CHARGE Syndrome via the SEMA3E Gene

  • Summary and Pricing
  • Clinical Features and Genetics
  • Citations
  • Methods
  • Ordering/Specimens
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NGS Sequencing

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
4819 SEMA3E$640.00 81479 Add to Order
Pricing Comment

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 targeted known variants, please proceed to our Targeted Variants landing page.

Turnaround Time

The great majority of tests are completed within 26 days.

Clinical Sensitivity

Pathogenic variants in the SEMA3E gene appear to be a rare cause of CHARGE syndrome (Keyte and Hutson 2012; Janssen et al. 2012).

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Clinical Features

CHARGE syndrome is a severe developmental disorder characterized by multiple congenital defects involving sensory and mediastinal organs. It is a clinically heterogeneous disorder in regards to symptoms and severity. Hallmark features include ocular coloboma; choanal atresia; cranial nerve abnormalities leading to facial palsy, loss of sense of smell, feeding, swallowing and breathing difficulties; and external and inner ear malformations resulting in hearing loss and reduced sense of balance. Additional features include hypogonadotropic hypogonadism, which manifests as incomplete or absent puberty and infertility; genital hypoplasia; growth and developmental delay; a wide variety of heart defects; cleft lip or palate; and distinctive facial features. CHARGE syndrome is usually diagnosed during childhood. Diagnosis is made based on the presence of a combination of major and minor clinical features (Blake et al. 1998; Verloes et al. 2005). Magnetic resonance imaging (MRI) of the temporal bones reveals abnormalities in the semicircular canal (Amiel et al 2001). In rare cases, CHARGE syndrome has been detected in adult individuals only after the birth of a child with the major characteristic features of the disease (Hughes et al. 2014). It has also been diagnosed antenatally (Legendre et al. 2012). CHARGE syndrome affects individuals worldwide with an incidence of approximately 1 case in 12,500 live births (Källén et al. 1999). Higher incidences have been reported in the Atlantic provinces of Newfoundland and Labrador and the Maritime Provinces (Issekutz et al. 2005). See also (Lalani et al. 2012) and the CHARGE Syndrome Foundation (


CHARGE syndrome is an autosomal dominant condition that is most often caused by pathogenic variants in the CHD7 gene. Pathogenic variants in the SEMA3E gene appear to be a rare cause of CHARGE syndrome. To date, only two patients were reported to have defects in SEMA3E. They consist of a missense variant that is predicted to result in the amino acid change p.Ser703Leu; and a balanced translocation between chromosomes 2 and 7 that disrupts the SEMA3E gene (Lalani et al. 2004).

The semaphorin 3E protein is involved in the control of vascular patterning, which is critical for normal organogenesis (Gu et al. 2005).

Testing Strategy

For this NextGen test, the full coding regions plus ~10 bp of non-coding DNA flanking each exon are sequenced for the gene listed below. 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 any regions not captured or with insufficient number of sequence reads. All pathogenic, likely pathogenic, or variants of uncertain significance are confirmed by Sanger sequencing.

Indications for Test

Patients presenting with the major clinical criteria or a combination of minor and major criteria for CHARGE syndrome as described (Blake et al. 1998; Verloes et al. 2005) and no disease-causing variants in CHD7.


Official Gene Symbol OMIM ID
SEMA3E 608166
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT


Name Inheritance OMIM ID
CHARGE Association 214800

Related Tests

Autism Spectrum Disorders and Intellectual Disability (ASD-ID) Comprehensive Sequencing Panel with CNV Detection
CHARGE and Kallmann Syndromes via the CHD7 Gene


Genetic Counselors
  • Amiel J, Attieé-Bitach T, Marianowski R, Cormier-Daire V, Abadie V, Bonnet D, Gonzales M, Chemouny S, Brunelle F, Munnich A, Manach Y, Lyonnet S. 2001. Temporal bone anomaly proposed as a major criteria for diagnosis of CHARGE syndrome. Am. J. Med. Genet. 99: 124–127. PubMed ID: 11241470
  • Blake KD, Davenport SL, Hall BD, Hefner MA, Pagon RA, Williams MS, Lin AE, Graham JM Jr. 1998. CHARGE association: an update and review for the primary pediatrician. Clin Pediatr (Phila) 37: 159–173. PubMed ID: 9545604
  • CHARGE Syndrome Foundation
  • Gu C, Yoshida Y, Livet J, Reimert DV, Mann F, Merte J, Henderson CE, Jessell TM, Kolodkin AL, Ginty DD. 2005. Semaphorin 3E and plexin-D1 control vascular pattern independently of neuropilins. Science 307: 265–268. PubMed ID: 15550623
  • Hughes SS, Welsh HI, Safina NP, Bejaoui K, Ardinger HH. 2014. Family history and clefting as major criteria for CHARGE syndrome. American Journal of Medical Genetics Part A 164: 48–53. PubMed ID: 24214489
  • Issekutz KA, Graham JM, Prasad C, Smith IM, Blake KD. 2005. An epidemiological analysis of CHARGE syndrome: Preliminary results from a Canadian study. American Journal of Medical Genetics Part A 133A: 309–317. PubMed ID: 15637722
  • Janssen N, Bergman JEH, Swertz MA, Tranebjaerg L, Lodahl M, Schoots J, Hofstra RMW, Ravenswaaij‐Arts V, A CM, Hoefsloot LH. 2012. Mutation update on the CHD7 gene involved in CHARGE syndrome. Human Mutation 33: 1149–1160. PubMed ID: 22461308
  • Källén K, Robert E, Mastroiacovo P, Castilla EE, Källén B. 1999. CHARGE Association in newborns: a registry-based study. Teratology 60: 334–343. PubMed ID: 10590394
  • Kaplan JD, Bernstein JA, Kwan A, Hudgins L. 2010. Clues to an early diagnosis of Kallmann syndrome. Am. J. Med. Genet. A 152A: 2796–2801. PubMed ID: 20949504
  • Keyte A, Hutson MR. 2012. The Neural Crest in Cardiac Congenital Anomalies. Differentiation 84: 25–40. PubMed ID: 22595346
  • Lalani S, Safiullah A, Molinari L, Fernbach S, Martin D, Belmont J. 2004. SEMA3E mutation in a patient with CHARGE syndrome. J Med Genet 41: e94. PubMed ID: 15235037
  • Lalani SR, Hefner MA, Belmont JW, Davenport SL. 2012. CHARGE 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: 20301296
  • Legendre M, Gonzales M, Goudefroye G, Bilan F, Parisot P, Perez M-J, Bonnière M, Bessières B, Martinovic J, Delezoide A-L, Jossic F, Fallet-Bianco C, Bucourt M, Tantau J, Loget P, Loeuillet L, Laurent N, Leroy B, Salhi H, Bigi N, Rouleau C, Guimiot F, Quélin C, Bazin A, Alby C, Ichkou A, Gesny R, Kitzis A, Ville Y, Lyonnet S, Razavi F, Gilbert-Dussardier B, Vekemans M, Attié-Bitach T. 2012. Antenatal spectrum of CHARGE syndrome in 40 fetuses with CHD7 mutations. J. Med. Genet. 49: 698–707. PubMed ID: 23024289
  • Verloes A. 2005. Updated diagnostic criteria for CHARGE syndrome: A proposal. American Journal of Medical Genetics Part A 133A: 306–308. PubMed ID: 15666308
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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 ~20 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 covered by Sanger sequencing.  All pathogenic, likely pathogenic, or variants of uncertain significance are confirmed 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, Common 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 ~20 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.

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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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