Connexin 26: GJB2 Sequencing

Clinical Information

The frequency of childhood deafness is estimated to be 1/500. Half of this hearing loss is genetic and approximately 80% of genetic hearing loss is nonsyndromic and inherited in an autosomal recessive manner. Approximately 50% of childhood nonsyndromic recessive hearing loss is caused by variants in the connexin 26 (Cx26) gene, GJB2, making it the most common form of autosomal recessive nonsyndromic hearing loss with a carrier rate estimated to be as high as 1 in 36 (2.8%). Newborns with confirmed hearing loss should have Cx26 testing. Cx26 testing will help define a group in which approximately 60% will have profound or severe-profound hearing loss requiring aggressive language intervention.

Technical Information

• This test is performed by Sanger sequencing.
• Genomic DNA is used to amplify the coding exon of the GJB2 gene by polymerase chain reaction, and the amplified product is sequenced bi-directionally, when possible, using the Sanger dideoxy method.
• Targeted sequencing analysis of the GJB2 non-coding exon 1 is also performed in order to detect the c.-3438C>T pathogenic variant in the basal promoter and the c.-3170G>A (IVS1+1G>A) donor splice site pathogenic variant (Denoyelle et al. (1999) Lancet 353, 1298-303).

Specimen Requirements

• The preferred sample type is 3-4 ml of peripheral blood collected in an EDTA (purple top) tube. Dried blood spots, extracted DNA, and saliva are also accepted for this test. Blood and saliva kits are available by request.
• Approximately 75 µL of blood should be applied to each of the five circles on a filter paper dried blood spot card. Allow blood to dry for at least 4 hours before shipping.
• Send approximately 5 µg of extracted DNA at a requested concentration of 90-130 ng/µl.
• Saliva samples must be submitted in an approved saliva kit.

Transport Instructions

• The blood specimen should be kept at room temperature and delivered via overnight shipping. If shipment is delayed by one or two days, the specimen should be refrigerated and shipped at room temperature. Do not freeze the specimen. Samples collected on Friday can be safely designated for Monday delivery.
• DBS cards should be shipped at room temperature.
• Extracted DNA should be sent at room temperature via overnight delivery.
• Saliva is stable at room temperature and can be delivered via overnight or ground shipping.