Congenital Stationary Night Blindness and Retinal Degeneration via the SLC24A1 Gene
- Summary and Pricing
- Clinical Features and Genetics
|Test Code||Test||Individual Gene Price||CPT Code Copy CPT Codes|
Our most cost-effective testing approach is NextGen sequencing with Sanger sequencing supplemented as needed to ensure sufficient coverage and to confirm NextGen calls that are pathogenic, likely pathogenic or of uncertain significance. If, however, full gene Sanger sequencing only is desired (for purposes of insurance billing or STAT turnaround time for example), please see link below for Test Code, pricing, and turnaround time information.
For ordering targeted known variants, please proceed to our Targeted Variants landing page.
The great majority of tests are completed within 28 days.
Predicting clinical sensitivity for the SLC24A1 gene is challenging due to genetic heterogeneity of optic atrophy. However, analytical sensitivity should be high because nearly all reported mutations are detectable by this method (Human Gene Mutation Database).
Deletion/Duplication Testing via aCGH
|Test Code||Test||Individual Gene Price||CPT Code Copy CPT Codes|
# of Genes Ordered
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The great majority of tests are completed within 28 days.
Congenital Stationary Night Blindness (CSNB) is a retinal condition with negative bright-flash dark-adapted electroretinogram response, and optic atrophy or dysplastic changes, or both, in the optic nerve head. Typical findings of CSNB include impaired night vision, no pigmentary retinopathy, full visual fields consistent with myopia and strabismus, which appear from early childhood and are nonprogressive (Heckenlively et al. 1983; Nakamura et al. 2001).Early stages of retinitis pigmentosa (RP) and CSNB are difficult to distinguish. RP and Leber Congenital Amaurosis (LCA) are inherited degenerative diseases of the retina. RP is characterized by night blindness, with age of onset varying from childhood to middle age, progression to constriction of the peripheral visual field and, eventually, to loss of central vision. LCA is characterized by bilateral congenital blindness. Several clinical features of LCA overlap with those of RP. These include attenuated retinal vessels, abnormal ERG findings and a variable amount of retinal pigmentation (Perrault et al. 1996; Daiger et al. 2007; Gu et al. 1999). Both LCA and RP are clinically and genetically heterogeneous.
Maintenance of calcium and sodium concentrations within the rod and cone outer segments is essential for phototransduction, and is regulated by the two similar exchanger or solute carrier proteins in the rod and cones encoded by SLC24A1 and SLC24A2, respectively. Mutations in SLC24A1 are involved with autosomal recessive (AR) CSNB or retinal degeneration (Sharon et al. 2002; Riazuddin et al. 2010). A mutation analysis in a large multigeneration family found five CSNB affected individuals who were homozygous for a SLC24A1 small deletion (c.1613_1614del, p.Phe538Cysfs*23), which segregated with the disease and was not found in 384 control chromosomes (Riazuddin et al. 2010). Mouse studies have shown that SLC24A1 is predominantly expressed in the inner segment, outer and inner nuclear layers of the retina. It is also expressed at low levels in the cornea, lens, and optic nerve (Riazuddin et al. 2010). About twenty causative mutations have been reported in this rod exchanger gene in patients with retinal degeneration (Sharon et al. 2002; Human Gene Mutation Database).
For this NextGen test, the full coding regions plus ~20 bp of non-coding DNA flanking each exon are sequenced for the gene 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
Patients with symptoms suggestive of inherited optic neuropathy in CSNB or retinal degeneration are candidates.
|Official Gene Symbol||OMIM ID|
|Comprehensive Inherited Retinal Dystrophies (includes RPGR ORF15) Sequencing Panel|
|Congenital Stationary Night Blindness Sequencing Panel|
|Optic Atrophy Sequencing Panel|
- Genetic Counselor Team - firstname.lastname@example.org
- Diane Allingham-Hawkins, PhD, FCCMG, FACMG - email@example.com
- Daiger SP, Bowne SJ, Sullivan LS. 2007. Perspective on genes and mutations causing retinitis pigmentosa. Archives of ophthalmology 125: 151-158. PubMed ID: 17296890
- Gu S. et al. 1999. Journal of Medical Genetics. 36: 705-7. PubMed ID: 10507729
- Heckenlively JR, Martin DA, Rosenbaum AL. 1983. Loss of electroretinographic oscillatory potentials, optic atrophy, and dysplasia in congenital stationary night blindness. Am. J. Ophthalmol. 96: 526–534. PubMed ID: 6605090
- Human Gene Mutation Database (Bio-base).
- Nakamura M, Ito S, Terasaki H, Miyake Y. 2001. Novel CACNA1F mutations in Japanese patients with incomplete congenital stationary night blindness. Investigative ophthalmology & visual science 42: 1610–1616. PubMed ID: 11381068
- Perrault I, Rozet JM, Calvas P, Gerber S, Camuzat A, Dollfus H, Châtelin S, Souied E, Ghazi I, Leowski C, Bonnemaison M, Paslier D Le, et al. 1996. Retinal-specific guanylate cyclase gene mutations in Leber’s congenital amaurosis. Nat. Genet. 14: 461–464. PubMed ID: 8944027
- Riazuddin SA, Shahzadi A, Zeitz C, Ahmed ZM, Ayyagari R, Chavali VRM, Ponferrada VG, Audo I, Michiels C, Lancelot M-E, Nasir IA, Zafar AU, et al. 2010. A Mutation in SLC24A1 Implicated in Autosomal-Recessive Congenital Stationary Night Blindness. The American Journal of Human Genetics 87: 523–531. PubMed ID: 20850105
- Sharon D, Yamamoto H, McGee TL, Rabe V, Szerencsei RT, Winkfein RJ, Prinsen CF, Barnes CS, Andreasson S, Fishman GA, others. 2002. Mutated alleles of the rod and cone Na-Ca+ K-exchanger genes in patients with retinal diseases. Investigative ophthalmology & visual science 43: 1971–1979. PubMed ID: 12037007
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.
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.
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
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.
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.
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.
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.