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Deafness, Autosomal Dominant 3B (DFNA3B) and Deafness, Autosomal Recessive 1B (DFNB1B) via the GJB6 Gene

  • Summary and Pricing
  • Clinical Features and Genetics
  • Citations
  • Methods
  • Ordering/Specimens
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TEST METHODS

Sequencing

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
1551 GJB6$490.00 81479 Add to Order
Targeted Testing

For ordering targeted known variants, please proceed to our Targeted Variants landing page.

Turnaround Time

The great majority of tests are completed within 18 days.

Clinical Sensitivity

The sensitivity of our sequencing assay for this gene is currently unknown. In total, only 18 GJB6 variants have been documented as pathogenic for hearing loss. However, since only missense variants have been described for this gene in association with DFNA3, we believe that our sequencing assay will be able to identify all of these variants. The only documented causative DFNB1 variants involving GJB6 are large deletions, which typically are not detected by this type of sequencing assay. For both DFNA3 and DFNB1, variants in the related gene GJB2 represent the vast majority of documented causative variants.

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 GJB6$690.00 81479 Add to Order
Pricing Comment

# of Genes Ordered

Total Price

1

$690

2

$730

3

$770

4-10

$840

11-30

$1,290

31-100

$1,670

Over 100

Call for quote

Turnaround Time

The great majority of tests are completed within 28 days.

Clinical Sensitivity

The sensitivity of our aCGH assay for this gene is currently unknown. In total, only 18 GJB6 variants have been documented as pathogenic for hearing loss. However, since only large deletions have been described for this gene in association with DFNB1, we believe that our aCGH assay will be able to identify a significant majority of these variants. The only documented causative DFNA3 variants involving GJB6 are missense variants, which typically are not detected by this type of aCGH assay. For both DFNA3 and DFNB1, variants in the related gene GJB2 represent the vast majority of documented causative variants.

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Clinical Features

Nonsyndromic hearing loss is characterized by difficulty or inability to hear that is not associated with any visible defects involving the external ear, other organs, or any other medical condition. Nonsyndromic hearing loss may be associated with abnormalities involving the middle ear and/or the inner ear (Ciuman 2013; Dodson et al. 2011; Hilgert et al. 2009). At least 70% of cases involving hearing loss are nonsyndromic (Van Camp et al. 1997).

Nonsyndromic hearing loss and deafness due to pathogenic variants in the GJB6 gene are associated with prelingual, progressive, mild-to-severe high-frequency sensorineural hearing impairment. Audioprofiles may vary significantly, even within a family. No vestibular or temporal bone abnormalities are typically found (Smith et al. 1993).

Genetics

The GJB6 gene encodes the gap junction protein beta 6, more commonly known as connexin 30. Connexins form gap junctions that facilitate metabolite and ion transport between adjacent cells. The role of connexin 30 in proper auditory function remains unclear, although it appears to be necessary for normal repair following sensory cell loss in the inner ear (Jagger and Forge 2015).

The different gene loci for nonsyndromic hearing loss are designated DFN (for DeaFNess) and named on the mode of inheritance; DFNA for autosomal dominant, DFNB for autosomal recessive and DFNX for X-linked inheritance, respectively. The GJB6 gene is located adjacent to the GJB2 gene in chromosomal region 13q12. GJB2 and GJB6 are both associated with nonsyndromic hearing loss inherited in both an autosomal dominant (DFNA3) and autosomal recessive (DFNB1) manner.

DFNA3 is caused by missense variants in either GJB2 (>90%) or GJB6 (<10%). DFNA3 has been further defined as DFNA3A (GJB2) and DFNA3B (GJB6). Both are characterized by prelingual, progressive, mild-to-severe high-frequency sensorineural hearing impairment, although DFNA3A is also reported to cause postlingual hearing loss (Smith et al. 1993). However, the association of GJB6 missense variants with DFNA3B remains uncertain due to limited evidence of dominant inheritance across multigenerational families (Grifa et al. 1999; Yang et al. 2007).

DFNB1 is caused by missense, nonsense, splicing and regulatory variants in GJB2 (~98%) or large deletions of GJB2 and/or upstream regions including GJB6 (~2%). DFNB1 has been further defined as DFNB1A (GJB2) and DFNB1B (GJB6). DFNB1 is characterized by congenital, non-progressive, mild to profound sensorineural hearing impairment. All DFNB1 documented pathogenic variants involving GJB6 are large deletions. Missense or nonsense variants in GJB6 have not been associated with DFNB1 (Smith et al. 1993).

Previous reports of individuals with hearing loss that were heterozygous for a variant in GJB2 and also heterozygous for a large deletion encompassing sequences upstream of GJB2 including all or part of GJB6 were thought to indicate digenic inheritance of DFNB1. However, subsequent experiments have shown that the mechanism of action of the deletions encompassing GJB6 is through altered GJB2 expression via loss of cis-regulatory elements in proximity to GJB6 (Rodriguez-Paris and Schrijver 2009; Rodriguez-Paris et al. 2011).

Aside from nonsyndromic hearing loss, pathogenic variants in the GJB6 gene can also cause Clouston syndrome or hidrotic ectodermal dysplasia, which is an autosomal dominant inherited skin disorder characterized by marked thickening of the palms and soles (palmoplantar hyperkeratosis), patchy or total hair loss (alopecia), thinning or thickening of nails (nail hypoplasia), as well as other nail deformities (Essenfelder et al. 2004).

Testing Strategy

This test involves bidirectional Sanger DNA sequencing of the single coding exon of GJB6. The entire coding region and ~20 bp of flanking non-coding DNA on either side of each splice site are sequenced. We will also sequence any portion (Test #100) or portions of the exon (Test #200) in family members of patients with known mutations or to confirm research results.

Indications for Test

Nonsyndromic hearing loss and deafness due to variants in GJB6 is suspected in individuals with the following: pre- or postlingual, mild to profound, progressive sensorineural hearing impairment; no related systemic findings identified by medical history and physical examination; or a family history of nonsyndromic hearing loss consistent with autosomal recessive or more rarely, autosomal dominant inheritance.

Gene

Official Gene Symbol OMIM ID
GJB6 604418
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Test

Name
Nonsyndromic Hearing Loss and Deafness Sequencing Panel

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Ciuman R.R. 2013. Medical Science Monitor. 19: 1195-210. PubMed ID: 24362017
  • Dodson K.M. et al. 2011. American Journal of Medical Genetics. Part A. 155A: 993-1000. PubMed ID: 21465647
  • Essenfelder G.M. et al. 2004. Human Molecular Genetics. 13: 1703-14. PubMed ID: 15213106
  • Grifa A. et al. 1999. Nature Genetics. 23: 16-8. PubMed ID: 10471490
  • Hilgert N. et al. 2009. Mutation Research. 681: 189-96. PubMed ID: 18804553
  • Jagger D.J., Forge A. 2015. Cell and Tissue Research. 360: 633-44. PubMed ID: 25381570
  • Rodriguez-Paris J. et al. 2011. Plos One. 6: e21665. PubMed ID: 21738759
  • Rodriguez-Paris J., Schrijver I. 2009. Biochemical and Biophysical Research Communications. 389: 354-9. PubMed ID: 19723508
  • Smith et al. 1993. Nonsyndromic Hearing Loss and Deafness, DFNA3. In: Pagon RA, Adam MP, Bird TD, Dolan CR, Fong C-T, Smith RJ, and Stephens K, editors. GeneReviews™, Seattle (WA): University of Washington, Seattle. PubMed ID: 20301708
  • Van Camp G. et al. 1997. American Journal of Human Genetics. 60: 758-64. PubMed ID: 9106521
  • Yang J.J. et al. 2007. Audiology & Neuro-otology. 12: 198-208. PubMed ID: 17259707
Order Kits
TEST METHODS

Bi-Directional Sanger Sequencing

Test Procedure

Nomenclature for sequence variants was from the Human Genome Variation Society (http://www.hgvs.org).  As required, DNA is extracted from the patient specimen.  PCR is used to amplify the indicated exons plus additional flanking non-coding sequence.  After cleaning of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit.  Products are resolved by electrophoresis on an ABI 3730xl capillary sequencer.  In most cases, sequencing is performed in both forward and reverse directions; in some cases, sequencing is performed twice in either the forward or reverse directions.  In nearly all cases, the full coding region of each exon as well as 20 bases of non-coding DNA flanking the exon are sequenced.

Analytical Validity

As of March 2016, we compared 17.37 Mb of Sanger DNA sequence generated at PreventionGenetics to NextGen sequence generated in other labs. We detected only 4 errors in our Sanger sequences, and these were all due to allele dropout during PCR. For Proficiency Testing, both external and internal, in the 12 years of our lab operation we have Sanger sequenced roughly 8,800 PCR amplicons. Only one error has been identified, and this was due to sequence analysis error.

Our Sanger sequencing is capable of detecting virtually all nucleotide substitutions within the PCR amplicons. Similarly, we detect essentially all heterozygous or homozygous deletions within the amplicons. Homozygous deletions which overlap one or more PCR primer annealing sites are detectable as PCR failure. Heterozygous deletions which overlap one or more PCR primer annealing sites are usually not detected (see Analytical Limitations). All heterozygous insertions within the amplicons up to about 100 nucleotides in length appear to be detectable. Larger heterozygous insertions may not be detected. All homozygous insertions within the amplicons up to about 300 nucleotides in length appear to be detectable. Larger homozygous insertions may masquerade as homozygous deletions (PCR failure).

Analytical Limitations

In exons where our sequencing did not reveal any variation between the two alleles, we cannot be certain that we were able to PCR amplify both of the patient’s alleles. Occasionally, a patient may carry an allele which does not amplify, due for example to a deletion or a large insertion. In these cases, the report contains no information about the second allele.

Similarly, our sequencing tests have almost no power to detect duplications, triplications, etc. of the gene sequences.

In most cases, only the indicated exons and roughly 20 bp of flanking non-coding sequence on each side are analyzed. Test reports contain little or no information about other portions of the gene, including many regulatory regions.

In nearly all cases, we are unable to determine the phase of sequence variants. In particular, when we find two likely causative mutations for recessive disorders, we cannot be certain that the mutations are on different alleles.

Our ability to detect minor sequence variants, due for example to somatic mosaicism is limited. Sequence variants that are present in less than 50% of the patient’s nucleated cells may not be detected.

Runs of mononucleotide repeats (eg (A)n or (T)n) with n >8 in the reference sequence are generally not analyzed because of strand slippage during PCR and cycle sequencing.

Unless otherwise indicated, the sequence data that we report are based on DNA isolated from a specific tissue (usually leukocytes). Test reports contain no information about gene sequences in other tissues.

Deletion/Duplication Testing Via Array Comparative Genomic Hybridization

Test Procedure

Equal amounts of genomic DNA from the patient and a gender matched reference sample are amplified and labeled with Cy3 and Cy5 dyes, respectively. To prevent any sample cross contamination, a unique sample tracking control is added into each patient sample. Each labeled patient product is then purified, quantified, and combined with the same amount of reference product. The combined sample is loaded onto the designed array and hybridized for at least 22-42 hours at 65°C. Arrays are then washed and scanned immediately with 2.5 µM resolution. Only data for the gene(s) of interest for each patient are extracted and analyzed.

Analytical Validity

PreventionGenetics' high density gene-centric custom designed aCGH enables the detection of relatively small deletions and duplications within a single exon of a given gene or deletions and duplications encompassing the entire gene. PreventionGenetics has established and verified this test's accuracy and precision.

Analytical Limitations

Our dense probe coverage may allow detection of deletions/duplications down to 100 bp; however due to limitations and probe spacing this cannot be guaranteed across all exons of all genes. Therefore, some copy number changes smaller than 100-300 bp within a targeted large exon may not be detected by our array.

This array may not detect deletions and duplications present at low levels of mosaicism or those present in genes that have pseudogene copies or repeats elsewhere in the genome.

aCGH will not detect balanced translocations, inversions, or point mutations that may be responsible for the clinical phenotype.

Breakpoints, if occurring outside the targeted gene, may be hard to define.

The sensitivity of this assay may be reduced when DNA is extracted by an outside laboratory.

Order Kits

Ordering Options


myPrevent - Online Ordering
  • The test can be added to your online orders in the Summary and Pricing section.
  • Once the test has been added log in to myPrevent to fill out an online requisition form.
REQUISITION FORM
  • A completed requisition form must accompany all specimens.
  • Billing information along with specimen and shipping instructions are within the requisition form.
  • All testing must be ordered by a qualified healthcare provider.

SPECIMEN TYPES
WHOLE BLOOD

(Delivery accepted Monday - Saturday)

  • Collect 3 ml -5 ml (5 ml preferred) of whole blood in EDTA (purple top tube) or ACD (yellow top tube). For Test #500-DNA Banking only, collect 10 ml -20 ml of whole blood.
  • For small babies, we require a minimum of 1 ml of blood.
  • Only one blood tube is required for multiple tests.
  • Ship blood tubes at room temperature in an insulated container. Do not freeze blood.
  • During hot weather, include a frozen ice pack in the shipping container. Place a paper towel or other thin material between the ice pack and the blood tube.
  • In cold weather, include an unfrozen ice pack in the shipping container as insulation.
  • At room temperature, blood specimen is stable for up to 48 hours.
  • If refrigerated, blood specimen is stable for up to one week.
  • Label the tube with the patient name, date of birth and/or ID number.

DNA

(Delivery accepted Monday - Saturday)

  • Send in screw cap tube at least 5 µg -10 µg of purified DNA at a concentration of at least 20 µg/ml for NGS and Sanger tests and at least 5 µg of purified DNA at a concentration of at least 100 µg/ml for gene-centric aCGH, MLPA, and CMA tests, minimum 2 µg for limited specimens.
  • For requests requiring more than one test, send an additional 5 µg DNA per test ordered when possible.
  • DNA may be shipped at room temperature.
  • Label the tube with the composition of the solute, DNA concentration as well as the patient’s name, date of birth, and/or ID number.
  • We only accept genomic DNA for testing. We do NOT accept products of whole genome amplification reactions or other amplification reactions.

CELL CULTURE

(Delivery preferred Monday - Thursday)

  • PreventionGenetics should be notified in advance of arrival of a cell culture.
  • Culture and send at least two T25 flasks of confluent cells.
  • Some panels may require additional flasks (dependent on size of genes, amount of Sanger sequencing required, etc.). Multiple test requests may also require additional flasks. Please contact us for details.
  • Send specimens in insulated, shatterproof container overnight.
  • Cell cultures may be shipped at room temperature or refrigerated.
  • Label the flasks with the patient name, date of birth, and/or ID number.
  • We strongly recommend maintaining a local back-up culture. We do not culture cells.
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