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Ehlers-Danlos syndrome via the TNXB Gene, Exon 35

  • 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
877 TNXB$250.00 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

In a group of 151 patients with the classic, hypermobility, or vascular types of the Ehlers-Danlos syndrome, two patients (1.3%) were found to have the common contiguous 30kb deletion allele (i.e. nonfunctional chimera TNXA/TNXB gene that contains the 120bp deletion crossing exon 35 and intron 35) (Schalkwijk et al. 2001). In a cohort of 80 Dutch patients with hypermobility type Ehlers-Danlos syndrome, one patient (1.25%) was found to have the common contiguous 30kb deletion allele (i.e. nonfunctional chimera TNXA/TNXB gene that contains the 120bp deletion crossing exon 35 and intron 35) (Zweers et al. 2003). In a study of 192 patients with congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency (Merke et al. 2013), the common contiguous 30kb deletion allele (i.e. nonfunctional chimera TNXA/TNXB gene that contains the 120bp deletion crossing exon 35 and intron 35) was found in a heterozygous state in 12 (6.25%). 

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 TNXB$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 Features

Ehlers-Danlos syndrome, due to complete tenascin-X deficiency (OMIM# 606408) is an autosomal recessive disorder of connective tissues characterized by hyperextensible skin, hypermobile joints, and tissue fragility (Burch et al. 1997; Schalkwijk et al. 2001; Lindor et al. 2005; Bristow et al. 2005). The major clinical features of this condition resemble the classic type of Ehlers-Danlos syndrome except for lack of poor wound healing and atrophic scarring. Large joint dislocations are the most frequent debilitating finding and easy bruising is a prominent feature. Some patients were also affected by congenital adrenal hyperplasia due to 21-hydroxylase deficiency because these patients were homozygous for a contiguous 30kb deletion disrupting both the CYP21A2 and TNXB genes. Patients also have features that are not typically found in Ehlers-Danlos syndrome such as spina bifida occulta, mitral valve prolapse, gastrointestinal bleeding, severe diverticular intestinal disease with ruptured diverticula, pancolonic diverticulitis, rectal prolapse, premature arteriosclerosis, stroke and obstructive airway disease (Schalkwijk et al. 2001; Lindor et al. 2005).

Heterozygous TNXB deficiency has also been associated with hypermobility type Ehlers-Danlos syndrome (OMIM# 130020), which is an autosomal dominant disorder of connective tissues characterized by joint hypermobility often in association with joint subluxations and chronic musculoskeletal pain (Levy 2013; Zweers et al. 2003; Merke et al. 2013). Compared with other types of Ehlers-Danlos syndrome, the hypermobility type is more common but much less severe. Its clinical presentations are extremely variable and its molecular etiology is largely unknown. Tenascin-X haploinsufficiency has been reported to explain the causes in some patients, and TNXB is the only known gene to date associated with hypermobility type Ehlers-Danlos syndrome (Levy 2013; Zweers et al. 2003; Zweers et al. 2005). The contiguous 30kb deletion disrupting both CYP21A2 and TNXB genes is common in patients with congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency, and tenascin-X haploinsufficiency has been associated with Ehlers-Danlos syndrome features in those patients who are heterozygous for this contiguous deletion (Chen et al. 2009; Merke et al. 2013). In addition, cardiac abnormalities such as quadricuspid aortic valve have been reported in this group of CAH patients.

Genetics

Tenascin-X encoded by the TNXB gene is an extracellular matrix protein acting as an important regulator of collagen deposition in vivo (Bristow et al. 2005; Mao et al. 2002). Defects in the TNXB gene can cause both autosomal dominant and recessive type of Ehlers-Danlos syndrome (Schalkwijk et al. 2001; Zweers et al. 2003). TNXB is one of the constitutional genes of the RCCX module (RP-C4-CYP21-TNX) in the human leukocyte antigen histocompatibility complex on chromosome 6. The RCCX module represents a complicated genomic region featured by frequent homologous recombination events due to existence of highly homologous pseudogenes in tandem (Lee 2005; Chen et al. 2012). The pseudogene TNXA is a partially duplicated segment that corresponds to intron 31 to exon 44 of TNXB. A contiguous 30kb deletion disrupting both CYP21A2 and TNXB genes, resulting in a nonfunctional chimera TNXA/TNXB gene that contains the 120bp deletion crossing exon 35 and intron 35, is the most common pathogenic allele found in patients with Ehlers-Danlos syndrome due to tenascin-X deficiency (Schalkwijk et al. 2001; Koppens et al. 2002; Lindor et al. 2005; Merke et al. 2013). It is also the only known pathogenic chimera TNXA/TNXB gene to date. Other documented genetic defects in TNXB are missense substitutions and small indels (Human Gene Mutation Database). Pathogenic exon-level copy number changes have not been reported within the unique portion (exons 1 to 31) of TNXB.

Testing Strategy

This test involves bidirectional Sanger DNA sequencing of exon 35 of TNXB plus -25 and +50 bp of flanking non-coding DNA. In particular, this test can identify the nonfunctional chimera TNXA/TNXB gene that contains the 120bp deletion crossing exon 35 and intron 35. In most reported cases, this particular chimera allele represents the contiguous 30kb deletion disrupting both CYP21A2 and TNXB genes (Schalkwijk et al. 2001; Koppens et al. 2002; Lindor et al. 2005; Merke et al. 2013). However, it should be noted that such a contiguous deletion involving CYP21A2 should be confirmed via Sanger DNA sequencing of CYP21A2 (Test # 1419), which can determine a carrier status of congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency. Sequencing of the remaining exons of TNXB is available as a separate test (Test #843).

Indications for Test

Candidates for this test are patients with Ehlers-Danlos syndrome due to complete tenascin-X deficiency; or patients with hypermobility type Ehlers-Danlos syndrome (especially those have significant reduced serum tenascin-X level). This test is also for patients affected by congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency with clinical evidence of hypermobility type Ehlers-Danlos syndrome (Chen et al. 2009; Merke et al. 2013). Additionally, testing is indicated for family members of patients who are known to have the nonfunctional chimera TNXA/TNXB gene that contains the 120bp deletion crossing exon 35 and intron 35.

Gene

Official Gene Symbol OMIM ID
TNXB 600985
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Test

Name
Ehlers-Danlos syndrome via the TNXB Gene

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Bristow J, Carey W, Egging D, Schalkwijk J. 2005. Tenascin-X, collagen, elastin, and the Ehlers-Danlos syndrome. Am J Med Genet C Semin Med Genet 139C: 24–30. PubMed ID: 16278880
  • Burch GH, Gong Y, Liu W, Dettman RW, Curry CJ, Smith L, Miller WL, Bristow J. 1997. Tenascin-X deficiency is associated with Ehlers-Danlos syndrome. Nat. Genet. 17: 104–108. PubMed ID: 9288108
  • Chen W, Kim MS, Shanbhag S, Arai A, VanRyzin C, McDonnell NB, Merke DP. 2009. The phenotypic spectrum of contiguous deletion of CYP21A2 and tenascin XB: quadricuspid aortic valve and other midline defects. Am. J. Med. Genet. A 149A: 2803–2808. PubMed ID: 19921645
  • Chen W, Xu Z, Sullivan A, Finkielstain GP, Ryzin C Van, Merke DP, McDonnell NB. 2012. Junction site analysis of chimeric CYP21A1P/CYP21A2 genes in 21-hydroxylase deficiency. Clin. Chem. 58: 421–430. PubMed ID: 22156666
  • Human Gene Mutation Database (Bio-base).
  • Koppens PFJ, Hoogenboezem T, Degenhart HJ. 2002. Carriership of a defective tenascin-X gene in steroid 21-hydroxylase deficiency patients: TNXB-TNXA hybrids in apparent large-scale gene conversions. Hum. Mol. Genet. 11: 2581–2590. PubMed ID: 12354783
  • Lee H-H. 2005. Chimeric CYP21P/CYP21 and TNXA/TNXB genes in the RCCX module. Mol. Genet. Metab. 84: 4-8. PubMed ID: 15639189
  • Levy HP. 2013. Ehlers-Danlos Syndrome, Hypermobility Type. In: Pagon RA, Adam MP, Bird TD, Dolan CR, Fong C-T, and Stephens K, editors. GeneReviews™, Seattle (WA): University of Washington, Seattle. PubMed ID: 20301456
  • Lindor NM, Bristow J. 2005. Tenascin-X deficiency in autosomal recessive Ehlers-Danlos syndrome. Am. J. Med. Genet. A 135: 75–80. PubMed ID: 15793839
  • Mao JR, Taylor G, Dean WB, Wagner DR, Afzal V, Lotz JC, Rubin EM, Bristow J. 2002. Tenascin-X deficiency mimics Ehlers-Danlos syndrome in mice through alteration of collagen deposition. Nat. Genet. 30: 421–425. PubMed ID: 11925569
  • Merke DP, Chen W, Morissette R, Xu Z, Ryzin C Van, Sachdev V, Hannoush H, Shanbhag SM, Acevedo AT, Nishitani M, Arai AE, McDonnell NB. 2013. Tenascin-X haploinsufficiency associated with Ehlers-Danlos syndrome in patients with congenital adrenal hyperplasia. J. Clin. Endocrinol. Metab. 98: E379–387. PubMed ID: 23284009
  • Schalkwijk J, Zweers MC, Steijlen PM, Dean WB, Taylor G, Vlijmen IM van, Haren B van, Miller WL, Bristow J. 2001. A recessive form of the Ehlers-Danlos syndrome caused by tenascin-X deficiency. N. Engl. J. Med. 345: 1167–1175. PubMed ID: 11642233
  • Zweers MC, Bristow J, Steijlen PM, Dean WB, Hamel BC, Otero M, Kucharekova M, Boezeman JB, Schalkwijk J. 2003. Haploinsufficiency of TNXB is associated with hypermobility type of Ehlers-Danlos syndrome. Am. J. Hum. Genet. 73: 214–217. PubMed ID: 12865992
  • Zweers MC, Dean WB, Kuppevelt TH van, Bristow J, Schalkwijk J. 2005. Elastic fiber abnormalities in hypermobility type Ehlers-Danlos syndrome patients with tenascin-X mutations. Clin. Genet. 67: 330–334. PubMed ID: 15733269
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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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