*'Almost-perfect baby' era is coming*
DNA Deletion And Duplication And The Associated Genetic Disorders
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The Duplication Of Chromosomes
Scientific American is part of Springer Nature, which owns or has commercial relations with thousands of scientific publications (many of them can be found at www.Springernature.Com/us). Scientific American maintains a strict policy of editorial independence in reporting developments in science to our readers.NEXT GEN SEQUENCING AND DELETION/DUPLICATION ANALYSIS OF NF1 ONLY (NF1-NG)
NEXT GEN SEQUENCING AND DELETION/DUPLICATION ANALYSIS OF NF1 ONLY (NF1-NG) Acceptable specimen types:Candidates for this test:
TEST DESCRIPTION
The NF1-only by NGS involves sequencing as well as deletion/duplication analysis of the entire coding NF1 region plus the alternatively spliced exons 9br, 23a and 48a (60 exons total). The test uses an extensively 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 >99.8% of the NF1 coding region ≥350x and 100% ≥200x, allowing detection of very low level mosaicism, down to 3-5% MAF respectively (regions covered by ≥350x respectively ≥200x). 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/ duplication up to 112bp.
Based on >15 years of experience with comprehensive RNA-based NF1 testing, we designed the customized and optimized NGS NF1-component of the assay to comprise all regions encountered through analysis of >15,000 unrelated individuals including >8,100 NF1-mutation-positive individuals carrying 1 out of >3,100 different unique NF1 mutations identified in the UAB MGL cohort. Included in the NGS assay are the regions covering >65 different deep intronic splice mutations (which reside beyond the +/-50 intronic base pairs that flank all exons). Validation of the full panel included, besides substitutions (missense, nonsense, splice variants), the most challenging mutations such as insertions/deletions/duplications of 1-112bp (~25% of the UAB NF1 cohort) and one-to-multiple exon deletions/duplications (~2.8% of the UAB NF1 cohort). The analytical sensitivity of our NGS testing approach was 100% for substitutions as well as insertion/deletions up to 112bp. This panel has not yet been validated to identify deletions/duplications >112bp and <1 exon, but such mutations have not yet been found in the UAB cohort, and therefore are likely very rare. The panel has been validated for the detection of germline (heterozygous) single-exon deletions/duplications as well as multi-exon deletions/duplications, however mosaic single-exon deletion/duplications validation is still pending. Single exon deletions/duplications are present in ~0.45% of NF1-positive patients from the UAB cohort with 9% of these individuals being mosaic (~0.045% of all in the UAB NF1-positive cohort). Detection of Alu/LINE insertions, identified in 0.25% of patients from the UAB NF1-positive cohort, has not yet been validated using the current NGS approach.
With the largest dataset of NF1 genotypes matched with phenotypes, any genotype-phenotype correlations identified will be reported in real time.
Confirmatory testing of reportable variants is performed by Sanger sequencing or other orthogonal methods.
For novel NF1 variants of unknown significance, we offer free of charge targeted RNA-based testing to assess the effect of the variant on splicing and enhance the correct classification/ interpretation.
Relevant family members of a proband with any (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.
Mosaicism is often present in sporadic patients with an NF1 microdeletion and has important repercussions for counseling. Free of charge evaluation by FISH analysis on 200 interphase chromosomes is offered in such cases.
SPECIMEN SHIPPING AND HANDLING:
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:
For more information, test requisition forms, or sample collection and mailing kits, please call: 205-934-5562.
Other related test options:
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