Fragile X Syndrome

Genetic instability of triplet repeat sequences is an initial step in the etiology of hereditary neurodegenerative diseases and the cause of fragile X syndrome, the most common form of inherited mental retardation. This new paradigm of genetic disease, the transmission of expanded triplet repeat sequences, is associated with approximately 15 other human hereditary diseases.[6,7] Fragile X syndrome is caused by the progressive expansion of CGG repeats in the FMR1 gene, and there is a direct correlation between the number of CGG repeats and the frequency of inheriting an expanded CGG tract. The FMR1 gene in normal individuals harbors between 5 and 37 CGG repeats, and three other size categories are recognized that confer clinical information. Full mutations (>230 repeats) are associated with disease, whereas premutations (55-200 repeats) provide at-risk predictive information. The fourth category includes repeat lengths that provide inconclusive diagnostic and predictive information (45-54 repeats) and is generally referred to as the gray zone. Accurate measurements of the length of the CGG tract are critical to the clinical diagnostics community.

The National Institute of Standards and Technology (NIST) has established a quantitative measurement program for trinucleotide repeats[8] through the development of accurate protocols and reference materials for the analysis of the FMR1 gene after polymerase chain reaction (PCR) amplification of the CGG repeats. Amplification is a commonly used technique for FMR1 gene analysis and is faster than the traditionally used Southern blot. However, interlaboratory comparisons on the accuracy of amplifying CGG repeats have shown tremendous variation. This is because, in part, of the secondary structure properties of triplet repeats and the lack of standardized protocols. We have determined the metrics in need of standardization for fragile X repeat measurements and have reported the specific measurement variability associated with slab-PAGE and capillary electrophoresis (CE) separations, interlane variability, intercapillary variability, and the variability associated with the amplification of CGG repeat elements.

As reported,[8] the measurement errors associated with slab gels ranged from 0.05 to 0.35 standard deviation (SD). There was also little variation between gels and amplification reactions. The CE measurements were more precise with SD between 0.02 and 0.29. This measurement error is equivalent to one nucleotide of a single triplet repeat. The accuracy in determining the number of CGG repeats was in agreement for the normal and gray-zone length alleles using both slab-PAGE and CE. However, accuracy decreased for the measurement of larger alleles, where repeat size varied ± 3 repeats for measuring 85-95 repeats and ±4 repeats for measuring 105-110 repeats, our largest amplified allele. The size of the CGG repeat element in each sample was confirmed by automated fluorescent DNA sequencing.

Getting Started With Dumbbells

Getting Started With Dumbbells

The use of dumbbells gives you a much more comprehensive strengthening effect because the workout engages your stabilizer muscles, in addition to the muscle you may be pin-pointing. Without all of the belts and artificial stabilizers of a machine, you also engage your core muscles, which are your body's natural stabilizers.

Get My Free Ebook

Post a comment