Molecular Genetics Holoprosencephaly Panel

Disorders causing brain malformations are phenotypically heterogeneous; mutations in certain genes have been associated inherited conditions causing brain developmental issues.  Holoprosencephaly is a brain malformation condition whereby during development, the brain fails to divide properly into two hemispheres.  Individuals with holoprosencephaly display various birth defects including hypotelorism, facial dysmorphism, microcephaly or macrocephaly.  This NGS test assess for single nucleotide variations and small insertion and deletions in coding regions, however up to 50% of individuals with HPE are caused by chromosomal abnormalities detectable by microarray. The inheritance pattern for HPE can either be autosomal dominant or recessive.

Reasons for Referral:

• Suspected genetic etiology of a brain malformation
• Carrier testing
• Positive family history

Next generation sequencing will analyze the exons or coding regions of the 14 genes associated with holoprosencephaly listed below using Illumina NextSeq 500/550 technology. Samples are prepared using hybridization probes to enrich exonic regions.  This assay does not assess regions of insufficient coverage, introns and promoter regions; pseudogenes; where the reference genome is inaccurate or contains gaps and insertions; and regions of high GC or polynucleotide repeats, but may contain variants that impact gene function.

Exon-level deletion/duplication analysis is performed by running the NGS data through the Genome Analysis Toolkit (GATK) Germline Copy Number Variation best practices pipeline from GATK, version 4.1.4.1. A Bayesian model was validated clinically in our lab. The model can detect copy changes at a resolution of three (3) or more probe targets (exons) for deletions and duplications in genes that do not have pseudogenes, and is not designed to detect low-level mosaicism or balanced alterations.

Holoprosencephaly Panel (14 genes):
CDON, DISP1, DLL1, FGF8, FGFR1, FOXH1, GAS1, GLI2, NODAL, PTCH1, SHH, SIX3, TGIF1, ZIC2

 Blood: EDTA or ACD (Solution A or B):

• Adult: 5 mL
• Child: 5 mL
• Infant: 2-3 mL

Saliva: 2 ORAgene™ Saliva Collection Kits (OGR-500) used according to manufacturer instructions.  Please contact KDL Client Services for a Saliva Collection Kit for patients that cannot provide a blood sample.

Assisted Saliva: 4 ORAgene™ Assisted Saliva Collection Kits (OGR-575) used according to manufacturer instructions.  Please contact KDL Client Services for an Assisted Saliva Collection Kit for patients that cannot provide a blood sample.

Skin Fibroblast: Punch Biopsy (Cell cultures will be prepared at KDL and used for testing), or 2 T-25 confluent flasks

DNA: 5-10µg at 60-100ng/uL (DNA must be extracted in a CLIA-certified laboratory or a laboratory meeting equivalent requirements as determined by the CAP and/or CMS)

Prenatal:

Direct Amniotic Fluid (10-20mL)

Direct CVS

Direct POC

Cultured Amniocytes (2 T-25 flasks)

Cultured CVS (2 T-25 flasks)

Cultured Fetal Tissue: Product of Conception (2 T-25 flasks)

Cord Blood (1-2mL)

Notice Regarding Molecular Genetic Testing on CVS or Amniotic Fluid Specimens:

Maternal cell rule-out testing will be performed on all prenatal specimens received. Please provide maternal blood in addition to the fetal specimen. Additional charges apply for the maternal cell rule-out test.

For routine testing of blood and saliva (or DNA extracted from them), KDL does NOT accept samples from patients within two (2) weeks of a packed cell/platelet transfusion or within four (4) weeks of a whole blood transfusion. For extraordinary circumstances, where testing must be performed outside of the above windows, please contact our lab.

A REQUISITION FORM MUST ACCOMPANY ALL SAMPLES.  Please include detailed clinical information, including ethnicity, clinical history, and family history.

Weekly

8 weeks

• Package and ship specimen to remain cold, but not frozen. 
• Ship via overnight express, using the FedEx priority overnight label provided. 
• Contact Client Services for shipping kits and instructions at (855) 535-1522.