BRAF-Related Disorders via the BRAF 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
114 BRAF$990.00 81406 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
This test will detect causative mutations in ~75% of CFCS patients, ~5% of LS patients, and ~2% of NS patients.

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 BRAF$690.00 81479 Add to Order
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Over 100

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Turnaround Time

The great majority of tests are completed within 28 days.

Clinical Features
Cardio-Facio-Cutaneous Syndrome (CFCS, OMIM 115150) is a rare developmental disorder characterized by distinctive facial appearance; congenital cardiac and ectodermal abnormalities; postnatal growth failure; feeding difficulties with failure to thrive and neurological findings.  Facial features include high forehead, short, upturned nose with a low nasal bridge, prominent external ears that are posteriorly angulated and ocular hypertelorism.  The most common cardiac abnormalities include pulmonic  stenosis and atrial septal defects.  Ectodermal abnormalities are heterogeneous in features and severity.  They include café au lait spots, erythema, keratosis, ichthyosis, eczema, sparse and brittle hair, and nail dystrophy.  The neurological findings include seizures, hypotonia, macrocephaly and various degrees of mental and cognitive delay (Reynolds et al. Am J Med Genet 25:413-427, 1986).

Noonan Syndrome (NS, OMIM 163950) is a relatively common developmental disorder.  NS is characterized by dysmorphic facial features, growth and congenital heart defects, and musculoskeletal abnormalities.  Cardiac abnormalities are found in up to 80% of patients and include pulmonary valve stenosis, atrial septal defect, atrioventricular canal defect, and hypertrophic cardiomyopathy.  Musculoskeletal abnormalities include short stature, chest deformity with sunken or raised sternum, and short webbed neck.  Several additional abnormalities have been described and include renal, genital, hematological, neurologic, cognitive, behavioral, gastrointestinal, dental, and lymphatic findings.  Intelligence is usually normal; however, learning disabilities may be present.  NS is characterized by an extensive clinical heterogeneity, even among members of the same family.  Diagnosis is often made in infancy or early childhood.  Symptoms often change and lessen with advancing age.  Infants with NS are at risk of developing juvenile myelomonocytic leukemia (JMML OMIM 607785).  The prevalence of NS is estimated at 1 in 1000 to 1 in 2,500 births worldwide (Allanson et al. Am J Med Genet 21:507-514, 1985; Romano et al. Pediatrics 126:746-759, 2010). 

LEOPARD Syndrome (multiple Lentigines, Electrocardiographic-conduction abnormalities, Ocular hypertelorism, Pulmonary stenosis, Abnormal genitalia, Retardation of growth, sensorineural Deafness, OMIM 151100) is a rare congenital developmental disorder characterized by skin pigmentation anomalies including multiple lentigines and café au lait spots, hypertrophic cardiomyopathy, pulmonary valve stenosis, and deafness.  Other less common features include short stature, mild mental retardation, and abnormal genitalia (Legius et al. J Med Genet 39:571-574, 2002; Sarkozy et al. J Med Genet 41:e68, 2004). 
CFCS is caused by mutations in four genes within the RAS/MAPK pathway: BRAF, MAP2K1, MAP2K2 and KRAS (Rodriguez-Viciana et al. Science 311:1287-1290, 2006; Niihori et al. Nat Genet 38:294-296, 2006).  Over 37 BRAF mutations were detected in patients with CFCS.  They are the most common cause of CFCS, and account for ~75% of all cases genotyped.  Most causative mutations are missense resulting in amino acid substitutions.  A few small deletions have also been reported.  All mutations reported to date have been de novo.  The penetrance is complete in patients with CFCS (Rauen, GeneReviews, 2010).

NS is caused by gain of function mutations in various genes within the RAS/MAPK pathway, including  BRAF.  These mutations appear to activate the gene product (SHP2 protein).  To date, seven RAS/MAPK genes (PTPN11, SOS1, RAF1, KRAS, SHOC2, BRAF and NRAS) have been involved in patients with NS.  BRAF mutations were reported in ~2% of all NS cases genotyped (Allanson and Roberts, GeneReviews, 2011).  Only four different missense mutations were reported (Sarkozy et al. Hum Mutat 30:695-702, 2009).  Although de novo mutations are found in a substantial fraction of patients, familial cases have been reported.  In these families, NS is inherited in an autosomal dominant manner with variable expressivity (Romano, Pediatrics 126:746-759, 2010). 

LEOPARD Syndrome is caused by defects in three genes within the RAS/MAPK: PTPN11, RAF1 and BRAF (Digilio et al. Am J Hum Genet 71:389-394, 2002; Pandit et al. Nat Genet 39:1007-1012, 2007; Sarkozy, Hum Mutat 30: 95-702, 2009).  Unlike NS, LEOPARD Syndrome mutations act through a dominant negative  effect, which appears to disrupt the function of the wild-type gene Version 1.6   April 25, 2012 product (SHP2 protein) (Jopling et al. PLOS Genetics 3:e225, 2007).  Only two different BRAF missense mutations were reported in patients with LEOPARD syndrome (Sarkozy, 2009; Koudova et al. Eur J Med Genet 52:337-340, 2009).  Parents of LEOPARD patients are often asymptomatic, and de novo mutations are common.  However, familial cases have been reported.  In these families, affected relatives are diagnosed only after the birth of a visibly affected child, and the disease is transmitted in an autosomal dominant manner with variable penetrance and expressivity (Gelb and Tartaglia, GeneReviews, 2010). Genotype-phenotype correlations have been proposed (see for example Limongelli et al. Am J Med Genet A 146:620-628,2008).  Somatic BRAF mutations have been implicated in several human cancers.
Testing Strategy
BRAF testing for CFCS, NS and LS involves bidirectional DNA sequencing of all coding exons and splice sites of the BRAF gene.  The full coding sequence of each exon plus ~ 20 bp of flanking DNA on either side are sequenced.  We will also sequence and single exon (Test #100) in family members of patients with a known mutation or to confirm research results.
Indications for Test
Patients with clinical features of CFCS, NS and LS are candidates for this test. 

In addition to BRAF, PreventionGenetics also offers sequencing of each of the PTPN11, SOS1, RAF1, KRAS, SHOC2, NRAS, MAP2K1 and MAP2K2 genes individually, or in Panels.


Official Gene Symbol OMIM ID
BRAF 164757
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT


Genetic Counselors
  • Allanson JE, Hall JG, Hughes HE, Preus M, Witt RD. 1985. Noonan syndrome: the changing phenotype. Am. J. Med. Genet. 21: 507-514. PubMed ID: 4025385
  • Allanson, Judith E MD , Roberts, Amy E MD. (2011). "Noonan Syndrome." PubMed ID: 20301303
  • Digilio MC, Conti E, Sarkozy A, Mingarelli R, Dottorini T, Marino B, Pizzuti A, Dallapiccola B. 2002. Grouping of Multiple-Lentigines/LEOPARD and Noonan Syndromes on the PTPN11 Gene. The American Journal of Human Genetics 71: 389–394. PubMed ID: 12058348
  • Gelb BD, Tartaglia M. 2010. LEOPARD 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: 20301557
  • Jopling C, Geemen D van, Hertog J den. 2007. Shp2 knockdown and Noonan/LEOPARD mutant Shp2–induced gastrulation defects. PLoS genetics 3: e225. PubMed ID: 18159945
  • Koudova, M., (2009). "Novel BRAF mutation in a patient with LEOPARD syndrome and normal intelligence." Eur J Med Genet 52(5): 337-40. PubMed ID: 19416762
  • Legius E, Schrander-Stumpel C, Schollen E, Pulles-Heintzberger C, Gewillig M, Fryns J-P. 2002. PTPN11 mutations in LEOPARD syndrome. J. Med. Genet. 39: 571–574. PubMed ID: 12161596
  • Niihori T, Aoki Y, Narumi Y, Neri G, Cavé H, Verloes A, Okamoto N, Hennekam RCM, Gillessen-Kaesbach G, Wieczorek D, Kavamura MI, Kurosawa K, Ohashi H, Wilson L, Heron D, Bonneau D, Corona G, Kaname T, Naritomi K, Baumann C, Matsumoto N, Kato K, Kure S, Matsubara Y. 2006. Germline KRAS and BRAF mutations in cardio-facio-cutaneous syndrome. Nat. Genet. 38: 294–296. PubMed ID: 16474404
  • Pandit B, Sarkozy A, Pennacchio LA, Carta C, Oishi K, Martinelli S, Pogna EA, Schackwitz W, Ustaszewska A, Landstrom A, Bos JM, Ommen SR, Esposito G, Lepri F, Faul C, Mundel P, López Siguero JP, Tenconi R, Selicorni A, Rossi C, Mazzanti L, Torrente I, Marino B, Digilio MC, Zampino G, Ackerman MJ, Dallapiccola B, Tartaglia M, Gelb BD. 2007. Gain-of-function RAF1 mutations cause Noonan and LEOPARD syndromes with hypertrophic cardiomyopathy. Nature Genetics 39: 1007–1012. PubMed ID: 17603483
  • Rauen KA. 2010. Cardiofaciocutaneous 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: 20301365
  • Reynolds JF, Neri G, Herrmann JP, Blumberg B, Coldwell JG, Miles PV, Opitz JM. 1986. New multiple congenital anomalies/mental retardation syndrome with cardio-facio-cutaneous involvement--the CFC syndrome. Am. J. Med. Genet. 25: 413–427. PubMed ID: 3789005
  • Rodriguez-Viciana P, Tetsu O, Tidyman WE, Estep AL, Conger BA, Cruz MS, McCormick F, Rauen KA.. 2006. Germline Mutations in Genes Within the MAPK Pathway Cause Cardio-facio-cutaneous Syndrome. Science 311: 1287–1290. PubMed ID: 16439621
  • Romano AA, Allanson JE, Dahlgren J, Gelb BD, Hall B, Pierpont ME, Roberts AE, Robinson W, Takemoto CM, Noonan JA. 2010. Noonan syndrome: clinical features, diagnosis, and management guidelines. Pediatrics 126: 746-759. PubMed ID: 20876176
  • Sarkozy A, Carta C, Moretti S, Zampino G, Digilio MC, Pantaleoni F, Scioletti AP, Esposito G, Cordeddu V, Lepri F, Petrangeli V, Dentici ML, Mancini GM, Selicorni A, Rossi C, Mazzanti L, Marino B, Ferrero GB, Silengo MC, Memo L, Stanzial F, Faravelli F, Stuppia L, Puxeddu E, Gelb BD, Dallapiccola B, Tartaglia M. 2009. Germline BRAF mutations in Noonan, LEOPARD, and cardiofaciocutaneous syndromes: Molecular diversity and associated phenotypic spectrum. Human Mutation 30: 695–702. PubMed ID: 19206169
  • Sarkozy A, Conti E, Digilio MC, Marino B, Morini E, Pacileo G, Wilson M, Calabrò R, Pizzuti A, Dallapiccola B. 2004. Clinical and molecular analysis of 30 patients with multiple lentigines LEOPARD syndrome. Journal of Medical Genetics 41: e68–e68. PubMed ID: 15121796
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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 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.
  • 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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