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Congenital Myopathy Sequencing Panel

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

NGS Sequencing

Test Code Test Copy GenesCPT Code Copy CPT Codes
1365 ACTA1 81479 Add to Order
BICD2 81479
BIN1 81479
CCDC78 81479
CFL2 81479
CNTN1 81479
COL12A1 81479
COL6A1 81407
COL6A2 81407
COL6A3 81407
DNM2 81479
KBTBD13 81479
KLHL40 81479
KLHL41 81479
LMNA 81406
LMOD3 81479
MEGF10 81479
MICU1 81479
MTM1 81406
MYH7 81407
MYO18B 81479
NEB 81408
RYR1 81408
SELENON 81479
STAC3 81479
TNNT1 81479
TPM2 81479
TPM3 81479
TTN 81479
Full Panel Price* $2190.00
Test Code Test Copy Genes Total Price CPT Codes Copy CPT Codes
1365 Genes x (29) $2190.00 81406(x2), 81407(x4), 81408(x2), 81479(x21) Add to Order
Pricing Comment

If you would like to order a subset of these genes contact us to discuss pricing.

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 28 days.

Clinical Sensitivity

A survey of a defined pediatric population (southeastern Michigan) in the United States found the prevalence of congenital myopathy (CM) to be 1:26,000 (Amburgey et al. 2011). A genetic cause was identified in 35% of cases, and pathogenic variants in the RYR1 gene were the single most common genetic cause of CM in this cohort. Non-specific findings was the most frequently encountered muscle biopsy result followed by minicores, central cores, fiber type disproportion, central nuclei, and nemaline rods (Amburgey et al. 2011).

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Deletion/Duplication Testing via aCGH

Test Code Test Copy GenesIndividual Gene PriceCPT Code Copy CPT Codes
600 ACTA1$690.00 81479 Add to Order
BIN1$690.00 81479
CCDC78$690.00 81479
CFL2$690.00 81479
CNTN1$690.00 81479
COL12A1$690.00 81479
COL6A1$690.00 81479
COL6A2$690.00 81406
COL6A3$690.00 81406
DNM2$690.00 81479
KBTBD13$690.00 81479
KLHL40$690.00 81479
KLHL41$690.00 81479
LMNA$690.00 81479
MEGF10$690.00 81479
MTM1$690.00 81479
MYH7$690.00 81479
NEB$690.00 81479
RYR1$690.00 81479
SELENON$690.00 81479
STAC3$690.00 81479
TNNT1$690.00 81479
TPM2$690.00 81479
TPM3$690.00 81479
TTN$690.00 81479
Full Panel Price* $1290.00
Test Code Test Copy Genes Total Price CPT Codes Copy CPT Codes
600 Genes x (25) $1290.00 81406(x2), 81479(x23) 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 Sensitivity

Clinical sensitivity for del/dup testing is expected to be low. One exception is the Ashkenazi Jewish nebulin exon 55 deletion pathogenic variant.

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Clinical Features

Congenital myopathy (CM) refers to a genetically and clinically heterogeneous group of disorders characterized by muscle weakness and hypotonia at birth or in infancy. Five forms of congenital myopathy are recognized by the International Standards of Care Committee for Congenital Myopathies (North et al. 2014): Nemaline Myopathy, Core Myopathy, Centronuclear Myopathy, Myosin Storage Myopathy, and Congenital Fiber Type Disproportion. The clinical course of congenital myopathy is typically static or slowly progressive. Lower facial weakness leading to an open mouth is often the first sign of congenital myopathy in the newborn. Ptosis and ophthalmoplegia are also common. See North et al. (2014) for diagnostic strategies and a comprehensive review of the congenital myopathies.

Genetics

The genetics of the congenital myopathies are complex because of extensive genetic heterogeneity and overlapping clinical and histopathological findings among the five different subtypes (North et al. 2014).

The following genes have been reported with autosomal recessive inheritance: BIN1, CFL2, CNTN1, KLHL40, KLHL41, LMOD3, MEGF10, MICU1, MYO18B, NEB, STAC3, and TNNT1. The following genes are reported with autosomal dominant inheritance: BICD2, CCDC78, COL12A1, DNM2, KBTBD13, and TPM2. The following genes can exhibit both recessive and dominant inheritance: ACTA1, COL6A1, COL6A2, COL6A3, LMNA, MYH7, RYR1, SELENON/SEPN1, TPM3, and TTN. Of note, some dominant RYR1 central core myopathy variants can exhibit reduced penetrance (for example the c.14818G>A variant, Davis et al 2003). Pathogenic variants in the MTM1 gene cause X-linked centronuclear myopathy.

See individual gene test descriptions for information on molecular biology of gene products.

Testing Strategy

For this Next Generation Sequencing (NGS) test, 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 reported pathogenic, likely pathogenic, and 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.

This panel provides at least 99% coverage of all coding exons of the genes listed, plus ~20 bases of flanking noncoding DNA. We define coverage as >20X NGS reads for coding regions and 0-10 bases of flanking DNA, >10X NGS reads for 11-20 bases of flanking DNA, or Sanger sequencing.

Of note, this testing includes coverage of the SELENON (NM_20451.2) c.*1107T>C variant in the 3' UTR.

Exons 82-105 of the NEB gene are organized in three nearly identical repetitive blocks of 8 exons each making this region difficult to analyze. Since there are six highly homologous alleles, there is some limitation in variant and zygosity calling in this region. If an undocumented or pathogenic variant is detected in this region via NextGen Sequencing, a unique PCR and Sanger sequencing method will be used for confirmation.

This panel does not include coverage of TTN exons 85-95 and exons 153-155 (NM_133378.4) and NEB exon 84 (NM_001271208.1). Sanger coverage of these regions is available for an additional charge; however, very few pathogenic variants have been reported in these regions (Human Gene Mutation Database).

Indications for Test

Individuals with clinical symptoms consistent with congenital myopathy.

Diseases

Name Inheritance OMIM ID
Autosomal Recessive Centronuclear Myopathy AR 255200
Bethlem Myopathy AD,AR 158810
Bethlem Myopathy 2 AD 616471
Central Core Disease AR, AD 117000
Congenital Fiber Type Disproportion AR, AD 255310
Klippel-Feil Syndrome 4, Autosomal Recessive, with Myopathy and Facial Dysmorphism AR 616549
Muscular Dystrophy, Congenital, LMNA-Related AR, AD 613205
Myopathy with Extrapyramidal Signs AR 615673
Myopathy, Centronuclear AD 614807
Myopathy, Centronuclear, 1 AD 160150
Myopathy, Congenital, Compton-North AR 612540
Myopathy, Early-Onset, Areflexia, Respiratory Distress, And Dysphagia AD,AR 614399
Myopathy, Early-Onset, With Fatal Cardiomyopathy AR 611705
Myopathy, Myosin Storage AD 608358
Native American myopathy AR 255995
Nemaline Myopathy 1 AR, AD 609284
Nemaline Myopathy 10 AR 616165
Nemaline Myopathy 2 AR 256030
Nemaline Myopathy 3 AR, AD 161800
Nemaline Myopathy 4 AD 609285
Nemaline Myopathy 5 AR 605355
Nemaline Myopathy 6 AD 609273
Nemaline Myopathy 7 AR 610687
Nemaline Myopathy 8 AR 615348
Nemaline Myopathy 9 AR 615731
Rigid Spine Muscular Dystrophy 1 AR, AD 602771
Severe X-Linked Myotubular Myopathy XL 310400
Spinal Muscular Atrophy, Lower Extremity-Predominant, 2 AD 615290
Ullrich Congenital Muscular Dystrophy AD 254090

Related Tests

Name
RYR1-Related Congenital Myopathies via the RYR1 Gene
Autism Spectrum Disorders and Intellectual Disability (ASD-ID) Comprehensive Sequencing Panel with CNV Detection
Autosomal Dominant Limb Girdle Muscular Dystrophy (LGMD) Sequencing Panel
Autosomal Recessive Limb Girdle Muscular Dystrophy (LGMD) Sequencing Panel
Centronuclear Myopathy Sequencing Panel
Centronuclear Myopathy, X-Linked via the MTM1 Gene
Centronuclear Myopathy-2, Autosomal Recessive via the BIN1 Gene
Centronuclear Myopathy-4, Autosomal Dominant (CNM4) via the CCDC78 Gene
Charcot Marie Tooth - Axonal Neuropathy Sequencing Panel
Charcot Marie Tooth - Comprehensive Sequencing Panel
Charcot Marie Tooth - Demyelinating Neuropathy Sequencing Panel
Complex Hereditary Spastic Paraplegia Sequencing Panel with CNV Detection
Comprehensive Cardiology Sequencing Panel with CNV Detection
Comprehensive Neuromuscular Sequencing Panel
Comprehensive Neuropathy Sequencing Panel
Compton-North Congenital Myopathy via the CNTN1 Gene
Congenital Fiber Type Disproportion Sequencing Panel
Congenital Muscular Dystrophy Sequencing Panel
Congenital Myopathy (Native American Myopathy) via the STAC3 Gene
Core Myopathy Sequencing Panel
Dilated Cardiomyopathy Sequencing Panel with CNV Detection
Distal Arthrogryposis Sequencing Panel
Distal Hereditary Myopathy Sequencing Panel
Dynamin-2 Related Disorders via the DNM2 Gene
Early-Onset Myopathy, Areflexia, Respiratory Distress, and Dysphagia (EMARDD) via the MEGF10 Gene
Fetal Akinesia Deformation Sequence/Lethal Multiple Pterygium Syndrome Sequencing Panel
Fetal Concerns Sequencing Panel with CNV Detection
Hereditary Spastic Paraplegia Comprehensive Sequencing Panel with CNV Detection
Hutchinson-Gilford Progeria Syndrome (HGPS) via the LMNA Gene
Hypertrophic Cardiomyopathy and other MYH7-Related Disorders via the MYH7 Gene
Hypertrophic Cardiomyopathy Sequencing Panel with CNV Detection
Laminopathies via the LMNA Gene
Left Ventricular Noncompaction (LVNC) Sequencing Panel with CNV Detection
Limb Girdle Muscular Dystrophy, Type 2J and Tibial Muscular Dystrophy via the TTN Gene (exons 307 - 312)
Limb-Girdle Muscular Dystrophy (LGMD) Sequencing Panel
Malignant Hyperthermia Susceptibility Sequencing Panel
Metabolic Myopathies, Rhabdomyolysis and Exercise Intolerance Sequencing Panel
Myofibrillar Myopathy Sequencing Panel
Myopathy, Congenital via the TPM3 Gene
Nemaline Myopathy (NEM2) via the Nebulin (NEB) Gene
Nemaline Myopathy 10 via the LMOD3 Gene
Nemaline Myopathy 5 (Amish Nemaline Myopathy) via the TNNT1 Gene
Nemaline Myopathy 7 via the CFL2 Gene
Nemaline Myopathy 9 via the KLHL41 Gene
Nemaline Myopathy Sequencing Panel
Nemaline Myopathy via the KLHL40 Gene
Nemaline Myopathy via the NEB Exon 55 Deletion
Nemaline Myopathy With Cores (NEM6) via the KBTBD13 Gene
Nemaline Myopathy NEB Triplicate Repeat Region, Exons 82-105
Pure Hereditary Spastic Paraplegia Sequencing Panel with CNV Detection
Selenoprotein N, 1 via the SELENON/SEPN1 Gene
Sudden Cardiac Arrest Sequencing Panel with CNV Detection
Tropomyosin 2-Related Disorders via the TPM2 Gene
Type VI Collagenopathy via the COL6A1 Gene
Type VI Collagenopathy via the COL6A2 Gene
Type VI Collagenopathy via the COL6A3 Gene
Type VI-Related Collagenopathy Sequencing Panel
Type VI-Related Collagenopathy via the COL12A1 Gene

CONTACTS

Genetic Counselors
Geneticist
Citations
  • Amburgey K. et al. 2011. Annals of Neurology. 70: 662-5. PubMed ID: 22028225
  • Davis M.R. et al. 2003. Neuromuscular Disorders. 13: 151-7. PubMed ID: 12565913
  • Human Gene Mutation Database (Bio-base).
  • North K.N. et al. 2014. Neuromuscular Disorders. 24: 97-116. PubMed ID: 24456932
Order Kits
TEST METHODS

NextGen Sequencing using PG-Select Capture Probes

Test Procedure

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.

Analytical Validity

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.   

Analytical Limitations

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

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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