Molecular Genetics Fragile X Syndrome

Fragile X syndrome is the most common form of inherited intellectual and developmental disabilities (IDDs).  Fragile X syndrome is the result of an expansion of a trinucleotide repeat (CGG) in the Fragile X Mental Retardation 1 (FMR1) gene located on the X chromosome.  Full size repeat expansions result in methylation of the FMR1 gene and the lack of a gene product.  Fragile X is inherited as an X-linked disorder and affects both males and females.  All mothers of males affected with fragile X syndrome are premutation carriers, and all full mutations arise from premutations.  Premutation carriers may be affected with Fragile X-associated Tremor Ataxia Syndrome (FXTAS), which is generally found in older premutation male carriers and consists of tremor and ataxia.  Premutation female carriers may be affected with FMR1-associated primary ovarian insufficiency (POI), the hallmark clinical feature being cessation of menses before age 40 years, and occasionally with tremor and ataxia.  

Fragile X-associated Tremor Ataxia syndrome (FXTAS) occurs in 40-45% of male FMR1 premutation carriers over the age of 50 and less frequently in female FMR1 premutation carriers. FXTAS is the result of a premutation, >54 trinucleotide (CGG) repeats, in the FMR1 gene. Core features of FXTAS are intention tremor and/or ataxia; peripheral neuropathy, autonomic dysfunction, and gradual cognitive decline may be present as well. Common psychiatric features include anxiety, dysinhibition, depression, and apathy.

Genetic counseling is recommended for all premutation and full mutation carriers.

Reason for Referral

• Testing is indicated for mentally retarded individuals and for individuals with a family history of mental retardation or learning disability of unknown etiology. 
• Testing is indicated for females with clinical features of early menopause, particularly those with a family history of Fragile X syndrome.

• Testing is indicated for individuals with clinical features of FXTAS, particularly with a family history of Fragile X syndrome.

PCR and capillary electrophoresis  analysis is used to measure the number of CGG trinucleotide repeats in the 5’ untranslated region of the FMR1 gene at Xq27.3; additionally, CGG repeat size and methylation status is determined by Southern blot for large premutations (>CGG₁₅₀) and for fully expanded alleles.

Allele Types #CGG Repeats:
       Normal: 5 - 44
       Intermediate: 45 - 54
       Premutation: 55 – 200
       Full mutation (i.e. disease-causing): >200

Blood: EDTA (purple-top) tube or ACD (yellow-top) tube

• Adult: 5 mL
• Child: 5 mL
• Infant: 2-3 mL

DNA: 20µg  at a minimum of 100ng/µL(DNA must be extracted in a CLIA-certified laboratory or a laboratory meeting equivalent requirements as determined by the CAP and/or CMS).

Saliva:2 ORAgene™ Saliva Collection Kits (OGR-500) used according to manufacturer instructions.  Please contact KDL Client Services for a Saliva Collection Kit for patients that cannot provide a blood sample.

Assisted Saliva: 4 ORAgene™ Assisted Saliva Collection Kits (OGR-575) used according to manufacturer instructions.  Please contact KDL Client Services for an Assisted Saliva Collection Kit for patients that cannot provide a blood sample.

Buccal Swab: Please contact KDL Client Services for Buccal Swab Collection Kit(s) for patients that cannot provide a blood sample.

Prenatal testing:

• Direct Amniotic Fluid (10-20mL)
• Direct CVS
• Direct POC
• Cultured Amniocytes (2-T25 flasks)
• Cultured CVS (2-T25 flasks)
• Cultured Fetal Tissue: Product of Conception (2 T25 flasks)
• Cord Blood (1-2mL)
• When testing CVS, sizing can be performed and expansions will be detected but methylation status may not be reliably determined. If methylation status is requested or required by the referring provider, testing of CVS is not recommended.

Notice Regarding Molecular Genetic Testing on Prenatal Specimens:

• Maternal cell rule-out testing will be performed on all prenatal specimens received. Please provide maternal blood (or saliva) in addition to the fetal specimen. Additional charges apply for the maternal cell rule-out test.

For routine testing of blood and saliva (or DNA extracted from them), KDL does NOT accept samples from patients within two (2) weeks of a packed cell/platelet transfusion or within four (4) weeks of a whole blood transfusion.  For extraordinary circumstances, where testing must be performed outside of the above windows, please contact our lab.

A REQUISITION FORM MUST ACCOMPANY ALL SAMPLES.  Please include detailed clinical information, including ethnicity, clinical history, and family history.

Weekly

7-10 days. If expansion found, add 14 days to turn around time.

• Package and ship specimen to remain cold. but not frozen.
• Contact Client Services for shipping kits and instructions. 
• Please use the cold pack provided in the KDL shipping kit. 
• Ship the specimen overnight express, using the FedEx priority overnight label provided. 

1. NCBI GeneReviews: FMR1-Related Disorders by Robert A. Saul and Jack C. Tarleton (2012) http://www.ncbi.nlm.nih.gov/books/NBK1384/
2. Monaghan et al. 2013 Genet Med 15(7)

Prior to any genetic testing we recommend genetic counseling. To receive forms and information about prenatal diagnostic testing, please contact Client Services at (855) 535-1522.