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Maple Syrup Urine Disease Type IB via the BCKDHB Gene

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

Sequencing

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
527 BCKDHB$810.00 81406 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
It has been reported that mutations in the BCKDHB gene are responsible of approximately 38% of the MSUD cases, while mutations in the BCKDHA and the DBT genes are responsible for approximately 33% and 19% of MSUD cases, respectively (Nellis and Danner. Am J Hum Genet 68:232-237, 2001).

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 BCKDHB$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
Maple syrup urine disease (MSUD; OMIM 248600) is a heterogeneous organic aciduria disorder caused by the impairment of the branched-chain α-keto acid dehydrogenase complex (BCKD). BCKD is a mitochondrial complex, encoded by four nuclear genes (BCKDHA, BCKDHB, DBT and DLD), which is involved in the metabolism of branched-chain amino acids (leucine, isoleucine, and valine) (Morton et al. Pediatrics 109:999-1008, 2002; Nellis et al. Molec Genet Metab 80:189-195, 2003; Chuang et al. J Biol Chem 279:17792-17800, 2004). Defective BCKD complex activity leads to the accumulation of the branch-chain amino acids to toxic levels (Chuang et al. 2004). MSUD, in untreated neonates, is characterized by mental and physical retardation, maple syrup odor in cerumen and urine, poor feeding, ketonuria, irritability, lethargy, intermittent apnea, opisthotonus, stereotyped movements such as “fencing” and “bicycling”, coma and respiratory failure. Biochemically, MSUD is characterized by elevated plasma concentrations of branched-chain amino acids (leucine, isoleucine, and valine) and allo-isoleucine, as well as a generalized disturbance of plasma amino acid concentration ratios (Schadewaldt et al. Clin Chem 45:1734-1740, 1999; Morton et al. 2002; Nellis et al. 2003; Chuang et al. 2004).
Genetics
MSUD is an autosomal recessive genetically heterogeneous disorder caused by mutations in one of the four BCKD complex encoded genes (BCKDHA, BCKDHB, DBT and DLD). MSUD Type IB is caused by mutations in the BCKDHB gene, which encodes the 2-oxoisovalerate dehydrogenase subunit beta of the of the BCKD complex (Nobukuni et al. J Clin Invest 87:1862-1866, 1991; Mitsubuchi et al. J Biol Chem 266:14686-14691, 1991; Chuang et al. Am J Hum Genet 58: 1373-1377, 1996). A mix of missense, nonsense, splicing, small insertion and deletion mutations as well as gross deletion mutations within the BCKDHB gene have been reported (Nobukuni et al. 1991; Henneke et al. Hum Mutat 22:417-422, 2003; Chuang et al. 2004; Rodriguez et al. Hum Mutat 27:715-727, 2006).
Testing Strategy
This test involves bidirectional sequencing using genomic DNA of the 10 coding exons (exons 1-10) of the BCKDHB gene. The full coding region of each exon plus ~20 bp of flanking non-coding DNA on each 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
Candidates for this test are patients with symptoms consistent with MSUD and family members of patients who have known BCKDHB mutations.

Gene

Official Gene Symbol OMIM ID
BCKDHB 248611
Inheritance Abbreviation
Autosomal Dominant AD
Autosomal Recessive AR
X-Linked XL
Mitochondrial MT

Disease

Name Inheritance OMIM ID
Maple Syrup Urine Disease 248600

Related Test

Name
Maple Syrup Urine Disease Sequencing Panel

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Chuang, J. L., et.al. (1996). "Maple syrup urine disease: the E1beta gene of human branched-chain alpha-ketoacid dehydrogenase complex has 11 rather than 10 exons, and the 3' UTR in one of the two E1beta mRNAs arises from intronic sequences." Am J Hum Genet 58(6): 1373-7. PubMed ID: 8651316
  • Chuang, J. L., et.al. (2004). "Structural and biochemical basis for novel mutations in homozygous Israeli maple syrup urine disease patients: a proposed mechanism for the thiamin-responsive phenotype." J Biol Chem 279(17): 17792-800. PubMed ID: 14742428
  • Henneke, M., et.al. (2003). "Identification of twelve novel mutations in patients with classic and variant forms of maple syrup urine disease." Hum Mutat 22(5): 417 - 422. PubMed ID: 14517957
  • Mitsubuchi, H., et.al. (1991). "Structural organization and chromosomal localization of the gene for the E1 beta subunit of human branched chain alpha-keto acid dehydrogenase." J Biol Chem 266(22): 14686-91. PubMed ID: 1860867
  • Morton, D. H., et.al. (2002). "Diagnosis and treatment of maple syrup disease: a study of 36 patients." Pediatrics 109(6): 999-1008. PubMed ID: 12042535
  • Nellis, M. M., Danner, D. J. (2001). "Gene preference in maple syrup urine disease." Am J Hum Genet 68(1): 232-7. PubMed ID: 11112664
  • Nellis, M. M., et.al. (2003). "Relationship of causative genetic mutations in maple syrup urine disease with their clinical expression." Mol Genet Metab 80(1-2): 189-95. PubMed ID: 14567968
  • Nobukuni, Y., et.al. (1991). "Maple syrup urine disease. Complete defect of the E1 beta subunit of the branched chain alpha-ketoacid dehydrogenase complex due to a deletion of an 11-bp repeat sequence which encodes a mitochondrial targeting leader peptide in a family with the disease." J Clin Invest 87(5): 1862-6. PubMed ID: 2022752
  • Rodriguez-Pombo, P., et.al. (2006). "Mutational spectrum of maple syrup urine disease in Spain." Hum Mutat 27(7): 715-727. PubMed ID: 16786533
  • Schadewaldt, P., et.al. (1999). "Significance of L-alloisoleucine in plasma for diagnosis of maple syrup urine disease." Clin Chem 45(10): 1734-40. PubMed ID: 10508118
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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