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Joubert and Meckel-Gruber Syndromes Sequencing Panel
- Summary and Pricing
- Clinical Features and Genetics
- Citations
- Methods
- Ordering/Specimens
Order Kits
TEST METHODS
- NextGen Sequencing using PG-Select Capture Probes
- Deletion/Duplication Testing via Array Comparative Genomic Hybridization
NGS Sequencing
| Test Code | Test Copy Genes | CPT Code Copy CPT Codes | |
|---|---|---|---|
| 1057 | AHI1 | 81407 | Add |
| ARL13B | 81479 | ||
| B9D1 | 81479 | ||
| B9D2 | 81479 | ||
| C5orf42 | 81479 | ||
| CC2D2A | 81479 | ||
| CEP290 | 81408 | ||
| CEP41 | 81479 | ||
| CSPP1 | 81479 | ||
| INPP5E | 81479 | ||
| KIF14 | 81479 | ||
| KIF7 | 81479 | ||
| MKS1 | 81479 | ||
| NPHP1 | 81406 | ||
| NPHP3 | 81479 | ||
| OFD1 | 81479 | ||
| PDE6D | 81479 | ||
| RPGRIP1L | 81479 | ||
| TCTN1 | 81479 | ||
| TCTN2 | 81479 | ||
| TCTN3 | 81479 | ||
| TMEM138 | 81479 | ||
| TMEM216 | 81479 | ||
| TMEM231 | 81479 | ||
| TMEM237 | 81479 | ||
| TMEM67 | 81407 | ||
| TTC21B | 81479 | ||
| ZNF423 | 81479 | ||
| Full Panel Price* | $640.00 | ||
| Test Code | Test Copy Genes | Total Price | CPT Codes Copy CPT Codes | |
|---|---|---|---|---|
| 1057 | Genes x (28) |
$640.00 | 81406, 81407(x2), 81408, 81479(x24) | Add |
Pricing Comment
We are happy to accommodate requests for single genes or a subset of these genes. The price will remain the list price. If desired, free reflex testing to remaining genes on panel is available. Alternatively, a single gene or subset of genes can also be ordered on our PGxome Custom Panel.
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
Clinical sensitivity for this NextGen test for JSRD and MKS is ~ 50% (Parisi and Glass 2013).
Deletion/Duplication Testing via aCGH
| Test Code | Test Copy Genes | Individual Gene Price | CPT Code Copy CPT Codes | |
|---|---|---|---|---|
| 600 | AHI1 | $990.00 | 81479 | Add |
| ARL13B | $990.00 | 81479 | ||
| B9D1 | $990.00 | 81479 | ||
| C5orf42 | $990.00 | 81479 | ||
| CC2D2A | $990.00 | 81479 | ||
| CEP290 | $990.00 | 81479 | ||
| CEP41 | $990.00 | 81479 | ||
| INPP5E | $990.00 | 81479 | ||
| KIF7 | $990.00 | 81479 | ||
| MKS1 | $990.00 | 81479 | ||
| NPHP1 | $990.00 | 81405 | ||
| NPHP3 | $990.00 | 81479 | ||
| OFD1 | $990.00 | 81479 | ||
| RPGRIP1L | $990.00 | 81479 | ||
| TCTN1 | $990.00 | 81479 | ||
| TCTN2 | $990.00 | 81479 | ||
| TMEM138 | $990.00 | 81479 | ||
| TMEM216 | $990.00 | 81479 | ||
| TMEM237 | $990.00 | 81479 | ||
| TMEM67 | $990.00 | 81479 | ||
| Full Panel Price* | $1490.00 | |||
| Test Code | Test Copy Genes | Total Price | CPT Codes Copy CPT Codes | |
|---|---|---|---|---|
| 600 | Genes x (20) |
$1490.00 | 81405, 81479(x19) | Add |
Pricing Comment
|
# of Genes Ordered |
Total Price |
|
1 |
$990 |
|
2-5 |
$1190 |
|
6-10 |
$1290 |
|
11-100 |
$1490 |
|
Over 100 |
Call for quote |
Turnaround Time
The great majority of tests are completed within 20 days.
Clinical Sensitivity
Gross deletions or duplications not detectable by Sanger sequencing have been reported in AHI1, CC2D2A, CEP290, MKS1, NPHP1, OFD1, and TMEM67 (Human Gene Mutation Database). Clinical sensitivity is difficult to predict due to the paucity of documented cases.
Clinical Features
Joubert Syndrome and related disorders (JSRD) are marked by hypotonia, abnormal ocular movements, neonatal respiratory difficulties, intellectual disability, hypoplasia of the cerebellar vermis, and malformation of the brainstem. The brain malformations lead to the "molar tooth sign" on cranial MRI, which is pathognomonic for JSRD. Other variable JSRD features include cystic kidneys, nephronophthisis, retinal dystrophy, ocular coloboma, occipital encephalocele, polydactyly, ataxia, and hepatic fibrosis. For more information, see (Parisi and Glass 2013; Doherty 2009; Parisi et al. 2007).
Meckel-Gruber Syndrome (MKS) is a lethal autosomal recessive condition, also marked by brain malformation, cystic renal disease and polydactyly (Alexiev et al. 2006). In MKS, the pathognomonic feature is occipital encephalocele, which is generally identified during routine sonography between 12 and 20 weeks of gestation. MKS is a common cause of prenatal echogenic kidneys (Chaumoitre et al. 2006). Nearly all MKS infants are stillborn or die shortly after birth.
Genetics
Both JSRD and MKS are genetically heterogeneous; JSRD is known to be caused by pathogenic variants in at least 20 different genes and MKS is caused by pathogenic variants in at least 12 different genes (Parisi and Glass 2013). Most of the genes reported to cause MKS have also been found to cause JSRD, with the exception of B9D2, KIF14, NPHP3, and TTC21B. In addition, all genes reported to cause MKS and JSRD play some role in the structure, function and maintenance of the primary cilia and/or basal body organelle (Hildebrandt et al. 2009). Thus, MKS and JSRD have been proposed to represent a single clinical entity, with a spectrum of overlapping symptoms and causative genes. JSRD and MKS are inherited in an autosomal recessive manner with the exception of OFD1, which is inherited in an X-linked dominant manner. More information about the molecular biology of the gene products may be found in the individual gene test descriptions.
Testing Strategy
For this NextGen panel, the full coding regions plus ~10 bp of non-coding DNA flanking each exon are sequenced for each of the genes 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
This test is for individuals with symptoms suggestive of MKS or JSRD and their families.
Genes
| Official Gene Symbol | OMIM ID |
|---|---|
| AHI1 | 608894 |
| ARL13B | 608922 |
| B9D1 | 614144 |
| B9D2 | 611951 |
| C5orf42 | 614571 |
| CC2D2A | 612013 |
| CEP290 | 610142 |
| CEP41 | 610523 |
| CSPP1 | 611654 |
| INPP5E | 613037 |
| KIF14 | 611279 |
| KIF7 | 611254 |
| MKS1 | 609883 |
| NPHP1 | 607100 |
| NPHP3 | 608002 |
| OFD1 | 300170 |
| PDE6D | 602676 |
| RPGRIP1L | 610937 |
| TCTN1 | 609863 |
| TCTN2 | 613846 |
| TCTN3 | 613847 |
| TMEM138 | 614459 |
| TMEM216 | 613277 |
| TMEM231 | 614949 |
| TMEM237 | 614423 |
| TMEM67 | 609884 |
| TTC21B | 612014 |
| ZNF423 | 604557 |
| Inheritance | Abbreviation |
|---|---|
| Autosomal Dominant | AD |
| Autosomal Recessive | AR |
| X-Linked | XL |
| Mitochondrial | MT |
Diseases
Related Tests
CONTACTS
Genetic Counselors
- Genetic Counselor Team - clinicaldnatesting@preventiongenetics.com
Geneticist
- Anthony Krentz, PhD - anthony.krentz@preventiongenetics.com
Citations
- Alexiev B.A. et al. 2006. Archives of Pathology & Laboratory Medicine. 130: 1236-8. PubMed ID: 16879033
- Chaumoitre K. et al. 2006. Ultrasound in Obstetrics & Gynecology. 28: 911-7. PubMed ID: 17094077
- Doherty D. 2009. Seminars in Pediatric Neurology. 16: 143-54. PubMed ID: 19778711
- Hildebrandt F. et al. 2009. Journal of the American Society of Nephrology. 20: 23-35. PubMed ID: 19118152
- Human Gene Mutation Database (Bio-base).
- Parisi M, Glass I. 2013. Joubert Syndrome and Related Disorders. 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: 20301500
- Parisi M.A. et al. 2007. European Journal of Human Genetics. 15: 511-21. PubMed ID: 17377524
Order Kits
TEST METHODS
- NextGen Sequencing using PG-Select Capture Probes
- Deletion/Duplication Testing via Array Comparative Genomic Hybridization
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 (http://www.hgvs.org). 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.
Deletion/Duplication Testing via Array Comparative Genomic Hybridization
Test Procedure
Equal amounts of genomic DNA from the patient and a gender matched reference sample are amplified and labeled with Cy3 and Cy5 dyes, respectively. To prevent any sample cross contamination, a unique sample tracking control is added into each patient sample. Each labeled patient product is then purified, quantified, and combined with the same amount of reference product. The combined sample is loaded onto the designed array and hybridized for at least 22-42 hours at 65°C. Arrays are then washed and scanned immediately with 2.5 µM resolution. Only data for the gene(s) of interest for each patient are extracted and analyzed.
Analytical Validity
PreventionGenetics' high density gene-centric custom designed aCGH enables the detection of relatively small deletions and duplications within a single exon of a given gene or deletions and duplications encompassing the entire gene. PreventionGenetics has established and verified this test's accuracy and precision.
Analytical Limitations
Our dense probe coverage may allow detection of deletions/duplications down to 100 bp; however due to limitations and probe spacing this cannot be guaranteed across all exons of all genes. Therefore, some copy number changes smaller than 100-300 bp within a targeted large exon may not be detected by our array.
This array may not detect deletions and duplications present at low levels of mosaicism or those present in genes that have pseudogene copies or repeats elsewhere in the genome.
aCGH will not detect balanced translocations, inversions, or point mutations that may be responsible for the clinical phenotype.
Breakpoints, if occurring outside the targeted gene, may be hard to define.
The sensitivity of this assay may be reduced when DNA is extracted by an outside laboratory.
Order Kits
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.
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.
SPECIMEN TYPES
WHOLE BLOOD
(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.
DNA
(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.
CELL CULTURE
(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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