Multiple Congenital Anomalies-Hypotonia-Seizures Syndrome 2 via PIGA Gene Sequencing
- Summary and Pricing
- Clinical Features and Genetics
|Test Code||Test Copy Genes||Price||CPT Code Copy CPT Codes|
CNV detection may be ordered through Test #600.
For ordering sequencing of targeted known variants, please proceed to our Targeted Variants landing page.
The great majority of tests are completed within 18 days.
Clinical sensitivity of PIGA in a large cohort of patients with Multiple congenital anomalies-hypotonia-seizures syndrome 2 relevant phenotypes is unavailable in the literature, because most of the studies are case reports. Analytical sensitivity is expected to be high because the pathogenic variants reported should be detectable by sequencing. PIGA pathogenic variants appear to be a rare cause of disease.
No large deletions and duplications have been reported involving the PIGA gene (Johnston et al. 2012. PubMed ID: 22305531; Kato et al. 2014. PubMed ID: 24706016; Tarailo-Graovac et al. 2015. PubMed ID: 25885527; Human Gene Mutation Database).
Multiple congenital anomalies-hypotonia-seizures syndrome 2 is a rare neurodevelopmental disorder with onset in utero or early infancy. It may be lethal in infancy. The major features include dysmorphic features, neonatal hypotonia, myoclonic seizures, and variable congenital anomalies involving the central nervous, cardiac, and urinary systems (Johnston et al. 2012. PubMed ID: 22305531; Kato et al. 2014. PubMed ID: 24706016; Tarailo-Graovac et al. 2015. PubMed ID: 25885527).
Females with a heterozygous pathogenic variant in PIGA are normally unaffected. Somatic pathogenic variants in PIGA are considered to be causative for paroxysmal nocturnal hemoglobinuria (Johnston et al. 2012. PubMed ID: 22305531; Belet et al. 2014. PubMed ID: 24357517).
Multiple congenital anomalies-hypotonia-seizures syndrome 2 is inherited in an X-linked recessive manner and caused by pathogenic variants in the PIGA gene which encodes the phosphatidylinositol glycan anchor biosynthesis class A protein. This protein is one of the enzymes involved in the first step of biosynthesis of the glycosylphosphatidylinositol (GPI) anchor, a glycolipid structure embedded in the plasma membrane. PIGA pathogenic variants are one genetic cause of early onset epileptic encephalopathy. Pathogenic variants in PIGA gene include missense, nonsense, splicing, and small deletion/duplications (both frameshift and in-frame). No large deletions and duplications have been reported (Johnston et al. 2012. PubMed ID: 22305531; Kato et al. 2014. PubMed ID: 24706016; Tarailo-Graovac et al. 2015. PubMed ID: 25885527; Human Gene Mutation Database).
This test involves bidirectional Sanger sequencing of all coding exons and splice sites of the PIGA gene. The full coding sequence of each exon plus ~10 bp of flanking DNA on either side are sequenced. We will also sequence any single exon (Test #100) or pair of exons (Test #200) in family members of patients with known mutations or to confirm research results.
Indications for Test
PIGA sequencing is recommended for patients who are suspected to have Multiple congenital anomalies-hypotonia-seizures syndrome 2.
|Official Gene Symbol||OMIM ID|
|Multiple Congenital Anomalies-Hypotonia-Seizures Syndrome 2||XL||300868|
|Paroxysmal Nocturnal Hemoglobinuria||XL||300818|
- Genetic Counselor Team - email@example.com
- Li Fan, MD, PhD, FCCMG, FACMG - firstname.lastname@example.org
Bi-Directional Sanger Sequencing
Nomenclature for sequence variants was from the Human Genome Variation Society (http://www.hgvs.org). As required, DNA is extracted from the patient specimen. PCR is used to amplify the indicated exons plus additional flanking non-coding sequence. After cleaning of the PCR products, cycle sequencing is carried out using the ABI Big Dye Terminator v.3.0 kit. Products are resolved by electrophoresis on an ABI 3730xl capillary sequencer. In most cases, sequencing is performed in both forward and reverse directions; in some cases, sequencing is performed twice in either the forward or reverse directions. In nearly all cases, the full coding region of each exon as well as 10 bases of non-coding DNA flanking the exon are sequenced.
Human Genome Variation Society (HGVS) recommendations are used to describe sequence variants (http://www.hgvs.org).All differences from the reference sequences 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. PubMed ID: 25741868). 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.Benign variants are not listed in the reports, but are available upon request.
As of February 2018, we compared 26.8 Mb of Sanger DNA sequence generated at PreventionGenetics to NextGen sequence generated in other labs. We detected only 4 errors in our Sanger sequences, and these were all due to allele dropout during PCR. For Proficiency Testing, both external and internal, in the 14 years of our lab operation we have Sanger sequenced roughly 14,300 PCR amplicons. Only one error has been identified, and this was an error in analysis of sequence data.
Our Sanger sequencing is capable of detecting virtually all nucleotide substitutions within the PCR amplicons. Similarly, we detect essentially all heterozygous or homozygous deletions within the amplicons. Homozygous deletions which overlap one or more PCR primer annealing sites are detectable as PCR failure. Heterozygous deletions which overlap one or more PCR primer annealing sites are usually not detected (see Analytical Limitations). All heterozygous insertions within the amplicons up to about 100 nucleotides in length appear to be detectable. Larger heterozygous insertions may not be detected. All homozygous insertions within the amplicons up to about 300 nucleotides in length appear to be detectable. Larger homozygous insertions may masquerade as homozygous deletions (PCR failure).
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.
In exons where our sequencing did not reveal any variation between the two alleles, we cannot be certain that we were able to PCR amplify both of the patient’s alleles. Occasionally, a patient may carry an allele which does not amplify, due for example to a deletion or a large insertion. In these cases, the report contains no information about the second allele.
Similarly, our sequencing tests have almost no power to detect duplications, triplications, etc. of the gene sequences.
Only the indicated exons and roughly 10 bp of flanking non-coding sequence on each side are analyzed. Test reports contain little or no information about other portions of the gene, including many regulatory regions.
In nearly all 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 for example 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 and cycle sequencing.
Unless otherwise indicated, the sequence data that we report are based on DNA isolated from a specific tissue (usually leukocytes). Test reports contain no information about gene sequences in other tissues.
We cannot be certain that the reference sequence(s) are correct. Exons, for example, may be misidentified. In cases where the genomic and mRNA sequences disagree, we use the genomic sequence as our reference.
We have confidence in our ability to track a specimen once it has been received by PreventionGenetics. However, we take no responsibility for 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 or medical options is recommended.
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.