Neonatal Crisis Sequencing Panel with CNV Detection
- Summary and Pricing
- Clinical Features and Genetics
Comprehensive Sequencing Panel with CNV Detection
|Test Type||Test Code||Total Price|
|Duo (Proband + Family Member)||7383, 7382||$3490.00|
|Trio (Proband + 2 Family Members)||7383, 7382(x2)||$4490.00|
|Genes x(1156)||81161(x1), 81222(x1), 81223(x1), 81252(x1), 81302(x1), 81304(x1), 81321(x1), 81323(x1), 81403(x6), 81404(x35), 81405(x52), 81406(x59), 81407(x17), 81408(x18), 81479(x2117)|
For a full list of genes click here.
We prioritize all neonatal illness tests in order to decrease our turnaround time as much as possible. Barring technical issues, we anticipate a maximum of 28 days for sequencing results, although we strive to report results by 15 days whenever possible. In order to allow faster reporting of results, we may issue preliminary reports of the sequencing results in certain cases (please see the Reporting section below for further details).
Please note that in order to offer the fastest possible turnaround time for this test, results of copy number variant (CNV) analysis may be reported at a later date than the results of the sequencing portion of the test.
Due to the genetic heterogeneity of the disorders tested in this panel, the clinical sensitivity of this specific grouping of genes is difficult to estimate. We are currently unaware of any reports in the literature in which these genes have been sequenced together.
Reports will consist of two different sections:
Variants in genes known to be associated with phenotype
Variants in genes possibly associated with phenotype
All differences from the reference sequences (sequence variants) are assigned to one of five interpretation categories (Pathogenic, Likely Pathogenic, Variant of Uncertain Significance, Likely Benign and Benign) per ACMG Guidelines (Richards et al. 2015). Pathogenic, Likely Pathogenic and Variants of Uncertain Significance considered to contribute to the proband's phenotype will be reported in the first and second sections (1 & 2).
Human Genome Variation Society (HGVS) recommendations are used to describe sequence variants (http://www.hgvs.org).
CNVs that are found to encompass all or part of a gene(s) that is known to be associated with, or is possibly associated with, the patient’s phenotype will also be reported. In addition, we will report CNVs of clinical relevance that are >0.5M Mb for deletions or >1 Mb for duplications. Please note that our analysis method may detect CNVs below these sizes. Such CNVs will be reported only if they seem to be of clinical relevance to the patient. Please note that in order to offer the fastest possible turnaround time for this test, results of copy number variant (CNV) analysis may be reported at a later date than the results of the sequencing portion of the test.
When possible, a preliminary report may be issued. This will only occur for cases with a clearly positive outcome, and for which the sequencing data meet our internal pre-determined quality metrics. In such cases, the preliminary report will be issued prior to Sanger confirmation of the reported variants. Once Sanger results have been completed, a final report will be issued with any updates and information regarding other identified variants that might be relevant to the patient’s condition.
Limitations and Other Test Notes:
Interpretation of the test results is limited by the information that is currently available. Enhanced interpretation should be possible in the future as more data and knowledge about human genetics and this specific disorder accumulate. A negative finding does not rule out a genetic diagnosis.
When sequencing does not reveal any heterozygous differences from the reference sequence, we cannot be certain that we were able to detect both patient alleles. Occasionally, a patient may carry an allele which does not capture or amplify, due to a large deletion or insertion. In these cases, the report will contain no information about the second allele.
For technical reasons, the Newborn Crises test is not 100% sensitive. Some exons cannot be efficiently captured, and some genes cannot be accurately sequenced because of the presence of multiple copies in the genome. Therefore, a small fraction of sequence variants relevant to the patient's health will not be detected.
CNVs of four exons or more in size are detected with sensitivity approaching 100% through analysis of NextGen sequence data. However, sensitivity for detection of CNVs smaller than four exons is lower (we estimate ~75%).
In order to offer the fastest possible turnaround time for this test, results of copy number variant (CNV) analysis may be reported at a later date than the results of the sequencing portion of the test.
We sequence coding exons for most given transcripts, plus ~10 bp of flanking non-coding DNA for each exon. Unless specifically indicated, test reports contain no information about other portions of the gene, such as regulatory domains, deep intronic regions, uncharacterized alternative exons, chromosomal rearrangements, repeat expansions, epigenetic effects, and mitochondrial genome variants.
In most cases, we are unable to determine the phase of sequence variants. In particular, when we find two likely causative variants for recessive disorders, we cannot be certain that the variants are on different alleles, unless parental specimens are also tested.
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 amplification.
Unless otherwise indicated, DNA sequence data is obtained from a specific cell-type (usually leukocytes if taken 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.
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.
Genetic counseling to help to explain test results to the patients and to discuss reproductive options is recommended.
Many known monogenic diseases may present early in life as severe neonatal or early childhood (<2 years old) illness. In the United States, such diseases account for ~20% of infant deaths and ~18% of pediatric hospitalizations (Kingsmore. 2012. PubMed ID: 22872815; Saunders et al. 2012. PubMed ID: 23035047). Recent studies have shown that genetic testing in such patients can be a very beneficial first-line test, as the results may provide a comprehensive molecular diagnosis that allows for early intervention for the patient, proper genetic counseling of the family, and may reduce the time spent in the diagnostic odyssey (Kingsmore. 2012. PubMed ID: 22872815; Saunders et al. 2012. PubMed ID: 23035047; Ceyhan-Birsoy et al. 2017. PubMed ID: 28079900).
In recent studies, thorough curation of clinically relevant genes has shown that only ~500-1000 genes are typically associated with the majority of the severe infant or early childhood onset genetic disorders (Kingsmore. 2012. PubMed ID: 22872815; Ceyhan-Birsoy et al. 2017. PubMed ID: 28079900). Clinically, such patients may present with a wide-variety of non-specific symptoms, including but not limited to: respiratory distress, hypotonia, gastrointestinal distress, difficulty feeding, failure to thrive, lethargy, seizures, encephalopathy, cardiac defects, organomegaly, unusual facial features, abnormal odor, and metabolic disturbances (Saudubray and Cazorla. 2016).
This panel includes 1156 genes. The majority of the genes in this test have been included in large gene panels designed for testing of disorders with severe neonatal or early childhood onset (Kingsmore. 2012. PubMed ID: 22872815; Ceyhan-Birsoy et al. 2017. PubMed ID: 28079900). In the more recent study by Ceyhan-Birsoy et al. (2017), genes were divided into several categories. For this test, we have included the majority of the genes classified as Category A (genes with definitive or strong evidence to cause a highly penetrant childhood-onset disorder) in their study. In addition, we have included a number of nuclear genes that have been recently associated with mitochondrial disorders (Craven et al. 2017. PubMed ID: 28415858).
The full list of genes sequenced in this test is available under the “Summary and Pricing” tab.
For the Newborn Crisis panel, Next Generation Sequencing (NGS) technologies are used to cover the coding regions of targeted genes plus ~10 bases of non-coding DNA flanking each exon. Genomic DNA is extracted from patient specimens, as required. Patient DNA corresponding to the targeted genes is captured using Agilent Clinical Research Exome hybridization probes. Captured DNA is sequenced on the NovaSeq 6000 using 2x150 bp paired-end reads (Illumina, San Diego, CA, USA). The following quality control metrics are generally achieved: >97% of target bases are covered at >20x, and mean coverage of target bases >120x. Data analysis and interpretation is performed by the internally developed software Titanium-Exome. Variant calls are made by the GATK Haplotype caller and annotated using in house software and SnpEff. Variants are filtered and annotated using VarSeq (www.goldenhelix.com). Common benign and low quality variants are filtered from analysis. All reported pathogenic, likely pathogenic, and 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.
Copy number variants (CNVs) are also detected from NGS data. We utilize a CNV calling algorithm that compares mean read depth and distribution for each target in the test sample against multiple matched controls. Neighboring target read depth and distribution and zygosity of any variants within each target region are used to reinforce CNV calls. All CNVs are confirmed using another technology such as aCGH, MLPA, or PCR before they are reported.
The report will not include all the observed variants in a patient due to the large number of genes included. However, the list of variants is available, along with our interpretations, upon request. Since this test is performed using exome capture probes, a reflex to exome sequencing may be ordered. Please see the PGxome page (Test #5000) for limitations and reporting criteria for this test.
Indications for Test
This test is primarily implicated for patients with neonatal, infantile or early childhood onset severe disorders with a suspected genetic cause.
- Genetic Counselor Team - email@example.com
- Ceyhan-Birsoy et al. 2017. PubMed ID: 28079900
- Craven et al. 2017. PubMed ID: 28415858
- Kingsmore. 2012. PubMed ID: 22872815
- Saudubray J-M and Cazorla AG. 2016. Clinical Approach to Inborn Errors of Metabolism in Pediatrics. In: Saudubray JM, Baumgartner MR, and Walter J, editors. Inborn Metabolic Diseases: Diagnosis and Treatment. Berlin: Springer, p 8-14.
- Saunders et al. 2012. PubMed ID: 23035047
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.