1217.

    Article  PubMed  CAS  Google Scholar 

  41. Smeets, H J M, Poddighe, J, Stuyt, P M. J., Stalenhoef, A. F. H, Ropers, H H, and Wieringa, B (1988) Identification of apolipoprotein E polymorphism by using synthetic oligonucleottdes J Lipid Res 29, 1231–1237

    PubMed  CAS  Google Scholar 

  42. Houlston, R. S., Snowden, C, Green, F, Alberti, G M M, and Humphries, S. E. (1989) Apolipoprotein (apo) E genotypes by polymerase chain reaction and allele-specific oligonucleotide probes no detectable linkage disequilibrium between apo E and apo ll Human Genet. 83, 364–368

    Article  CAS  Google Scholar 

  43. Innerarity, T L, Mahley, R W, Weisgraber, K. H, Bersot, T. P., Krauss, R M, Vega, G L, Grundy, S M., Frtedl, W, Davignon, J., and McCarthy, J (1990) Familial defective apoliprotein B-100. a mutation of apolipoprotein B that causes hypercholesterolemia J Lipid Res 31, 1337–1349

    PubMed  CAS  Google Scholar 

  44. Ruztcka, V, Màrz, W, Russ, A., and Gross, W. (1992) Apolipoprotein B(Arg3500→Gln) allele specific polymerase chain reaction. large-scale screening of pooled blood samples. J Lipid Res 33, 1563–1567.

    Google Scholar 

  45. Hansen, P S, Rudtger, N, Tybjaerg-Hansen, A., Faergeman, O., and Gregersen, N (1991) Detection of the apoB-3500 mutation (glutamine for arginine) by gene amplification and cleavage with MspI J Lipid Res 32, 1229–1233

    PubMed  CAS  Google Scholar 

  46. Syvanen, A.-C., Aalto-Setala, K., Kontula, K., and Soderlund, H. (1989) Direct sequencing of affinity-captured amplified human DNA application to the detection of apolipoprotin E polymorphism FEB S Lett 258, 7l–74.

    Google Scholar 

  47. Sanger, F., Nicklen, S., and Coulcon, A. R. (1977) DNA sequencing with chamtermmating inhibitors Proc Natl Acad Sci USA 74, 5463–5467.

    Article  PubMed  CAS  Google Scholar 

  48. Syvánen, A.-C., Aalto-Setalå K., Harju, L., Kontula, K., and Soderlund, H (1990) A primer-guided nucleotide incorporation assay in the genotyping of apolipoprotein E Genomics 8, 684–692

    Article  PubMed  Google Scholar 

  49. Zuliani, G. and Hobbs, H. H. (1990) Dinucleotide repeat polymorphism at the 3’ end of the LDL receptor gene. Nucleic Acids Res. 18, 4300.

    Article  PubMed  CAS  Google Scholar 

  50. Zuliani, G. and Hobbs, H. H. (1990) Tetranucleotide repeat polymorphism in the apolipoprotein C-III gene Nucleic Acids Res 18 4299.

    Google Scholar 

  51. Zuliain, G and Hobbs, H H. (1990) Tetranucleotide repeat polymorphism in the LPL gene. Nucleic Acids Res 18 4958.

    Article  Google Scholar 

  52. Knott, T J, Wallis, S C, Pease, R J, Powell, L M, and Scott, J (1986) A hypervariable region 3’ to the human apohpoprotein B gene. Nucleic Acids Res. 14, 9215–9216

    Article  PubMed  CAS  Google Scholar 

  53. Huang, L.-S and Breslow, J L (1987) A unique AT-rich hypervarlable mmlsatellite 3’ to the apoB gene defines a high information restriction fragment length polymorphism J Biol Chem 262, 8952–8955.

    PubMed  CAS  Google Scholar 

  54. Zuham, G and Hobbs, H. H. (1990) Tetranucleotide repeat polymorphism in the apohpoprotein B gene Nucleic Acids Res 18 4299.

    Google Scholar 

  55. Ludwig, E H., Haubold, K., and McCarthy, B. J. (1991) Analysis of two different tandem repetitive elements within the human apohpoprotein B gene J Lipid Res 32, 374–379.

    PubMed  CAS  Google Scholar 

  56. Hixson, J E, Powers, P K, and McMahan, C A (1993) The human apolipoprotein B 3’ hypervariable region: detection of eight new alleles and comparisons of allele frequencies in blacks and whites. Human Genet 91, 475–479

    Article  CAS  Google Scholar 

  57. Litt, M., Hauge, X, and Sharma, V. (1993) Shadow bands seen when typing polymorphic dinucleotide repeats. some causes and cures Biotechniques 15, 280–284

    PubMed  CAS  Google Scholar 

  58. Mellersh, C. and Sampson, J (1993) Simplifying detection of microsatellite length polymorphisms. Biotechniques 15, 582–584

    PubMed  CAS  Google Scholar 

  59. Hauge, X Y and Litt, M (1993) A study of the origin of’ shadow bands’ seen when typing dinucleotide repeat polymorphisms by the PCR Human Mol Genet 2, 411–415

    Article  CAS  Google Scholar 

  60. Paterson, A. H., Lander, E. S., Hewitt, J. D., Peterson, S, Lincoln, S E, and Tanksley, S. D. (1988) Resolution of quantitative traits into Mendehan factors by using a complete linkage map of restriction fragment length polymorphisms Nature 335, 72l–726

    Article  Google Scholar 

  61. Lander, E. S. and Botstem, D. (1989) Mapping mendelian factors underlying quantitative traits using RFLP linkage maps Genetics 121, 185–199

    PubMed  CAS  Google Scholar 

  62. Cooperative Human Linkage Center (CHLC): Murray, J., Buetow, H., Weber, J. L., Ludwtgsen, S., Scherpbrer-Heddema, T., Mamon, F, Qurllen, J, Sheffield, V C, Sunden, S., Duyk, G. M.; Gènèthon: Welssenbach, J., Gyapay, G, Dtb, C., Morrissette, J, Lathrop, G. M., Vtgnal, A, Universlty of Utah White, R., Matsunamt, N., Gerken, S., Mehs, R., Albertsen, H, Plaetke, R., Odelberg, S; Yale University Ward, D.; Centre d’Etude du Polymorphrsme Humain (CEPH): Dausset, J., Cohen, D., and Cann, H. (1994) A comprehensive human linkage map with centtmorgan density. Science 265, 2049–2054

    Article  PubMed  CAS  Google Scholar 

  63. Weber, J L and May, P E. (1989) Abundant class of human DNA polymorphisms which can be typed using the polymerase chain reaction Am J Human Genet 44, 388–396

    CAS  Google Scholar 

  64. Edwards, A., Clvltello, A., Hammond, H A, and Caskey, C T (1991) DNA typing and genetic mapping with trimeric and tetrameric tandem repeats Am. J Hum Genet 49, 746756

    Google Scholar 

  65. Mansfield, D. C, Brown, A. F., Green, D. K., Carothers, A. D., Morris, S. W, Evans, H. J., and Wright, A. F. (1994) Automation of genetic linkage analysis using fluorescent microsatellite markers. Genomics 24, 225–233

    Article  PubMed  CAS  Google Scholar 

  66. Reed, P W., Davies, J L., Copeman, J., Bennett, S. T., Palmer, S. M., Pritchard, L E., Gough, S. C L, Kawaguchr, Y., Cordell, H J., Balfour, K. M., Jenkins, S., Powell, E. E., Vignal, A., and Todd, J. A.(1994) Chromosomespecific microsatellite sets for fluorescence-based, semi-automated genome mapping Nat Genet 7, 390–395.

    Article  PubMed  CAS  Google Scholar 

  67. Schwengel, D. A, Jedlicka, A. E, Nanthakumar, E. J, Weber, J L., and Levitt, R. (1994) Comparrson of fluorescence-based semi-automated genotyping of multiple microsatellite loct with autoradiographic techniques Genomics 22, 46–54.

    Article  PubMed  CAS  Google Scholar 

  68. Weissenbach, J., Gyapay, G., Drb, C., Vignal, A, Mortssette, J., Mtllasseau, P, Vaysseix, G, and Lathrop, M (1992) A second-generation linkage map of the human genome. Nature 359, 794–801

    Article  PubMed  CAS  Google Scholar 

  69. Scheftield, V C., Weber, J L., Buetow, K. H., Murray, J C, Even, D A, Wiles, K., Gastter, J. M., Pulido, J. C, Yandava, C., Sunden, S L, Mattes, G., Busmga, T, McClam, A., Beck, J, Scherprer, T, Grlliam, J, Zhong, J., and Duyk, G M (1995) A collection of tri-and tetranucleottde repeat markers used to generate high quality, high resolution human genome-wade linkage maps. Human Mol Genet 4, 1837–1844

    Article  Google Scholar 

  70. Perlin, M. W., Lancta, G., and Ng, S.-K. (1995) Toward fully automated genotyping. genotyping microsatellite markers by deconvolution. Am J Human Genet 57, 1199–1210

    CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Genetics, Southwest Foundation for Biomedical Research, San Antonio, TX

    Shelley A. Cole & James E. Hixson

Authors
  1. Shelley A. Cole
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. James E. Hixson
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Tufts University, Boston, MA

    Jose M. Ordovas

Rights and permissions

Reprints and permissions

Copyright information

© 1998 Humana Press Inc., Totowa, NJ

About this protocol

Cite this protocol

Cole, S.A., Hixson, J.E. (1998). PCR Methodology Applied to Genetic Studies of Lipoprotein Metabolism and Atherosclerosis. In: Ordovas, J.M. (eds) Lipoprotein Protocols. Methods in Molecular Biology™, vol 110. Humana Press. https://doi.org/10.1385/1-59259-582-0:1

Download citation

  • DOI: https://doi.org/10.1385/1-59259-582-0:1

  • Publisher Name: Humana Press

  • Print ISBN: 978-0-89603-420-4

  • Online ISBN: 978-1-59259-582-2

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation