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CHILDREN'S HOSPITAL & RESEARCH CENTER

Molecular Genetics Diagnostic Tests Menu

The following are descriptions of diagnostic molecular genetics tests available to qualified professionals in the medical community. For more information, call Kathleen Houtchens, PhD, Supervisor Molecular Diagnostics/Genetics at (510) 428-3623.

For Molecular Diagnostics, please see our Molecular Diagnostics page.

Link to Requisition Form

Array Comparative Genomic Hybridization
Fragile-X Syndrome
Prader-Willi Syndrome
Angelman Syndrome
Spinal Muscular Atrophy Types 1, 2, and 3
GJB2 (Connexin 26) Sequencing Test
GJB6 (Connexin 30) Deletion Assay
Factor V Leiden Thrombophilia
Prothrombin Thrombophilia
MTHFR Thermolabile Variant

Array Comparative Genomic Hybridization
Microarray Comparative Genomic Hybridization (array CGH) detects copy number changes at a much finer resolution than traditional karyotyping. Current guidelines recommend array CGH as a first-line test in the evaluation of individuals with:

  1. Multiple anomalies not specific to a well-delineated genetic syndrome
  2. Apparently non-syndromic developmental delay/intellectual disability
  3. Autism spectrum disorders

Each microarray contains 105,000 unique oligonucleotide probes designed by the International Standard Cytogenomic Array (ISCA) Consortium, a collaborative of laboratories promoting standardization of microarray platform design and results reporting. A portion of the ISCA array CGH probes cover the genome with an average probe spacing of 35 kb. The remaining probes are positioned where copy number changes are associated with disease, thus providing increased array resolution at genomic regions of increased clinical importance.

Fragile-X Syndrome
Fragile X syndrome is the most common inherited cause of mental retardation, occurring in 1/1000-1500 males and 1/2000-2500 females. Expansion in the number of CGG repeats in an exon of the FMR-1 gene is the causative mutation in ~99% of cases of Fragile X syndrome.

We screen all patients for CGG repeat size using a PCR assay. Male patients with large CGG repeats, and female patients with large repeats or a single size normal fragment, are further analyzed by Southern blotting with a FMR-1 probe.

Prader-Willi Syndrome
Prader-Willi syndrome is characterized by diminished fetal activity, obesity, muscular hypotonia, mental retardation, short stature, hypogonadotropic hypogonadism, and small hands & feet. Approximately 99% of Prader-Willi syndrome is caused either by a paternally derived deletion or by maternal uniparental disomy (UPD) on the proximal long arm of the chromosome 15.

We use a methylation-specific polymerase chain reaction assay (M-PCR) for the detection of Prader-Willi syndrome on patients' DNA that has undergone a chemical modification. M-PCR distinguishes methylated from unmethylated DNA at a CpG island of SNRPN, a gene associated with Prader-Willi syndrome. This method detects, but does not differentiate, deletions, UPD, and imprinting mutations. M-PCR does not detect rare mutations in the Prader-Willi DNA region.

Angelman Syndrome
Angelman syndrome is characterized by severe mental retardation, absent speech, puppet-like movements, inappropriate laughter, and epilepsy. Either a maternal deletion of 15q11-q13 or paternal uniparental disomy of chromosome 15 occurs in ~75-80% of patients with Angelman syndrome.

We use a methylation-specific polymerase chain reaction assay (M-PCR) for the detection of Angelman Syndrome on patients' DNA that has undergone a chemical modification. M-PCR distinguishes methylated from unmethylated DNA at a CpG island of SNRPN, a gene whose methylation status is associated with Angelman syndrome. This method detects, but does not differentiate, deletions, UPD, and imprinting mutations. M-PCR does not detect rare mutations in the Angelman Syndrome gene, UBE3 region.

Spinal Muscular Atrophy Types 1, 2, and 3
Spinal Muscular Atrophy (SMA) is an autosomal recessive disorder of the spinal and bulbar motor neurons causing muscle atrophy and weakness. With SMA, the facial muscles are spared and intellect is normal. The estimated SMA carrier frequency is 1 in 50.

Over 95-98% of SMA patients have homozygous deletions in exons 7 and/or 8 of the SMN1 gene. We use PCR to amplify exons 7 and 8 of SMN1 and the highly homologous SMN2 gene. The PCR products are then subjected to a restriction enzyme assay that specifically detects SMN1 deletions. This method cannot be used for carrier testing.

GJB2 (Connexin 26) Sequencing Test
Mutations in the GJB2 gene, encoding the connexin 26 protein, are the most common cause of nonsyndromic sensorineural hearing loss (NSNHL), accounting for up to 50% of all cases of prelingual NSNHL.

Connexin 26 is a full-gene sequencing assay to identify mutations in the GJB2 gene. In almost all cases of NSNHL, it is suggested that GJB2 sequencing be performed because it is substantially more common than any other genetic or nongenetic cause of NSNHL, and the audiological characteristics can be quite variable.


GJB6 (Connexin 30) Deletion Assay
A 342-kb deletion in the GJB6 gene, which encodes the connexin 30 protein, is the second most frequent mutation causing prelingual deafness. The deletion in homozygosity, or in combination with a single GJB2 (connexin 26) gene mutation, causes hearing loss. As such, any person who is found to be heterozygous for a GJB2 mutation, or with congenital severe to profound hearing loss and no clear cause, is a candidate for the GJB6 deletion test. A multiplex PCR reaction is performed to detect the presence of the GJB6 deletion.

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Factor V Leiden Thrombophilia
Inherited thrombophilia predisposes an individual to thrombotic events such as venous thrombosis, the third most common cardiovascular disease. Activated protein C (APC) resistance is regarded as the most prevalent coagulation abnormality associated with venous thrombosis. A single point mutation in the Factor V gene, referred to as the Factor V Leiden mutation, renders the protein partially resistant to inactivation by APC. Genetic analysis has demonstrated that this mutation, which has a relatively high prevalence in the general population (e.g., about 5% in Caucasians), accounts for 85% to 95% of APC resistance cases.

Melting curve analysis on amplified Factor V gene product is used to detect the presence of a wild-type allele and/or the Factor V Leiden allele.

Prothrombin Thrombophilia
The Prothrombin (Factor II) G20210A mutation results in elevated circulating levels of Prothrombin and risks of venous thrombosis similar to those of Factor V Leiden. The incidence of heterozygous carriers of this mutation is estimated to be 1.0-2.5% in individuals of European and African descent. The incidence is much lower in individuals of Asian heritage.

Melting curve analysis on amplified Factor II gene product is used to detect the presence of the wild-type and/or the mutant alleles.

MTHFR Thermolabile Variant
Methylene Tetrahydrofolate Reductase (MTHFR) is a key enzyme that converts 5,10-methylenetetrahydrofolate to 5-methylenetetrahydrofolate, which provides the methyl group for methylation of homocysteine to methionine. Hyperhomocysteinemia is a risk factor for arterial disease and venous thrombosis. The C677T variant in the MTHFR gene is associated with reduced enzyme activity and increased levels of plasma homocysteine. The MTHFR A1298C variant alone does not significantly affect plasma homocysteine, but may do so when combined with the C677T variant.

The C677T and A1298C variants are detected by multiplex amplification the MTHFR gene. The PCR products are subsequently digested by restriction to identify the presence of the wild-type and/or mutant alleles.

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Revised: Thursday, June 16, 2016 11:46 AM

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