FOXC1-Related Disorders via the FOXC1 Gene

  • Summary and Pricing
  • Clinical Features and Genetics
  • Citations
  • Methods
  • Ordering/Specimens
Order Kits


Test Code Test Copy GenesPriceCPT Code Copy CPT Codes
7059 FOXC1$640.00 81479 Add to Order
Pricing Comments

Our most cost-effective testing approach is NextGen sequencing with Sanger sequencing supplemented as needed to ensure sufficient coverage and to confirm NextGen calls that are pathogenic, likely pathogenic or of uncertain significance. If, however, full gene Sanger sequencing only is desired (for purposes of insurance billing or STAT turnaround time for example), please see link below for Test Code, pricing, and turnaround time information.

For Sanger Sequencing click here.
Targeted Testing

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

Turnaround Time

The great majority of tests are completed within 20 days.

Clinical Sensitivity

Approximately 25%-60% of Axenfeld-Rieger syndrome cases are due to FOXC1 or PITX2 pathogenic variants (Tümer and Bach-Holm 2009; Reis et al. 2012; Alward 2000). In the remaining ~50% of ARS patients the genetic cause is unknown, which indicates that there are still more genes to be identified (Tümer and Bach-Holm 2009). Several gross deletions, duplications and complex rearrangements have been reported in FOXC1 that are associated with ARS and that are unlikely to be detected by sequencing (Human Gene Mutation Database).

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CNV via aCGH

Test Code Test Copy GenesPriceCPT Code Copy CPT Codes
600 FOXC1$990.00 81479 Add to Order
Pricing Comments

# of Genes Ordered

Total Price









Over 100

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Targeted Testing

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

Turnaround Time

The great majority of tests are completed within 20 days.

Clinical Features

Axenfeld-Rieger syndrome (ARS) is a rare, highly penetrant autosomal dominant disorder characterized by varying degrees of eye anterior segment anomalies with systemic malformations such as dental hypoplasia and a protuberant umbilicus (Hjalt and Semina 2005; Berry et al. 2006; Waldron et al. 2010; Tümer and Bach-Holm 2009; Chang et al. 2012). Dental abnormalities in this syndrome help in the diagnosis and to distinguish ARS from other eye anterior segment abnormalities. Early diagnosis of ARS through its dento-facial and systemic features is essential in treating or preventing the most serious consequence of ARS (O’Dwyer and Jones 2005). The major clinical concern is high risk of developing open-angle glaucoma, which represents the main challenge in terms of treatment. Approximately 50% of the ARS affected patients develop glaucoma (Shields et al. 1985; Alward 2000; Hjalt and Semina 2005; Chang et al. 2012). ARS affected patients also need surveillance and management of sensorineural hearing loss, and cardiac, endocrinological, craniofacial and orthopaedic defects (Chang et al. 2012).

Rieger syndrome (RIEG) and Axenfeld anomaly both are in the anterior chamber cleavage group of anomalies and are considered to be variations of a single developmental disorder and belong to the ARS group (Reese and Ellsworth 1966). RIEG is characterized by malformations of the eyes, teeth, and umbilicus; whereas, Axenfeld anomaly displays only ocular features (Amendt et al. 2000).


Linkage studies mapped four chromosomal loci (4q25, 6p25, 13q14 and 16q24) that are associated with ARS and related or overlapping phenotypes. FOXC1 (the forkhead box C1 gene, also known as FKHL7) is the causative gene at chromosome 6p25 (Alward 2000; Lines et al. 2002; Tümer and Bach-Holm 2009). FOXC1 causative mutations cause autosomal dominant ARS.

FOXC1 encodes a transcription factor that is expressed throughout eye ontogeny (Lines et al. 2002). This factor (FOXC1) is reported to maintain homeostasis in trabecular meshwork (TM) cells by regulating genes that play an important role in stress response (Ito et al. 2014; Paylakhi et al. 2013). TM helps in regulating intraocular pressure by acting as a drainage structure for aqueous humor (Tamm 2009). The mutations in FOXC1 that lead to FOXC1 dysfunction significantly decrease TM cell viability and subsequently contribute to the development of glaucoma. It's been reported that ~75% of ARS patients with FOXC1 mutations develop earlier-onset glaucoma (Ito et al. 2014; Paylakhi et al. 2013). A Genotype-Phenotype correlation study indicated that patients with FOXC1 duplications are at higher risk for development of glaucoma as compared to other FOXC1 mutations. PITX2 is the causative ARS gene at chromosome 4q25. PITX2 also encodes a transcription factor (PITX2). PITX2 and FOXC1 have been reported to interact with each other, which is essential for the regulation of common downstream target genes within specific cell lineages (Berry et al. 2006). Also, coinheritance of PITX2 and FOXC1 mutations has been reported in a family, which segregated with the disease and showed variable phenotypic expression. The most severely affected individual had mutations in both genes, whereas single heterozygous mutations caused milder ARS phenotypes (Kelberman et al. 2011). So far, over 100 mutations have been identified in FOXC1 that are associated with ARS and related phenotypes (Human Gene Mutation Database).

Testing Strategy

For this Next Generation Sequencing (NGS) test, sequencing is accomplished by capturing specific regions with an optimized solution-based hybridization kit, followed by massively parallel sequencing of the captured DNA fragments. Additional Sanger sequencing is performed for regions not captured or with insufficient number of sequence reads.

For Sanger sequencing, polymerase chain reaction (PCR) is used to amplify targeted regions. After purification of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit. PCR products are resolved by electrophoresis on an ABI 3730xl capillary sequencer. In nearly all cases, cycle sequencing is performed separately in both the forward and reverse directions.

This test provides full coverage of all coding exons of the FOXC1 gene, plus ~10 bases of flanking noncoding DNA. We define full coverage as >20X NGS reads or Sanger sequencing.

Indications for Test

All patients with symptoms suggestive of Axenfeld-Rieger syndrome and related phenotypes are candidates.


Official Gene Symbol OMIM ID
FOXC1 601090
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Tests

Axenfeld-Rieger Syndrome Sequencing Panel with CNV Detection
Glaucoma Sequencing Panel with CNV Detection
Neonatal Crisis Sequencing Panel with CNV Detection


Genetic Counselors
  • Alward WL. 2000. American Journal of Ophthalmology. 130: 107-15. PubMed ID: 11004268
  • Amendt BA. et al. 2000. Cellular and Molecular Life Sciences : Cmls. 57: 1652-66. PubMed ID: 11092457
  • Berry FB. et al. 2006. Human Molecular Genetics. 15: 905-19. PubMed ID: 16449236
  • Chang TC. et al. 2012. The British Journal of Ophthalmology. 96: 318-22. PubMed ID: 22199394
  • Hjalt TA., Semina EV. 2005. Expert Reviews in Molecular Medicine. 7: 1-17. PubMed ID: 16274491
  • Human Gene Mutation Database (Bio-base).
  • Ito YA. et al. 2014. Cell Death & Disease. 5: e1069. PubMed ID: 24556684
  • Kelberman D. et al. 2011. Human Mutatation. 32: 1144–52. PubMed ID: 21837767
  • Lines MA. et al. 2002. Human Molecular Genetics. 11: 1177–84. PubMed ID: 12015277
  • O'Dwyer E.M., Jones D.C. 2005. International Journal of Paediatric Dentistry. 15: 459-63. PubMed ID: 16238657
  • Paylakhi SH. et al. 2013. Experimental Eye Research. 111: 112-21. PubMed ID: 23541832
  • Reese AB., Ellsworth RM. 1966. Archives of Ophthalmology. 75: 307-18. PubMed ID: 5948260
  • Reis LM, Tyler RC, Volkmann Kloss BA, Schilter KF, Levin AV, Lowry RB, Zwijnenburg PJG, Stroh E, Broeckel U, Murray JC, Semina EV. 2012. PITX2 and FOXC1 spectrum of mutations in ocular syndromes. Eur. J. Hum. Genet. 20: 1224–1233. PubMed ID: 22569110
  • Shields MB. et al. 1985. Survey of Ophthalmology. 29: 387-409. PubMed ID: 3892740
  • Tamm ER. 2009. Experimental Eye Research. 88: 648-55. PubMed ID: 19239914
  • Tümer Z, Bach-Holm D. 2009. European Journal of Human Genetics : Ejhg. 17: 1527-39. PubMed ID: 19513095
  • Waldron JM. et al. 2010. Special Care in Dentistry : Official Publication of the American Association of Hospital Dentists, the Academy of Dentistry For the Handicapped, and the American Society For Geriatric Dentistry. 30: 218-22. PubMed ID: 20831741
Order Kits

NextGen Sequencing using PG-Select Capture Probes

Test Procedure

We use a combination of Next Generation Sequencing (NGS) and Sanger sequencing technologies to cover the full coding regions of the listed genes plus ~10 bases of non-coding DNA flanking each exon.  As required, genomic DNA is extracted from the patient specimen.  For NGS, patient DNA corresponding to these regions is captured using an optimized set of DNA hybridization probes.  Captured DNA is sequenced using Illumina’s Reversible Dye Terminator (RDT) platform (Illumina, San Diego, CA, USA).  Regions with insufficient coverage by NGS are often covered by Sanger sequencing.

For Sanger sequencing, Polymerase Chain Reaction (PCR) is used to amplify targeted regions.  After purification of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit.  PCR products are resolved by electrophoresis on an ABI 3730xl capillary sequencer.  In nearly all cases, cycle sequencing is performed separately in both the forward and reverse directions.

Patient DNA sequence is aligned to the genomic reference sequence for the indicated gene region(s). All differences from the reference sequences (sequence variants) are assigned to one of five interpretation categories, listed below, per ACMG Guidelines (Richards et al. 2015).

(1) Pathogenic Variants
(2) Likely Pathogenic Variants
(3) Variants of Uncertain Significance
(4) Likely Benign Variants
(5) Benign Variants

Human Genome Variation Society (HGVS) recommendations are used to describe sequence variants (  Rare variants and undocumented variants are nearly always classified as likely benign if there is no indication that they alter protein sequence or disrupt splicing.

Analytical Validity

As of March 2016, 6.36 Mb of sequence (83 genes, 1557 exons) generated in our lab was compared between Sanger and NextGen methodologies. We detected no differences between the two methods. The comparison involved 6400 total sequence variants (differences from the reference sequences). Of these, 6144 were nucleotide substitutions and 256 were insertions or deletions. About 65% of the variants were heterozygous and 35% homozygous. The insertions and deletions ranged in length from 1 to over 100 nucleotides.

In silico validation of insertions and deletions in 20 replicates of 5 genes was also performed. The validation included insertions and deletions of lengths between 1 and 100 nucleotides. Insertions tested in silico: 2200 between 1 and 5 nucleotides, 625 between 6 and 10 nucleotides, 29 between 11 and 20 nucleotides, 25 between 21 and 49 nucleotides, and 23 at or greater than 50 nucleotides, with the largest at 98 nucleotides. All insertions were detected. Deletions tested in silico: 1813 between 1 and 5 nucleotides, 97 between 6 and 10 nucleotides, 32 between 11 and 20 nucleotides, 20 between 21 and 49 nucleotides, and 39 at or greater than 50 nucleotides, with the largest at 96 nucleotides. All deletions less than 50 nucleotides in length were detected, 13 greater than 50 nucleotides in length were missed. Our standard NextGen sequence variant calling algorithms are generally not capable of detecting insertions (duplications) or heterozygous deletions greater than 100 nucleotides. Large homozygous deletions appear to be detectable.   

Analytical Limitations

Interpretation of the test results is limited by the information that is currently available.  Better interpretation should be possible in the future as more data and knowledge about human genetics and this specific disorder are accumulated.

When Sanger sequencing does not reveal any difference from the reference sequence, or when a sequence variant is homozygous, we cannot be certain that we were able to detect both patient alleles.  Occasionally, a patient may carry an allele which does not amplify, due to a large deletion or insertion.   In these cases, the report will contain no information about the second allele.  Our Sanger and NGS Sequencing tests are generally not capable of detecting Copy Number Variants (CNVs).

We sequence all coding exons for each given transcript, plus ~10 bp of flanking non-coding DNA for each exon.  Test reports contain no information about other portions of the gene, such as regulatory domains, deep intronic regions or any currently uncharacterized alternative exons.

In most 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 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.

Unless otherwise indicated, DNA sequence data is obtained from a specific cell-type (usually leukocytes from whole blood).   Test reports contain no information about the DNA sequence in other cell-types.

We cannot be certain that the reference sequences are correct.

Rare, low probability interpretations of sequencing results, such as for example the occurrence of de novo mutations in recessive disorders, are generally not included in the reports.

We have confidence in our ability to track a specimen once it has been received by PreventionGenetics.  However, we take no responsibility for any specimen labeling errors that occur before the sample arrives at PreventionGenetics.

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 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.

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.

In the case of duplications, aCGH will not determine the chromosomal location of the duplicated DNA. Most duplications will be tandem, but in some cases the duplicated DNA will be inserted at a different locus. This method will also not determine the orientation of the duplicated segment (direct or inverted).

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

The sensitivity of this assay is dependent upon the quality of the input DNA. In particular, highly degraded DNA will yield poor results.

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.
  • 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.


(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.


(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.


(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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