Branchio-Oculo-Facial Syndrome (BOFS) via the TFAP2A Gene

  • Summary and Pricing
  • Clinical Features and Genetics
  • Citations
  • Methods
  • Ordering/Specimens
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Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
1672 TFAP2A$680.00 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

A genotype-phenotype analysis done in 30 families (41 affected individuals) identified TFAP2A mutations in 90% of the families (Milunsky et al. 2011).

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 TFAP2A$690.00 81479 Add to Order
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Turnaround Time

The great majority of tests are completed within 28 days.

Clinical Features

Branchio-Oculo-Facial Syndrome (BOFS) is a rare autosomal dominant congenital disorder with highly variable expression. BOFS is characterized by Branchial defects, which include erythematous cutaneous defects in the cervical region. Ocular anomalies are more diverse and include microphthalmia, anophthalmia, lacrimal duct obstruction, coloboma, cataract and ptosis. Facial defects include cleft lip and/or palate (CL/P), pseudocleft or abnormal philtrum (Al-Dosari et al. 2010; Aliferis et al. 2011; Günes et al. 2014). Other common findings are malformed and prominent pinnae and conductive hearing loss from inner ear and/or petrous bone anomalies. Less frequent findings are premature hair graying, ectopic thymus, renal anomalies and mild developmental delay (Lin and Milunsky 2011; Li et al. 2013). BOFS patients with predominant ocular phenotypes may go undiagnosed. Upper lip examination would be helpful in such cases. TFAP2A gene testing can be useful in confirming the diagnosis and also for better genetic counseling (Lin et al. 1991; Aliferis et al. 2011).


Autosomal dominant BOFS is caused by mutations in the TFAP2A (transcription factor AP-2α) gene. TFAP2A encoded AP-2α is a member of the AP2 transcription factor family, which is an essential regulator of mammalian cranial neural-tube closure and craniofacial development (Schorle et al. 1996; Zhang et al. 1996). The retinoic acid-inducible AP-2 family members (AP-2α, AP-2β, AP-2δ, AP-2γ, and AP-2ε) are shown to be expressed in several embryonic areas such as the forebrain, face and limb buds during mouse embryogenesis (Chazaud et al. 1996). Specifically, AP-2α is expressed in several eye tissues, including the developing lens, and is required in early ocular morphogenesis (West-Mays et al. 1999). AP-2α knockout mice studies have shown that loss of AP-2α in multiple tissues in the craniofacial region leads to severe optic cup patterning defects and failure of optic stalk morphogenesis. Tissue-specific deletion of AP-2α in the surface ectoderm, neural crest or retina did not recapitulate the phenotype, which indicates that tissue–tissue interactions are required for ocular morphogenesis (Bassett et al. 2010). To date, genetic heterogeneity has not been reported in BOFS. The majority of the TFAP2A causative mutations are missense (~90%) and act in a dominant-negative manner, which affect the DNA-binding domain activity of AP-2α. An analysis done in 30 families identified TFAP2A mutations in ~90% (27/30) of the families (Milunsky et al. 2011). Several mutations were recurrent and all mutations occurred in exons 4 and 5, which have been reported as hotspot regions for missense mutations (Milunsky et al. 2011). De novo mutations have also been reported in this region (Milunsky et al. 2008). A clear correlation between genotype and phenotype has not been seen in the cases analyzed to date (less than 100 cases). Complete penetrance, variable expressivity and marked clinical variability have been reported within the affected family members (Li et al. 2013). So far, about 35 total mutations (missense, small and gross deletion and insertions/duplications) have been reported in TFAP2A that are associated with BOFS (Human Gene Mutation Database).

Testing Strategy

This test involves bidirectional DNA Sanger sequencing of all coding exons and ~10 bp of flanking noncoding sequence of the TFAP2A gene. We will also sequence any single exon (Test #100) in family members of patients with a known mutation or to confirm research results.

Indications for Test

All patients with symptoms suggestive of Branchio-Oculo-Facial Syndrome (BOFS) are candidates.


Official Gene Symbol OMIM ID
TFAP2A 107580
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT


Name Inheritance OMIM ID
Branchiooculofacial Syndrome 113620

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Genetic Counselors
  • Al-Dosari MS, Almazyad M, Al-Ebdi L, Mohamed JY, Al-Dahmash S, Al-Dhibi H, Al-Kahtani E, Al-Turkmani S, Alkuraya H, Hall BD, others. 2010. Ocular manifestations of branchio-oculo-facial syndrome: report of a novel mutation and review of the literature. Molecular vision 16: 813. PubMed ID: 20461149
  • Aliferis K, Stoetzel C, Pelletier V, Hellé S, Angioï-Duprez K, Vigneron J, Leheup B, Marion V, Dollfus H. 2011. A novel TFAP2A mutation in familial Branchio-Oculo-Facial Syndrome with predominant ocular phenotype. Ophthalmic Genet. 32: 250–255. PubMed ID: 21728810
  • Bassett EA, Williams T, Zacharias AL, Gage PJ, Fuhrmann S, West-Mays JA. 2010. AP-2 knockout mice exhibit optic cup patterning defects and failure of optic stalk morphogenesis. Human Molecular Genetics 19: 1791–1804. PubMed ID: 20150232
  • Chazaud C, Oulad-Abdelghani M, Bouillet P, Décimo D, Chambon P, Dollé P. 1996. AP-2.2, a novel gene related to AP-2, is expressed in the forebrain, limbs and face during mouse embryogenesis. Mechanisms of development 54: 83–94. PubMed ID: 8808408
  • Günes N, Cengiz FB, Duman D, Dervişoğlu S, Tekin M, Tüysüz B. 2014. Branchio-oculo-facial syndrome in a newborn caused by a novel TFAP2A mutation. Genet. Couns. 25: 41–47. PubMed ID: 24783654
  • Human Gene Mutation Database (Bio-base).
  • Li H, Sheridan R, Williams T. 2013. Analysis of TFAP2A mutations in Branchio-Oculo-Facial Syndrome indicates functional complexity within the AP-2 DNA-binding domain. Human Molecular Genetics 22: 3195–3206. PubMed ID: 23578821
  • Lin AE, Milunsky JM. 2011. Branchiooculofacial Syndrome. In: Pagon RA, Adam MP, Ardinger HH, Bird TD, Dolan CR, Fong C-T, Smith RJ, and Stephens K, editors. GeneReviews(®), Seattle (WA): University of Washington, Seattle. PubMed ID: 21634087
  • Milunsky JM, Maher TA, Zhao G, Roberts AE, Stalker HJ, Zori RT, Burch MN, Clemens M, Mulliken JB, Smith R, Lin AE. 2008. TFAP2A Mutations Result in Branchio-Oculo-Facial Syndrome. The American Journal of Human Genetics 82: 1171–1177. PubMed ID: 18423521
  • Milunsky JM, Maher TM, Zhao G, Wang Z, Mulliken JB, Chitayat D, Clemens M, Stalker HJ, Bauer M, Burch M, Chénier S, Cunningham ML, et al. 2011. Genotype-phenotype analysis of the branchio-oculo-facial syndrome. Am. J. Med. Genet. A 155A: 22–32. PubMed ID: 21204207
  • Schorle H, Meier P, Buchert M, Jaenisch R, Mitchell PJ. 1996. Transcription factor AP-2 essential for cranial closure and craniofacial development. Nature 381: 235–238. PubMed ID: 8622765
  • West-Mays JA, Zhang J, Nottoli T, Hagopian-Donaldson S, Libby D, Strissel KJ, Williams T. 1999. AP-2alpha transcription factor is required for early morphogenesis of the lens vesicle. Dev. Biol. 206: 46–62. PubMed ID: 9918694
  • Zhang J, Hagopian-Donaldson S, Serbedzija G, Elsemore J, Plehn-Dujowich D, McMahon AP, Flavell RA, Williams T. 1996. Neural tube, skeletal and body wall defects in mice lacking transcription factor AP-2. Nature 381: 238–241. PubMed ID: 8622766
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Bi-Directional Sanger Sequencing

Test Procedure

Nomenclature for sequence variants was from the Human Genome Variation Society (  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 10 bases of non-coding DNA flanking the exon are sequenced.

Analytical Validity

As of February 2018, we compared 26.8 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 14 years of our lab operation we have Sanger sequenced roughly 14,300 PCR amplicons. Only one error has been identified, and this was an error in analysis of sequence data.

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 10 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.

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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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