Skip to main content
SpringerLink
Log in

Clinical Genetic Testing for Fragile X Syndrome by Polymerase Chain Reaction Amplification and Southern Blot Analyses

  • Protocol
  • First Online:
Fragile-X Syndrome

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1942))

Abstract

Fragile X syndrome (FXS) is characterized by mental retardation and in the vast majority of cases it is caused by expansion of CGG trinucleotide repeats in the 5′ untranslated region (or UTR) in the FMR1 gene on the X chromosome. The size and methylation status of CGG repeats are correlated with the clinical phenotype of FMR1-related disorders. The methods used for clinical genetic testing of FXS include polymerase chain reaction (PCR) amplification and Southern blot analyses, which effectively detect alleles with repeats in the normal, intermediate, premutation, and full mutation size ranges, as well as the methylation status of FMR1 promoter region.

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

Access this chapter

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 139.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Hardcover Book
USD 179.99
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

Institutional subscriptions

References

  1. Monaghan KG, Lyon E, Spector EB et al (2013) ACMG standards and guidelines for fragile X testing: a revision to the disease-specific supplements to the standards and guidelines for Clinical Genetics Laboratories of the American College of Medical Genetics and Genomics. Genet Med 15(7):575–586

    Article  CAS  Google Scholar 

  2. Turner G, Webb T, Wake S et al (1996) Prevalence of fragile X syndrome. Am J Med Genet 64(1):196–197

    Article  CAS  Google Scholar 

  3. Saul RA, Tarleton JC (1998) FMR1-related disorders. GeneReviews. https://www.ncbi.nlm.nih.gov/books/NBK1384/

    Google Scholar 

  4. Yu S, Pritchard M, Kremer E et al (1991) Fragile X genotype characterized by an unstable region of DNA. Science 252:1179–1181

    Article  CAS  Google Scholar 

  5. Tassone F, Pan R et al (2008) A rapid polymerase chain reaction-based screening method for identification of all expanded alleles of the fragile X (FMR1) gene in newborn and high-risk populations. J Mol Diagn 10(1):43–49

    Article  CAS  Google Scholar 

  6. Chen L, Hadd A et al (2010) An information-rich CGG repeat primed PCR that detects the full range of fragile X expanded alleles and minimizes the need for southern blot analysis. J Mol Diagn 12:589–600

    Article  CAS  Google Scholar 

  7. Filipovic-Sadic S, Sah S et al (2010) A novel FMR1 PCR method for the routine detection of low abundance expanded alleles and full mutations in fragile X syndrome. Clin Chem 56:399–408

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Hantash FM, Goos DG et al (2010) Qualitative assessment of FMR1 (CGG)n triplet repeat status in normal, intermediate, premutation, full mutation, and mosaic carriers in both sexes: implications for fragile X syndrome carrier and newborn screening. Genet Med 12(3):162–173

    Article  CAS  Google Scholar 

  9. Lyon E, Laver T et al (2010) A simple, high-throughput assay for fragile X expanded alleles using triple repeat primed PCR and capillary electrophoresis. J Mol Diagn 12(4):505–511

    Article  CAS  Google Scholar 

  10. Nahhas FA, Monroe TJ et al (2012) Evaluation of the human fragile X mental retardation 1 polymerase chain reaction reagents to amplify the FMR1 gene: testing in a clinical diagnostic laboratory. Genet Test Mol Biomarkers 16(3):187–192

    Article  CAS  Google Scholar 

  11. Das S, Kubota T et al (1998) Methylation analysis of the fragile X syndrome by PCR. Genet Test 1(3):151–155

    Article  CAS  Google Scholar 

  12. Panagopoulos I, Lassen C et al (1999) A methylation PCR approach for detection of fragile X syndrome. Hum Mutat 14(1):71–79

    Article  CAS  Google Scholar 

  13. Chen L, Hadd A et al (2011) High resolution methylation PCR for fragile X analysis: evidence for novel FMR1 methylation patterns undetected in southern blot analyses. Genet Med 13:528–538

    Article  Google Scholar 

  14. Grasso M, Boon EMJ et al (2014) A novel methylation PCR that offers standardized determination of FMR1 methylation and CGG repeat length without southern blot analysis. J Mol Diagn 16:23–31

    Article  CAS  Google Scholar 

  15. Southern E (2006) Southern blotting. Nat Protoc 1(2):518–525

    Article  CAS  Google Scholar 

  16. Rousseau F, Heitz D et al (1991) Direct diagnosis by DNA analysis of the fragile X syndrome of mental retardation. N Engl J Med 325(24):1673–1681

    Article  CAS  Google Scholar 

  17. Nakahori Y, Knight SJ et al (1991) Molecular heterogeneity of the fragile X syndrome. Nucleic Acids Res 19(16):4355–4359

    Article  CAS  Google Scholar 

  18. Fragile X CGG repeat genotyping GeneProber™ GLFXDig1 manual Catalog No. 40-2004-41

    Google Scholar 

  19. Amos Wilson J, Pratt VM et al (2008) Consensus characterization of 16 FMR1 reference materials: a consortium study. J Mol Diagn 10:2–12

    Article  Google Scholar 

  20. Hawkins M, Boyle J et al (2010) Preparation and validation of the first WHO international genetic reference panel for fragile X syndrome. Eur J Hum Genet 19:10–17

    Article  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, NY, USA

    Xiaoqiang Cai, Mohammad Arif, Haolei Wan, Ruth Kornreich & Lisa J. Edelmann

  2. Sema4, a Mount Sinai Venture, Stamford, CT, USA

    Xiaoqiang Cai, Haolei Wan, Ruth Kornreich & Lisa J. Edelmann

  3. WuXi AppTec Group, Shanghai, China

    Xiaoqiang Cai

Authors
  1. Xiaoqiang Cai
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mohammad Arif
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Haolei Wan
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ruth Kornreich
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Lisa J. Edelmann
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Lisa J. Edelmann .

Editor information

Editors and Affiliations

  1. Wolfe PGD Stem Cell Lab, Racine IVF Unit at Lis Maternity Hospital, Tel Aviv Sourasky Medical Center, Tel Aviv University, Tel Aviv, Israel

    Dalit Ben-Yosef

  2. Wolfe PGD Stem Cell Lab, Racine IVF Unit at Lis Maternity Hospital, Tel Aviv Sourasky Medical Center, Tel Aviv University, Tel Aviv, Israel

    Yoav Mayshar

Rights and permissions

Reprints and permissions

Copyright information

© 2019 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Cai, X., Arif, M., Wan, H., Kornreich, R., Edelmann, L.J. (2019). Clinical Genetic Testing for Fragile X Syndrome by Polymerase Chain Reaction Amplification and Southern Blot Analyses. In: Ben-Yosef, D., Mayshar, Y. (eds) Fragile-X Syndrome. Methods in Molecular Biology, vol 1942. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-9080-1_2

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-9080-1_2

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-9079-5

  • Online ISBN: 978-1-4939-9080-1

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation