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Mitochondrial Phosphoenolpyruvate Carboxykinase 2 Deficiency via the PCK2 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
1433 PCK2$870.00 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
Clinical sensitivity cannot be estimated because only a single patient has been reported (DaRe et al. 2013; Robinson 2014). Analytical sensitivity may be high because all reported pathogenic variants are detectable by sequencing.

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 PCK2$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

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Turnaround Time

The great majority of tests are completed within 28 days.

Clinical Sensitivity
To date, no large deletions or duplications have been described in the PCK2 gene.

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Clinical Features
Patients with mitochondrial diseases or other diseases that mimic mitochondrial diseases present with a wide variety of phenotypes and age of onset, making diagnosis of these disorders challenging (DaRe et al. 2013). Clinical features can include encephalomyopathy, combinations of multisystemic symptoms affecting at least three organ symptoms, and a progressive clinical course with episodes of worsening symptoms. Enzymology studies reveal decreased respiratory chain complex (RCC) activity, and metabolic studies may show at least one indicator of impaired RCC activity. Additional diagnostic criteria are described by Bernier et al. (2002).

Disorders of phosphoenolpyruvate carboxykinase (PEPCK) are thought to be a rare cause of childhood onset lactic acidemia. In humans, two PEPCK isozymes are found. One is located in the mitochodria and is encoded by the PCK2 gene. The second is found in the cytosol and is encoded by the PCK1 gene (Robinson 2014). A few early reports have described children with lactic acidemia, hypoglycemia, hypotonia, hepatomegaly and failure to thrive who were also found to have decreased PEPCK enzyme activity (Robinson 2014). More recently, a single PEPCK deficient patient was identified as part of a study of individuals diagnosed with or suspected of having a mitochondrial disorder. A large Next Generation sequencing panel of 447 genes was used in attempt to diagnose these patients (DaRe et al. 2013). This particular patient was found to be a compound heterozygote for two variants in the PCK2 gene, which encodes the mitochondrial PEPCK isozyme. This patient was reported to have abnormal RCC activity and/or muscle pathology, in addition to clinical symptoms consistent with a mitochondrial disorder (DaRe et al. 2013).
Genetics
Mitochondrial PEPCK deficiency is thought to be inherited in an autosomal recessive manner, and is caused by pathogenic variants in the PCK2 gene (DaRe et al. 2013). To date, only missense causative variants have been reported in the PCK2 gene.

The PCK2 gene is a nuclear encoded gene containing 10 coding exons and located on chromosome 14 (14q11-q12). The PEPCK enzyme plays a critical role in gluconeogenesis by catalyzing the conversion of oxaloacetate to phosphoenolpyruvate. The PCK2 gene is expressed primarily in the pancreas, kidney and liver (Modaressi et al. 1998).
Testing Strategy
This test involves bidirectional Sanger sequencing using genomic DNA of all coding exons of the PCK2 gene plus ~20 bp of flanking non-coding DNA on each side. 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 a suspected mitochondrial disorder and lactic acidemia are good candidates for this test, particularly if other causes of disease have been ruled out. Family members of patients who have known PCK2 variants are also good candidates. We will sequence the PCK2 gene to determine carrier status.

Gene

Official Gene Symbol OMIM ID
PCK2 614095
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Related Test

Name
Cytosolic Phosphoenolpyruvate Carboxykinase 1 Deficiency via the PCK1 Gene

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Bernier F.P. et al. 2002. Neurology. 59: 1406-11.  PubMed ID: 12427892
  • DaRe J.T. et al. 2013. Bmc Medical Genetics. 14: 118.  PubMed ID: 24215330
  • Modaressi S. et al. 1998. The Biochemical Journal. 333 ( Pt 2): 359-66.  PubMed ID: 9657976
  • Robinson B.H. 2014. Lactic Acidemia: Disorders of Pyruvate Carboxylase and Pyruvate Dehydrogenase. Online Metabolic & Molecular Bases of Inherited Disease, New York, NY: McGraw-Hill.
Order Kits
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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