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Congenital Disorders of Glycosylation, Type Ia (CDG-Ia) via the PMM2 Gene

  • Summary and Pricing
  • Clinical Features and Genetics
  • Citations
  • Methods
  • Ordering/Specimens
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TEST METHODS

Sequencing

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
531 PMM2$650.00 81479 Add to Order
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 18 days.

Clinical Sensitivity
In cases with demonstrated reduced activity of phosphomannomutase and diagnostic serum transferrin glycoforms, PMM2 sequencing is nearly 100% sensitive (Sparks and Krasnewich, GeneReviews 2008).  Among 56 patients with proven enzyme deficiency Matthijs et al (Am J Hum Genet 62:542-550, 1998) found mutations on 99% of the affected alleles, and one mutation (p.Arg141His) accounted for almost 40% of all causative mutations.  The p.Arg141His homozygous genotype has never been observed, presumably because it is embryonic lethal (Matthijs et al. Molec Genet Metab 68:220-226, 1999).  The relatively common European p.Arg141His/p.Phe119Leu and p.Arg141His/p.Asp188Gly genotypes are associated with severe phenotypes, including a high mortality rate for the latter.

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 PMM2$690.00 81479 Add to Order
Pricing Comment

# of Genes Ordered

Total Price

1

$690

2

$730

3

$770

4-10

$840

11-30

$1,290

31-100

$1,670

Over 100

Call for quote

Turnaround Time

The great majority of tests are completed within 28 days.

Clinical Features
Congenital disorders of glycosylation (CDG) are a genetically heterogeneous group of disorders caused by defective synthesis of asparagine (N)-linked glycans. Abnormalities in these glycoconjugates result in disturbed metabolism, cell recognition, cell adhesion, protease resistance, host defense, cell migration, and antigenicity (Marquardt and Denecke. Eur J Pediat 162:359-379, 2003). Consequently, clinical presentations are characterized by multisystem involvement. Individuals with CDG Ia (OMIM 212065) have cerebellar hypoplasia, dysmorphic facies, coagulopathy, strabismus, psychomotor retardation and sometimes unusual fat distribution and inverted nipples (de Lonlay et al. J Med Genet 38:14-19, 2001; Sparks and Krasnewich GeneReviews 2008). Presentation and clinical course can be highly variable, and three stages have been delineated. The infantile multisystem stage is characterized by failure to thrive secondary to vomiting, diarrhea and feeding problems; hypotonia and hyporeflexia; high nasal bridge; prominent jaw and ears; and unusual fat distribution over the buttocks. Some patients who present in this stage exhibit fatal neurological and multi-organ involvement while others present with a non-fatal neurological form lacking visceral organ involvement (de Lonlay et al. J Med Genet 38:14-19, 2001). The late infantile and childhood ataxia-mental retardation stage is characterized by developmental delay, (especially language and motor skills), ataxia, joint contractures, and skeletal deformities. During the adult stable disability stage premature aging is observed along with stable mental capacity, progressive skeletal deformities and hypogonadism (Drouin-Garraud et al. Am J Med Genet 101: 46-49, 2001).
Genetics
All CDGs exhibit autosomal recessive inheritance. Thirteen forms of CDG have been characterized at the molecular level but only three, CDG Ia, CDG Ib, and CDG Ic, have been reported in more than a few individual patients. CDG Ia is the most common form with ~400 cases reported worldwide, followed by CDG 1b and CDG Ic, each with approximately 20 cases. The PMM2 gene (OMIM 601785) codes for phosphomannomutase-2, an enzyme involved in the synthesis of GDP-mannose. Mutations in PMM2 are distributed throughout the coding region and missense mutations are the most common type.
Testing Strategy
Phosphomannomutase-2 is encoded by exons 1 – 8 of the PMM2 gene on chr 16p13.3. Testing is accomplished by amplifying all coding exons and ~20 bp of adjacent noncoding sequence, then determining the nucleotide sequence using standard dideoxy sequencing methods and capillary electrophoresis. 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
Individuals with clinical symptoms consistent with CDG Ia. Individuals with diagnostic serum transferrin isoform results (decreased tetrasialotransferrin and increased asailotransferrin and disialotransferrin).

Gene

Official Gene Symbol OMIM ID
PMM2 601785
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Disease

Name Inheritance OMIM ID
Congenital Disorder Of Glycosylation Type 1A 212065

Related Test

Name
Congenital Disorders of Glycosylation (CDG) Sanger Sequencing Panel 1

CONTACTS

Genetic Counselors
Geneticist
Citations
  • de Lonlay, P., et.al. (2001). "A broad spectrum of clinical presentations in congenital disorders of glycosylation I: a series of 26 cases." J Med Genet 38(1): 14-9. PubMed ID: 11134235
  • Drouin-Garraud, V., et.al. (2001). "Neurological presentation of a congenital disorder of glycosylation CDG-Ia: implications for diagnosis and genetic counseling." Am J Med Genet 101(1): 46-9. PubMed ID: 11343337
  • Matthijs, G., et.al. (1998). "Lack of homozygotes for the most frequent disease allele in carbohydrate-deficient glycoprotein syndrome type 1A." Am J Hum Genet 62(3): 542-50. PubMed ID: 9497260
  • Matthijs, G., et.al. (1999). "Phosphomannomutase deficiency: the molecular basis of the classical Jaeken syndrome (CDGS type Ia)." Mol Genet Metab 68(2): 220-6. PubMed ID: 10527672
  • Susan E Sparks, Donna M Krasnewich (2008). "Congenital Disorder of Glycosylation Type 1a."
  • Susan E Sparks, Donna M Krasnewich (2008). "Congenital Disorders of Glycosylation Overview."
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TEST METHODS

Bi-Directional Sanger Sequencing

Test Procedure

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 20 bases of non-coding DNA flanking the exon are sequenced.

Analytical Validity

As of March 2016, we compared 17.37 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 12 years of our lab operation we have Sanger sequenced roughly 8,800 PCR amplicons. Only one error has been identified, and this was due to sequence analysis error.

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).

Analytical Limitations

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.

In most cases, only the indicated exons and roughly 20 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.

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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