Next Gen Sequencing of 16 (Non-NF1) RASopathy related genes and Deletion/Duplication analysis (NNP-NG)
Acceptable specimen types:
- Blood (2-3ml EDTA; no time limitations associated with receipt)
- Saliva (OGR-575 DNA Genotek; kits are provided upon request),
- DNA (extracted from lymphocyte cells, a minimum of 3μg, O.D. value at 260:280nm ≥1.8)
TURNAROUND TIME
- 25 working days
Candidates for this test:
- Patients with clinical features suggestive of either NS, NSML, CFC, Legius syndrome or Noonan-like syndrome with no mutation previously found by comprehensive RNA-based NF1+/- SPRED1 testing. patients with a clinical diagnosis of any of these syndromes that previously tested negative in a subset of the genes included in this panel; patients with a diagnosis of Costello syndrome but no HRAS mutation previously identified.
TEST DESCRIPTION
The Next Gen Sequencing of 16 RASopathy related genes and Deletion/Duplication analysis (RAS-NG) involves the simultaneous sequencing of 16 genes: SPRED1, PTPN11, PPP1CB, BRAF, CBL, HRAS, KRAS, NRAS, MAP2K1, MAP2K2, RAF1, RIT1, RASA2, SHOC2, SOS1 and SOS2. The test uses a customized and optimized set of Agilent Haloplex capture probes, followed by sequencing of overlapping amplicons within the regions of interest using Illumina sequencing chemistry. Each coding exon plus ~50bp of flanking intronic sequence are simultaneously sequenced. 5’ and 3’ untranslated sequences are not included.
The average coverage is 2000x with >99.5% of the coding region covered at ≥350x and 99.2% covered at 200x. The minimum coverage for any additional areas is >30x. Variant and copy number calls are made using a unique bioinformatics pipeline detecting all types of mutations including single nucleotide substitutions, indels and frameshifts caused by deletion or duplication up to 112bp.
Relevant family members of a proband with a (novel or previously identified) variant of unknown significance are offered free of charge targeted analysis as long as accurate phenotypic data are provided by a health care professional to enhance the interpretation. There is no limitation to the number of relatives that can be tested free of charge in such families.
Variant and copy number calls are made using a unique bioinformatics pipeline detecting all types of mutations including single nucleotide substitutions, indels and frameshifts caused by deletion or duplication up to 112bp. Deletion/duplication analysis for SPRED1 is included in this test, as such mutations are a part of the mutation spectrum for this gene. Deletion/duplication analysis for the other 15 genes on this panel is not offered as current empirical and biological evidence is not sufficient to allow the conclusion that an altered copy number of these genes is a mechanism critical for the phenotype associatedwith the Rasopathies. Nevertheless, deletion/duplication analysis through aCGH analysis can be offered as a reflex test through the UAB cytogenetic laboratory (directed by Drs. A. Carroll and F. Mikhail).
SPECIMEN SHIPPING AND HANDLING:
- Please find specimen requirement specifications above.
- All submitted specimens must be sent at room temperature. DO NOT ship on ice.
- Specimens must be packaged to prevent breakage and absorbent material must be included in the package to absorb liquids in the event that breakage occurs. Also, the package must be shipped in double watertight containers (e.g. a specimen pouch + the shipping company’s diagnostic envelope).
- To request a sample collection kit, please click here or email medgenomics@uabmc.edu to complete the specimen request form.
- Please contact the MGL (via email at medgenomics@uabmc.edu, or via phone at 205-934-5562) prior to sample shipment and provide us with the date of shipment and tracking number of the package so that we can better ensure receipt of the samples.
REQUIRED FORMS
Note: Detailed and accurate completion of this document is necessary for reporting purposes. The Medical Genomics Laboratory issues its clinical reports based on the demographic data provided by the referring institution on the lab requisition form. It is the responsibility of the referring institution to provide accurate information. If an amended report is necessary due to inaccurate or illegible documentation, additional reports will be drafted with charge.
Requests for Molecular Genetic testing will not be accepted for the following reasons:
- No label (patients full name and date of collection) on the specimens
- No referring physician’s or genetic counselor’s names and addresses
- No billing information
For more information, test requisition forms, or sample collection and mailing kits, please call: 205-934-5562.
Other related test options: