Skip to main content
PMC home page
Molecular Pathology : MP logoLink to Molecular Pathology : MP
. 1999 Oct;52(5):302–304. doi: 10.1136/mp.52.5.302

Variability in the interpretation of microsatellite patterns with different electrophoretic conditions.

A C Santos 1, Y Yamaoka 1, D Y Graham 1, A R Sepulveda 1
PMCID: PMC395715  PMID: 10748882

Abstract

Microsatellite markers permit the analysis of microsatellite instability and loss of heterozygosity. Frequently, the allelotypes of microsatellites are interpreted in the presence of numerous bands in gels. The importance of different gel electrophoresis conditions in the interpretation of microsatellite patterns was tested. Microsatellite markers were used to amplify DNA from gastric cancer samples and adjacent gastric mucosa. Polymerase chain reaction (PCR) products were separated by electrophoresis through 7% polyacrylamide gels containing either 5.6 M urea and 32% formamide or 7 M urea. PCR reactions separated on urea/formamide gels resulted consistently in clear allele definition (one or two bands), whereas 7 M urea gels resulted in allele patterns that comprised multiple bands. Analysis of microsatellite abnormalities using nonformamide gels gave false negative results in just under a third of cases (four of 13). In conclusion, the interpretation of microsatellite alterations in cancer DNA is improved by using electrophoresis conditions that result in complete DNA denaturation, such as urea/formamide/acrylamide gel electrophoresis.

Full Text

The Full Text of this article is available as a PDF (126.2 KB).

Selected References

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

  1. Balas A., Santos S., García-Sánchez F., Lillo R., Merino J. L., Vicario J. L. Complete coding sequence of HLA-B*2712: a serologic B27-negative antigen associated to Bw6. Tissue Antigens. 1998 Apr;51(4 Pt 1):394–397. doi: 10.1111/j.1399-0039.1998.tb02980.x. [DOI] [PubMed] [Google Scholar]
  2. Bein G., Gläser R., Kirchner H. Rapid HLA-DRB1 genotyping by nested PCR amplification. Tissue Antigens. 1992 Feb;39(2):68–73. doi: 10.1111/j.1399-0039.1992.tb01909.x. [DOI] [PubMed] [Google Scholar]
  3. Brewerton D. A., Hart F. D., Nicholls A., Caffrey M., James D. C., Sturrock R. D. Ankylosing spondylitis and HL-A 27. Lancet. 1973 Apr 28;1(7809):904–907. doi: 10.1016/s0140-6736(73)91360-3. [DOI] [PubMed] [Google Scholar]
  4. Chung D. C., Rustgi A. K. DNA mismatch repair and cancer. Gastroenterology. 1995 Nov;109(5):1685–1699. doi: 10.1016/0016-5085(95)90660-6. [DOI] [PubMed] [Google Scholar]
  5. Hamamoto T., Yokozaki H., Semba S., Yasui W., Yunotani S., Miyazaki K., Tahara E. Altered microsatellites in incomplete-type intestinal metaplasia adjacent to primary gastric cancers. J Clin Pathol. 1997 Oct;50(10):841–846. doi: 10.1136/jcp.50.10.841. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Hayden J. D., Martin I. G., Cawkwell L., Quirke P. The role of microsatellite instability in gastric carcinoma. Gut. 1998 Feb;42(2):300–303. doi: 10.1136/gut.42.2.300. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Lin J. T., Wu M. S., Shun C. T., Lee W. J., Wang J. T., Wang T. H., Sheu J. C. Microsatellite instability in gastric carcinoma with special references to histopathology and cancer stages. Eur J Cancer. 1995 Oct;31A(11):1879–1882. doi: 10.1016/0959-8049(95)00349-n. [DOI] [PubMed] [Google Scholar]
  8. Litt M., Hauge X., Sharma V. Shadow bands seen when typing polymorphic dinucleotide repeats: some causes and cures. Biotechniques. 1993 Aug;15(2):280–284. [PubMed] [Google Scholar]
  9. Liu B., Parsons R. E., Hamilton S. R., Petersen G. M., Lynch H. T., Watson P., Markowitz S., Willson J. K., Green J., de la Chapelle A. hMSH2 mutations in hereditary nonpolyposis colorectal cancer kindreds. Cancer Res. 1994 Sep 1;54(17):4590–4594. [PubMed] [Google Scholar]
  10. Martorell L., Johnson K., Boucher C. A., Baiget M. Somatic instability of the myotonic dystrophy (CTG)n repeat during human fetal development. Hum Mol Genet. 1997 Jun;6(6):877–880. doi: 10.1093/hmg/6.6.877. [DOI] [PubMed] [Google Scholar]
  11. Meltzer S. J., Yin J., Manin B., Rhyu M. G., Cottrell J., Hudson E., Redd J. L., Krasna M. J., Abraham J. M., Reid B. J. Microsatellite instability occurs frequently and in both diploid and aneuploid cell populations of Barrett's-associated esophageal adenocarcinomas. Cancer Res. 1994 Jul 1;54(13):3379–3382. [PubMed] [Google Scholar]
  12. Neumüller J., Fischer M., Eberl R. Failure of the serological determination of HLA-B27 due to antigen masking in patients with ankylosing spondylitis. Rheumatol Int. 1993;13(4):163–167. doi: 10.1007/BF00301265. [DOI] [PubMed] [Google Scholar]
  13. Neumüller J., Schwartz D. W., Dauber E., Mayr W. R. Evaluation of four monoclonal antibodies against HLA-B27 for their reliability in HLA-B27 typing with flow cytometry (FC): comparison with the classic microlymphocytotoxic test (MLCT). Cytometry. 1996 Sep 15;26(3):209–215. doi: 10.1002/(SICI)1097-0320(19960915)26:3<209::AID-CYTO5>3.0.CO;2-9. [DOI] [PubMed] [Google Scholar]
  14. Olerup O. HLA-B27 typing by a group-specific PCR amplification. Tissue Antigens. 1994 Apr;43(4):253–256. doi: 10.1111/j.1399-0039.1994.tb02334.x. [DOI] [PubMed] [Google Scholar]
  15. Panaccio M., Georgesz M., Lew A. M. FoLT PCR: a simple PCR protocol for amplifying DNA directly from whole blood. Biotechniques. 1993 Feb;14(2):238–243. [PubMed] [Google Scholar]
  16. Shinmura K., Tani M., Isogaki J., Wang Y., Sugimura H., Yokota J. RER phenotype and its associated mutations in familial gastric cancer. Carcinogenesis. 1998 Feb;19(2):247–251. doi: 10.1093/carcin/19.2.247. [DOI] [PubMed] [Google Scholar]
  17. Steffens-Nakken H. M., Zwart G., van den Bergh F. A. Validation of allele-specific polymerase chain reaction for DNA typing of HLA-B27. Clin Chem. 1995 May;41(5):687–692. [PubMed] [Google Scholar]
  18. Weissenbach J., Gyapay G., Dib C., Vignal A., Morissette J., Millasseau P., Vaysseix G., Lathrop M. A second-generation linkage map of the human genome. Nature. 1992 Oct 29;359(6398):794–801. doi: 10.1038/359794a0. [DOI] [PubMed] [Google Scholar]

Articles from Molecular Pathology are provided here courtesy of BMJ Publishing Group

RESOURCES