Hereditary Motor and Sensory Neuropathy 1A

Charcot-Marie-Tooth disease is a chronic motor and/or sensory neuropathy typically characterized by distal muscle weakness, sensory loss, and pes cavus deformity. Charcot-Marie-Tooth Type 1A (CMT1A) accounts for ~75% of Charcot-Marie-Tooth type 1 (CMT1), and CMT1 accounts for nearly half of all Charcot-Marie-Tooth disease.

CMT1A is caused by an 1.5 Mb duplication in 17p11.2, which results in the inheritance of three copies of the PMP22 gene. Inheritance is autosomal dominant; 20 – 33% of cases are due to de novo duplications. Rare cases of homozygous duplications (resulting in a total of four copies of PMP22) have been described and, in general, manifest a more severe phenotype than the typical three-copy cases.

1. Confirmation of diagnosis:
   1. In individuals with clinical features suggestive of CMT1.
2. Prenatal testing (technically feasible but not routinely performed – contact MGL to discuss):
   1. Pregnancies to couples in which one person has confirmed CMT1A .
3. Presymptomatic testing
   1. Adults at risk of inheriting CMT1A from a parent may be referred for presymptomatic testing for CMT1A.
   2. Requests to test asymptomatic children who are at risk of developing CMT1A are only accepted following genetic counselling by a recognized genetic service.

Multiplex ligation-dependent probe amplification (MLPA) analysis is carried out with the P033-B2 probe mix (MRC-Holland) to determine the gene dosage (i.e. number of copies) of the PMP22 gene. Note: This assay will detect both CMT1A and HNPP.

Blood: 4 mL EDTA is optimal (Minimum: 1 mL EDTA)
DNA: NOT ACCEPTED

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays). 

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  Payment is not required when requests are made for individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) AND eligible for testing according to the test utilization guidelines / policy. If the individual undergoing testing is not insured by these providers or does not meet utilization guidelines or policy, please complete a billing form; testing will only commence after receipt of billing informationTest prices can be found here.

Molecular genetic testing is limited by the current understanding of the genome and the genetics of a particular disease, as well as by the method of detection used. This method will not detect all mutations (e.g., point mutations in the coding region, promoter mutations, and regulatory element mutations). In rare cases, a point mutation could be detected.

For carrier/predictive testing due to family history, it is generally important to first document the gene mutation in an affected or carrier family member. This information should be provided to the laboratory for assessment of whether the assay is appropriate for detection of the familial mutation, and to aid in the interpretation of data.

In some cases, DNA alterations of undetermined or unclear clinical significance may be identified.

Rare single nucleotide variants or polymorphisms could lead to false-negative results. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

A previous bone marrow transplant from an allogenic donor will result in molecular data that reflects the donor genotype rather than the recipient (patient) genotype. Consult the on-service Molecular Geneticist for approach to testing in such individuals.

Transfusions performed with packed red blood cells will generally not affect the outcome of molecular genetic testing. However, if there is no clinical urgency, the cautious approach is to wait one week post packed red cell transfusion before collecting a sample for genetic testing. Consult the on-service Molecular Geneticist as needed.

Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in our interpretation of results may occur if information given is inaccurate or incomplete.

Familial Recurrent Polyneuropathy; Tomaculous Neuropathy

Hereditary Neuropathy with Liability to Pressure Palsies (HNPP) is characterized by repeated focal pressure neuropathies such as carpal tunnel syndrome and peroneal palsy with foot drop, typically with a family history consistent with autosomal dominant inheritance. Prolonged distal nerve conduction latencies is found on electrophysiologic studies of all individuals, symptomatic or not.

In the majority of cases (80%), HNPP is caused a 1.5 Mb contiguous gene deletion at 17p11.2, which includes the PMP22 gene. In the remaining 20%, the condition is caused by a point mutation in the PMP22 gene. Inheritance is autosomal dominant, although 20% of cases arise due to de novo mutations.

1. Confirmation of diagnosis:
   1. In individuals wtih clinical features suggestive of HNPP.
2. Prenatal diagnosis (technically feasible but not routinely performed – contact MGL to discuss):
   1. Pregnancies to couples in which one person has HNPP
3. Presymptomatic testing:
   1. Adults at risk of inheriting HNPP from a parent and who are not yet symptomatic may be referred for predictive testing for HNPP.
   2. Requests to test asymptomatic children who are at risk of developing HNPP are only accepted following genetic counselling by a recognized genetic service.

Multiplex ligation-dependent probe amplification (MLPA) analysis is carried out with the P033-B2 probe mix (MRC-Holland) to determine the gene dosage (i.e. number of copies) of the PMP22 gene. Note: This assay will detect both CMT1A and HNPP.

Blood: 4 mL EDTA is optimal (Minimum: 1 mL EDTA)
DNA: NOT ACCEPTED

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays). 

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  Payment is not required when requests are made for individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) AND eligible for testing according to the test utilization guidelines / policy. If the individual undergoing testing is not insured by these providers or does not meet utilization guidelines or policy, please complete a billing form; testing will only commence after receipt of billing informationTest prices can be found here.

Molecular genetic testing is limited by the current understanding of the genome and the genetics of a particular disease, as well as by the method of detection used. This method will not detect all mutations (e.g., point mutations in the coding region, promoter mutations, and regulatory element mutations). In rare cases, a point mutation could be detected.

For carrier/predictive testing due to family history, it is generally important to first document the gene mutation in an affected or carrier family member. This information should be provided to the laboratory for assessment of whether the assay is appropriate for detection of the familial mutation, and to aid in the interpretation of data.

In some cases, DNA alterations of undetermined or unclear clinical significance may be identified.

Rare single nucleotide variants or polymorphisms could lead to false-negative results. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

A previous bone marrow transplant from an allogenic donor will result in molecular data that reflects the donor genotype rather than the recipient (patient) genotype. Consult the on-service Molecular Geneticist for approach to testing in such individuals.

Transfusions performed with packed red blood cells will generally not affect the outcome of molecular genetic testing. However, if there is no clinical urgency, the cautious approach is to wait one week post packed red cell transfusion before collecting a sample for genetic testing. Consult the on-service Molecular Geneticist as needed.

Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in our interpretation of results may occur if information given is inaccurate or incomplete.

Hemoglobin H Disease; Hydrops Fetalis; Alpha Thalassemia Minor; Alpha Thalassemia Trait; Thalassemia Intermedia; Cooley’s Anemia; Mediterranean Anemia; Beta Thalassemia Major; Beta Thalassemia Minor; Beta Thalassemia Trait; Sickle Cell Disease; Sickle Cell Anemia; Hemoglobin C Trait; Hemoglobin E Trait

Thalassemias and hemoglobinopathies are conditions affecting the quantity and functionality, respectively, of hemoglobin within red blood cells.

The thalassemias are the result of mutations that decrease or eliminate the production of individual globin chains of the hemoglobin tetramer.

The sickle cell disorders are hemoglobinopathies caused by specific point mutations in the β globin gene (hemoglobins S, C, and E) that result in structural abnormalities of the protein rather than decreased production.  The clinical features of the sickle disorders can be quite variable, depending in part on the particular number and combination of α globin mutations.

In addition, since both the α- and β-globin chains comprise the primary adult hemoglobin, the co-inheritance of β globin gene mutations (for either thalassemia or hemoglobinopathies) and α globin mutations (for thalassemia) further increases the clinical variability encountered in this group of disorders.

Alpha thalassemia

Alpha thalassemia typically results from deletion of one or more of the four α globin genes.  Rare point mutations may also contribute to the condition.

Beta thalassemia

Beta thalassemia results most commonly from point mutations that lead to a reduction or complete loss of protein synthesis from one or both β globin genes.

Sickling disorders

The sickling disorders are the result of single point mutations in the β globin gene that result in the production of abnormal β globin chains.  HbS, the hemoglobin that causes sickle cell disease when present in the homozygous state, is caused by a p.Glu6Val β globin substitution (c.20A>T).  HbC is caused by a p.Glu6Lys (c.19G>A) β globin substitution .  HbE is caused by a p.Glu26Lys (c.79G>A) β globin substitution.  Notably, the HbE mutation results in the activation of a cryptic donor splice site, resulting in a thalassemia phenotype when co-inherited with another beta thalassemia mutation.

Other hemoglobinopathies result from various combinations of alpha and/or beta globin mutations as well as the other globin chain genes.

A hematology profile, including CBC and hemoglobin electrophoresis/HPLC, must be performed prior to ordering molecular genetic testing for the hemoglobin disorders unless an individual has a clinical diagnosis of one of the hemoglobin disorders.  If hematology investigations require follow up with molecular genetic testing, then these tests may be ordered.

1. Confirmation of diagnosis: 
   1. Testing ordered by a hematologist as relevant to the clinical presentation of the patient; to confirm a suspected or known clinical diagnosis or clarify unusual hemoglobinopathy cases.
2. Carrier testing:
   1. When ordered by a hematologist: as relevant to the clinical presentation/management of disease of the patient.
   2. Pediatric patients: to aid in the discrimination of carrier status from iron deficiency anemia.
   3. Adults of reproductive age: as per the SOGC-CCMG clinical practice guideline (2008).
   4. Specific for alpha thalassemia:
      1. In adults of reproductive age when:
         1. Both members of the couple have beta thalassemia trait and they may also be at risk of conceiving a child with Hemoglobin Barts hydrops fetalis syndrome.
         2. One member of the couple has beta thalassemia trait and the other has hematology suggestive of alpha thalassemia trait (i.e. their pregnancy may also be at risk of Hb Barts/hydrops fetalis)
      2. NB: Carrier screening to determine the reproductive risk for HbH disease is NOT an indication for molecular genetic testing that is eligible for coverage by BC MSP unless one member of the couple has hematology consistent with alpha thalassemia trait and the other has HPLC findings consistent with the HBA2 Constant Spring or Quong Sze mutations.
3. Prenatal testing (prenatal diagnosis requests are not normally accepted from physicians other than Medical Geneticists):
   1. Pregnancies known to be at risk based on parental carrier screening or ultrasound findings.

Carrier screening to determine the reproductive risk for HbH disease is NOT an indication for molecular genetic testing for alpha thalassemia except where one member of the couple has hematology consistent with alpha thalassemia trait and the other has HPLC findings consistent with a pathogenic HBA1 or HBA2 mutation (for example, hemoglobin Constant Spring). Genetic counselling is required prior to testing for couples in this scenario.

Alpha thalassemia: Gap junction PCR analysis is carried out to detect the –SEA, -α20.5, –MED, –FIL, –THAI, -α3.7, and -α4.2 deletions. Bidirectional Sanger sequencing across the region of the alpha-2 gene (HBA2) that contains the Constant Spring (c.427T>C, p.*143GlnextX32) and Quong Sze (c.377T>C, p.Leu126Pro) mutations is not routinely performed, but is available in certain clinical scenarios; consult on-service Molecular Geneticist.

Beta thalassemia & Hemoglobins S, C, E: Bidirectional Sanger sequencing across all exons of the HBB gene and intron sequences flanking each exon (exon 1: c.-105 to c.92+10; exon 2: c.93-25 to c.315+25; exon 3: c.316-200 to c*110). 

HBA: NM_000517.4  The ‘A’ within the initiation codon, ATG, is designated as nucleotide number 1.

HBB: NM_000518.4  The ‘A’ within the initiation codon, ATG, is designated as nucleotide number 1.

Pregnancy-related/Prenatal

If pregnancy management will be altered, 3 weeks; otherwise, routine TAT.

Blood: 4 mL EDTA is optimal (Minimum: 1 mL EDTA)
DNA: 100 μL at 200 ng/μL is optimal (Minimum: 30 μL at 200 ng/μL)

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays).  

Prenatal Specimens
Prenatal testing REQUIRES LABORATORY CONSULTATION PRIOR TO THE PROCEDURE and can only be ordered by a Medical Geneticist. Contact the laboratory at 604-875-2852 and choose the appropriate option for the Molecular Geneticist on service.
Chorionic Villi: 20 mg.
Direct Amniotic fluid: 25 mL collected in two separate tubes of equal volume.
Cultured Amniocytes: Two (2) 100% confluent T-25 flasks.
DNA extracted from prenatal specimens: 100 μL at 200 ng/μL is optimal (Minimum: 30 μL at 200 ng/μL)

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth. Ship samples by overnight courier with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays) as follows:

• Villi – on wet ice or in media at room temperature
• Amniocytes, Amniotic fluid, DNA – at room temperature

A hematology profile, including CBC and hemoglobin electrophoresis/HPLC MUST accompany the sample and requisition or be faxed separately to MGL when ordering testing for any of the hemoglobin disorders.

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  Payment is not required when requests are made for individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) AND eligible for testing according to the test utilization guidelines / policy. If the individual undergoing testing is not insured by these providers or does not meet utilization guidelines or policy, please complete a billing form; testing will only commence after receipt of billing informationTest prices can be found here.

Molecular genetic testing is limited by the current understanding of the genome and the genetics of a particular disease, as well as by the method of detection used.

Rare single nucleotide variants or polymorphisms could lead to false-negative or false-positive results. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

A previous bone marrow transplant from an allogenic donor will result in molecular data that reflects the donor genotype rather than the recipient (patient) genotype. Consult the on-service Molecular Geneticist for approach to testing in such individuals.

Transfusions performed with packed red blood cells will generally not affect the outcome of molecular genetic testing. However, if there is no clinical urgency, the cautious approach is to wait one week post packed red cell transfusion before collecting a sample for genetic testing. Consult the on-service Molecular Geneticist as needed.

Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in our interpretation of results may occur if information given is inaccurate or incomplete.

Pediatric monitoring post bone marrow transplant; same-sex donor and recipient. 

Donor, pre-transplant recipient, and post-transplant recipient samples are required; consult with on-service Molecular Geneticist if these requirements cannot be met.

Pediatric post-BMT monitoring in opposite-sex donor-recipient pairs is performed in the BCCH Cytogenetics laboratory. Generally, monitoring of post-BMT adults is performed in the BCCA molecular genetics laboratory. For contact information for these labs, please see our Can’t Find It? page.

Using the AmpFlSTR® Identifiler™ kit, multiplex PCR amplification of 15 genomic short tandem repeat (STR) loci is performed; patterns between donor, pre-transplant recipient, and post-transplant recipient are compared. The sizes of PCR products generated are used to infer their origin as recipient or donor. This allows for the post-transplant assessment of the donor contribution to the bone marrow production of the recipient. Chimerism is confirmed if recipient alleles are identified in the recipient’s post-transplant sample.

Blood: 4 mL EDTA is optimal (Minimum: 1 mL EDTA)
DNA: 100 μL at 200 ng/μL is optimal (Minimum: 30 μL at 200 ng/μL)
Bone Marrow: 0.5 mL marrow in EDTA is optimal (Minimum: 0.2 mL)

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays). 

Donor, pre-transplant recipient, and post-transplant recipient samples are required; consult with on-service Molecular Geneticist if these requirements cannot be met.

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  This test is only available to individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) and who meet test utilization guidelines or policy. For those without this coverage, contact the laboratory to discuss.

Test results should be interpreted in the context of clinical findings and other laboratory data.

Please refer to the BC Ministry of Health Guidelines and Gene Reviews for detailed background on the clinical features of HFE-HH.

Please refer to the BC Ministry of Health Guidelines and Gene Reviews for up-to-date information regarding the genetics of HFE-HH.

Genetic testing for HFE-HH is performed in accordance with BC Ministry of Health Guidelines.

There have been logistical changes to the ordering and laboratory algorithms (figures 1 and 2) that are not reflected in these Guidelines; the ordering algorithm (figure 1) has been updated here:

Changes to the approach to ordering

Please see HFE-HH Ordering Algorithm

All testing for HFE-HH should be ordered with the Standard Outpatient Laboratory Requisition (SOPLR), using the appropriate indications box under HFE-Hemochromatosis, as outlined in the algorithm.

The MGL general requisition is no longer used to order this test.

Changes to laboratory algorithm

Collection/Chemistry laboratories should follow their site-specific process for collection and processing, as established in consultation with BC chemistry laboratories and MGL.  This algorithm essentially follows figure 2 from the BC Ministry of Health Guidelines, with minor changes.

The C282Y mutation is detected by PCR-based RFLP analysis; this assay will not detect other rarer HFE alleles, including the H63D variant.

Blood: 0.5 – 4 mL EDTA

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays). 

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  Payment is not required when requests are made for individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) AND eligible for testing according to the test utilization guidelines / policy. If the individual undergoing testing is not insured by these providers or does not meet utilization guidelines or policy, please complete a billing form; testing will only commence after receipt of billing informationTest prices can be found here.

Molecular genetic testing is limited by the current understanding of the genome and the genetics of a particular disease, as well as by the method of detection used. This method will not detect all mutations (e.g., mutations outside the regions tested as described above, large genomic deletions, promoter mutations, regulatory element mutations).

Rare single nucleotide variants or polymorphisms could lead to false-negative results. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

A previous bone marrow transplant from an allogenic donor will result in molecular data that reflects the donor genotype rather than the recipient (patient) genotype. Consult the on-service Molecular Geneticist for approach to testing in such individuals.

Transfusions performed with packed red blood cells will generally not affect the outcome of molecular genetic testing. However, if there is no clinical urgency, the cautious approach is to wait one week post packed red cell transfusion before collecting a sample for genetic testing. Consult the on-service Molecular Geneticist as needed.

Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in our interpretation of results may occur if information given is inaccurate or incomplete.

Werdnig-Hoffmann Disease; Infantile Muscular Atrophy; Kugelberg-Welander Syndrome; Juvenile Muscular Atrophy; Proximal SMA

Spinal muscular atrophy (SMA) is an autosomal recessive condition characterized by progressive muscle weakness caused by the degeneration of anterior horn cells of the spinal cord and the brain stem nuclei. Onset ranges from before birth to young adulthood. Poor weight gain, sleep difficulties, pneumonia, scoliosis, and joint contractures are common complications. Subtypes include: SMA 0 (proposed name; also referred to as prenatal), with prenatal onset and severe joint contractures, facial diplegia, and respiratory failure; SMA 1, with onset before six months of age; SMA 2, with onset between six and 12 months; SMA3, with onset in childhood after 12 months; and SMA 4, with adult onset.

Two adjacent genes, SMN1 and SMN2, are associated with SMA. The two genes differ by only five base pairs and none of these base pair differences change the amino acids encoded by the genes. Nonetheless, the two genes do not encode identical proteins. SMN1 produces full-length transcripts while SMN2 primarily produces transcripts that lack exon 7 because one of the base pair changes in exon 7 disrupts SMN2 gene splicing.

SMN1 is the SMA disease gene. Approximately 95 – 98% of individuals with a clinical diagnosis of SMA are homozygous for an apparent deletion of exon 7 in SMN1. The remaining 2 – 5% are compound heterozygotes for an apparent deletion of exon 7 of SMN1 and an intragenic point mutation in SMN1.

The copy number of the SMN2 gene varies, ranging from zero to five. Although SMN2 does not produce the full length transcript with high efficiency, some full length transcript is produced. In some individuals with SMA who also have an increased copy number of the SMN2 gene, the small amount of full-length transcript generated SMN2 may help to produce a milder phenotype.

1. Confirmation of diagnosis:
   1. In individuals with clinical features suggestive of SMA.
2. Carrier testing:
   1. Adults at risk to be carriers of SMA due to a family history.
      NB: For the most accurate assessment of carrier status, please provide the results of SMN1 molecular analysis of the parents of the affected individual.
3. Prenatal testing (prenatal diagnosis requests are not normally accepted from physicians other than Medical Geneticists):
   1. Pregnancies at risk of SMA, where the parents have been confirmed by molecular analysis to each carry an SMN1 deletion.
      NB: If only one of the parents has been confirmed to be a carrier, contact the Molecular Geneticist on-service to discuss options.
4. Presymptomatic testing:
   1. Adults at risk of developing a milder form of SMA due to a family history confirmed to be due to SMN1 deletions.

The copy number of exons 7 and 8 of both the SMN1 and SMN2 genes is assessed by multiplex ligation-dependent probe amplification (MLPA) using the P060 probe mix (MRC-Holland).

Pregnancy-related/Prenatal

If pregnancy management will be altered, 3 weeks; otherwise, routine TAT.

Blood: 4 mL EDTA is optimal (Minimum: 1 mL EDTA)
DNA: NOT ACCEPTED

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays). 

Prenatal Specimens
Prenatal testing REQUIRES LABORATORY CONSULTATION PRIOR TO THE PROCEDURE and can only be ordered by a Medical Geneticist. Contact the laboratory at 604-875-2852 and choose the appropriate option for the Molecular Geneticist on service.
Chorionic Villi: 20 mg.
Direct Amniotic fluid: 25 mL collected in two separate tubes of equal volume.
Cultured Amniocytes: Two (2) 100% confluent T-25 flasks.

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth. Ship samples by overnight courier with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays) as follows:

• Villi – on wet ice or in media at room temperature
• Amniocytes, Amniotic fluid, DNA – at room temperature

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  Payment is not required when requests are made for individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) AND eligible for testing according to the test utilization guidelines / policy. If the individual undergoing testing is not insured by these providers or does not meet utilization guidelines or policy, please complete a billing form; testing will only commence after receipt of billing informationTest prices can be found here.

Molecular genetic testing is limited by the current understanding of the genome and the genetics of a particular disease, as well as by the method of detection used. This method will not detect all mutations (e.g., point mutations in the coding region, promoter mutations, and regulatory element mutations). In rare cases, a point mutation could be detected.

For carrier/predictive testing due to family history, it is generally important to first document the gene mutation in an affected or carrier family member. This information should be provided to the laboratory for assessment of whether the assay is appropriate for detection of the familial mutation, and to aid in the interpretation of data.

In some cases, DNA alterations of undetermined or unclear clinical significance may be identified.

Rare single nucleotide variants or polymorphisms could lead to false-negative results. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

A previous bone marrow transplant from an allogenic donor will result in molecular data that reflects the donor genotype rather than the recipient (patient) genotype. Consult the on-service Molecular Geneticist for approach to testing in such individuals.

Transfusions performed with packed red blood cells will generally not affect the outcome of molecular genetic testing. However, if there is no clinical urgency, the cautious approach is to wait one week post packed red cell transfusion before collecting a sample for genetic testing. Consult the on-service Molecular Geneticist as needed.

Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in our interpretation of results may occur if information given is inaccurate or incomplete.

Cystic fibrosis (CF) classically presents in infancy with clinical features that include chronic, debilitating lung infections and pancreatic insufficiency causing dietary malabsorption.

CFTR-related disorders (CFTR-RD) are defined as clinical entities associated with CFTR dysfunction that do not fulfill the diagnostic criteria for CF. This term has been ascribed to congenital bilateral absence of the vas deferens (CBAVD), recurrent pancreatitis, and disseminated bronchiectasis (PMID: 21658649). It is recommended that individuals with suspected CFTR-RD should have CFTR genetic testing performed in conjunction with sweat testing.

CF and CFTR-RD are autosomal recessive disorders caused by pathogenic variants in the CFTR gene, which encodes a chloride ion channel in epithelial cells. Over 2,000 different variants have been identified in CFTR; not all cause clinical symptoms and most are rare, with the exception of the CF-causing variant F508del which comprises approximately 70% of CF-causing alleles in individuals of Northern European ancestry.

1. Confirmation of diagnosis: The sweat chloride test is the gold standard test for confirming a diagnosis of cystic fibrosis (CF), and is recommended prior to or in conjunction with genetic testing in the investigation of a CFTR related disorder (CFTR-RD), depending on the clinical presentation.  Therefore, genetic testing should generally only be performed following or in conjunction with sweat testing except when:
   1. A sweat chloride test is not easily obtained (e.g., newborn with meconium ileus) OR
   2. A male has documented evidence of CBAVD and his partner is a known carrier of a CF-causing variant.
2. Carrier testing:
   1. Adults whose CF carrier risk due to a personal family history is greater than that of the general population OR their partner has a family history and CF carrier risk greater than that of the general population.
   2. Parents of a pregnancy where echogenic bowel has been detected on fetal ultrasound AND both of the parents are of an ancestry where the carrier frequency of CF is high and the detection rate of the assay is high (generally this applies to individuals of Northern-European ancestry or Ashkenazi Jewish ancestry).
3. Prenatal testing (prenatal diagnosis requests are not normally accepted from physicians other than Medical Geneticists):
   1. Pregnancies known to be at risk of CF AND the CF-causing variants segregating in the parents are known. 
4. Newborn screening:
   1. As part of the BC Newborn Screening Program, infants with an elevated immunoreactive trypsinogen (IRT) will undergo CFTR molecular genetic testing. These results are incorporated into the patient’s newborn screening report.

1. Population-based carrier screening for the purposes of reproductive planning is not covered by Health Insurance BC (BC MSP).
2. Testing of individuals with infertility who do not meet clinical criteria for CFTR-related CBAVD is not covered by Health Insurance BC (BC MSP)

One hundred and thirty (130) variants classified as CF-causing by the CFTR2 project are assessed using the MiSeqDx Cystic Fibrosis 139-Variant Assay (Illumina, Inc).   The length of the poly-T tract of intron 8 is reported according to published guidelines.  

The list of variants and the associated quality metrics are available here. 

If the clinical suspicion of CF is high, and two CF-causing variants are not identified by the targeted 130 variant assay, an expanded panel of variants or full gene sequencing may be performed; the clinical report methodology will indicate the analysis performed.

The list of expanded panel variants and the associated quality metrics are available here.

The target regions covered by full gene sequencing and the associated assay quality metrics are available here.

Pregnancy-related/Prenatal

3 weeks

Blood: 4 mL EDTA is optimal (Minimum: 1 mL EDTA)
DNA: 100 μL at 200 ng/μL is optimal (Minimum: 30 μL at 200 ng/μL)

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays).  

Prenatal Specimens
Prenatal testing REQUIRES LABORATORY CONSULTATION PRIOR TO THE PROCEDURE and can only be ordered by a Medical Geneticist. Contact the laboratory at 604-875-2852 and choose the appropriate option for the Molecular Geneticist on service.
Chorionic Villi: 20 mg.
Direct Amniotic fluid: 25 mL collected in two separate tubes of equal volume.
Cultured Amniocytes: Two (2) 100% confluent T-25 flasks.
DNA extracted from prenatal specimens: 100 μL at 200 ng/μL is optimal (Minimum: 30 μL at 200 ng/μL)

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth. Ship samples by overnight courier with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays) as follows:

• Villi – on wet ice or in media at room temperature
• Amniocytes, Amniotic fluid, DNA – at room temperature

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  Payment is not required when requests are made for individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) AND eligible for testing according to the test utilization guidelines / policy. If the individual undergoing testing is not insured by these providers or does not meet utilization guidelines or policy, please complete a billing form; testing will only commence after receipt of billing informationTest prices can be found here.

Molecular genetic testing is limited by the current understanding of the genome and the genetics of a particular disease, as well as by the method of detection used.

Rare single nucleotide variants or polymorphisms could lead to false-negative or false-positive results.  Some genetic abnormalities may not be detected by this assay including: some insertions and deletions, copy number variants, and chromosomal rearrangements.  This test cannot reliably detect mosaicism.  If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist

A previous bone marrow transplant from an allogenic donor will result in molecular data that reflects the donor genotype rather than the recipient (patient) genotype. Consult the on-service Molecular Geneticist for approach to testing in such individuals.

Transfusions performed with packed red blood cells will generally not affect the outcome of molecular genetic testing. However, if there is no clinical urgency, the cautious approach is to wait one week post packed red cell transfusion before collecting a sample for genetic testing. Consult the on-service Molecular Geneticist as needed.

Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in our interpretation of results may occur if information given is inaccurate or incomplete.

Huntington Chorea

Huntington disease (HD) is a progressive disorder of motor, cognitive, and psychiatric disturbances; onset is usually between 35 and 45 years of age. Early manifestations can include subtle changes in eye movements, coordination, minor involuntary movements, difficulty in mental planning, and a depressed or irritable mood. These evolve into more prominent chorea, with voluntary activity becoming increasingly difficult, and worsening dysarthria and dysphagia. The late stages are characterized by severe motor disability.

HD is caused exclusively by a trinucleotide (CAG) repeat expansion mutation in the HTT gene. The inheritance pattern is autosomal dominant with anticipation. Anticipation is the phenomenon of increased severity or decreased age of onset in successive generation due to expansion of the unstable repeat. In HD, anticipation is generally greater with paternal transmission of the expanded allele.

Alleles in the HTT gene are classified as:

• Normal: < 26 CAG repeats
• Normal (intermediate): 27 to 35 CAG repeats
• HD-causing with reduced penetrance: 36 to 39 repeats
• HD-causing with full penetrance: > 40 repeats

Normal (intermediate) alleles are not disease causing, but may be at risk of expansion into the pathogenic range in subsequent generations.  Alleles of 36-39 repeats show reduced penetrance; an individual with an allele in this range may or may not develop symptoms of Huntington disease during their lifetime.  Expanded alleles >100 repeats have been reported for infantile/juvenile onset HD.

1. Confirmation of diagnosis:
   1. In individuals with clinical features suggestive of HD.
2. Prenatal testing (prenatal diagnosis requests are not normally accepted from physicians other than Medical Geneticists):
   1. Pregnancies at risk of being affected with HD. Samples from both parents may be required to complete the prenatal diagnosis analysis.
3. Presymptomatic testing:
   1. Adults known to be at risk of developing symptoms due to a molecularly confirmed family history. Predictive testing will only be performed following genetic counselling by a recognized genetic service.

Sizing of the CAG repeat is performed on an ABI genetic analyzer following fluorescence-based PCR amplification. To aid in interpretation, PCR amplification is also performed to size the adjacent non-pathogenic CCG repeat, and the combined CAG/CCG repeat region. As required, triplet-primed (tp) PCR is performed.

Pregnancy-related/Prenatal

If pregnancy management will be altered, 3 weeks; otherwise, routine TAT.

Blood: 4 mL EDTA is optimal (Minimum: 1 mL EDTA)
DNA: 100 μL at 200 ng/μL is optimal (Minimum: 30 μL at 200 ng/μL)

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays). 

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  Payment is not required when requests are made for individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) AND eligible for testing according to the test utilization guidelines / policy. If the individual undergoing testing is not insured by these providers or does not meet utilization guidelines or policy, please complete a billing form; testing will only commence after receipt of billing informationTest prices can be found here.

Molecular genetic testing is limited by the current understanding of the genome and the genetics of a particular disease, as well as by the method of detection used. 

For carrier/predictive testing due to family history, it is generally important to first document the gene mutation in an affected or carrier family member. This information should be provided to the laboratory for assessment of whether the assay is appropriate for detection of the familial mutation, and to aid in the interpretation of data.

In some cases, DNA alterations of undetermined or unclear clinical significance may be identified.

In certain scenarios of repeat size mosaicism, false negative results may occur. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

Rare single nucleotide variants or polymorphisms could lead to false-negative or false-positive results. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

A previous bone marrow transplant from an allogenic donor will result in molecular data that reflects the donor genotype rather than the recipient (patient) genotype. Consult the on-service Molecular Geneticist for approach to testing in such individuals.

Transfusions performed with packed red blood cells will generally not affect the outcome of molecular genetic testing. However, if there is no clinical urgency, the cautious approach is to wait one week post packed red cell transfusion before collecting a sample for genetic testing. Consult the on-service Molecular Geneticist as needed.

Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in our interpretation of results may occur if information given is inaccurate or incomplete.

Kennedy Disease; X-linked Bulbospinal Neuropathy

Spinobulbar muscular atrophy (SBMA) is characterized by adolescent-onset mild androgen insensitivity (e.g., gynecomastia, small testes, oligo- or azoospermia) in males. This is followed by post-adolescent onset (age 20 – 50) proximal muscle weakness, fasciculations, and atrophy due to lower motor neuron degeneration. Eventually, most individuals with SBMA will have bulbar involvement. Life expectancy is not reduced. Carrier females are unaffected.

SBMA is caused by expansion of a CAG trinucleotide repeat in exon 1 of the androgen receptor (AR) gene on the X-chromosome. Inheritance is X-linked recessive. In general, repeat size roughly corresponds to the severity of disease and age of onset. Alleles in the AR gene are classified as:

• Normal: ≤ 34 repeats
• Questionable Significance: 35 repeats
  There is no consensus regarding the significance of an allele with 35 repeats
• Reduced Penetrance: 36 – 37 repeats
  It has been suggested that alleles with 36 or 37 repeats are reduced penetrance alleles, although this remains unclear.
• Full Penetrance: ≥ 38 repeats

1. Confirmation of diagnosis:
   1. In males with clinical features suggestive of SBMA.
2. Carrier Testing:
   1. Adult females at risk to be carriers of SBMA due to a family history. NB: Daughters of affected male individuals are obligate carriers of SBMA.
3. Prenatal testing (technically feasible, but not routinely performed – contact MGL to discuss):
   1. Male pregnancies at risk of SBMA. Prior to testing for SMBA, fetal sexing is performed; if the fetus is female, further testing is not indicated.
4. Presymptomatic testing:
   1. Adult males at risk of developing SBMA due to a family history. Predictive testing will only be performed following genetic counselling by a recognized genetic service.

The CAG repeat size is determined using an ABI genetic analyzer following fluorescence-based PCR amplification.

Blood: 4 mL EDTA is optimal (Minimum: 1 mL EDTA)
DNA: 100 μL at 200 ng/μL is optimal (Minimum: 30 μL at 200 ng/μL)

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays). 

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  Payment is not required when requests are made for individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) AND eligible for testing according to the test utilization guidelines / policy. If the individual undergoing testing is not insured by these providers or does not meet utilization guidelines or policy, please complete a billing form; testing will only commence after receipt of billing informationTest prices can be found here.

Molecular genetic testing is limited by the current understanding of the genome and the genetics of a particular disease, as well as by the method of detection used. This method will not detect all mutations (e.g., point mutations in the coding region of the gene, large genomic deletions, promoter mutations, regulatory element mutations). For some trinucleotide repeat disorders, repeat expansions have been described that cannot be amplified by PCR. Consideration should be given to this particularly in cases with severe clinical features or early onset; consult the on-service Molecular Geneticist to discuss specific repeat disorders.

For carrier/predictive testing due to family history, it is generally important to first document the gene mutation in an affected or carrier family member. This information should be provided to the laboratory for assessment of whether the assay is appropriate for detection of the familial mutation, and to aid in the interpretation of data.

In some cases, DNA alterations of undetermined or unclear clinical significance may be identified.

In certain scenarios of repeat size mosaicism, false negative results may occur. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

Rare single nucleotide variants or polymorphisms could lead to false-negative or false-positive results. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

A previous bone marrow transplant from an allogenic donor will result in molecular data that reflects the donor genotype rather than the recipient (patient) genotype. Consult the on-service Molecular Geneticist for approach to testing in such individuals.

Transfusions performed with packed red blood cells will generally not affect the outcome of molecular genetic testing. However, if there is no clinical urgency, the cautious approach is to wait one week post packed red cell transfusion before collecting a sample for genetic testing. Consult the on-service Molecular Geneticist as needed.

Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in our interpretation of results may occur if information given is inaccurate or incomplete.

Duchenne Muscular Dystrophy; Becker Muscular Dystrophy; DMD-Related Dilated Cardiomyopathy

The dystrophinopathies manifest as a spectrum of muscle diseases. The mildest of the phenotypes includes an asymptomatic increase in serum concentration of creatine phosphokinase (CK or CpK), muscle cramps with myoglobinuria, and isolated quadriceps myopathy. At the opposite end of the spectrum is Duchenne muscular dystrophy (DMD), usually presenting in early childhood with delay in the motor milestones. DMD is rapidly progressive; patients are usually wheelchair-bound by 12 years of age and death usually occurs before age 30, due most frequently to respiratory complications and/or cardiomyopathy. Becker muscular dystrophy (BMD) is characterized by later-onset skeletal muscle weakness; individuals with BMD usually remain ambulatory well into their 20s. Despite the milder skeletal muscle involvement in BMD, cardiomypathy is a common cause of morbidity and the most common cause of death (on average in the mid-40s). Finally, DMD-associated dilated cardiomyopathy (DCM) is characterized by left ventricular dilation and congestive heart failure. Female carriers of DMD mutations are at increased risk for cardiomyopathy.

The dystrophinopathies are due to mutations in dystrophin (DMD), an X-linked gene encoding a membrane-associated protein that is found in muscle and a subset of neurons. The Duchenne phenotype is almost invariably caused by mutations that disrupt the reading frame including: deletions or duplications; nonsense mutations, and splice-site mutations. These produce a dystrophin protein molecule that is degraded. The milder Becker phenotype, on the other hand, results from mutations that reduce but do not completely eliminate the production of functional dystrophin protein, including deletions or duplications that maintain the open reading frame of the transcript, some splicing mutations, and most non-truncating single-base changes that result in translation of a protein product with intact N and C termini. DMD-associated dilated cardiomyopathy is caused by mutations in DMD that affect the muscle promoter (PM) and the first exon (E1), resulting in no dystrophin transcript being produced in cardiac muscle; expression (under different promoters) is retained in skeletal muscle and the central nervous system.

1. Confirmation of diagnosis:
   1. Testing of males with a suspected diagnosis of DMD.
   2. Testing of females is warranted if there is a clinical presentation consistent with the disease.
2. Carrier testing:
   1. Testing of adult females at risk to be carriers because of a family history. NB: Carriers have the potential for health problems in addition to the ability to transmit disease to offspring; genetic counselling is recommended prior to testing.
3. Prenatal testing (prenatal diagnosis requests are not normally accepted from physicians other than Medical Geneticists):
   1. Pregnancies at risk of inheriting a known DMD deletion or duplication. Prior to testing for the DMD mutation, fetal sexing is performed; if the fetus is female, further testing is not indicated.
      NB: If the mutation segregating in the mother is not known, consult the on-service Molecular Geneticist for assessment of whether linkage analysis is available for prenatal diagnosis
4. Presymptomatic testing:
   1. Requests for presymptomatic testing are only accepted following genetic counselling by a recognized genetic service.

Multiplex ligation-dependant probe amplification (MLPA) is carried out with the P034-A2 and P035-A2 probe mixes (MRC-Holland) to detect whole exon deletions and duplications; each of the 79 exons of DMD and the alternate exon 1 (DP427c) are assessed.

Pregnancy-related/Prenatal

If pregnancy management will be altered, 3 weeks; otherwise, routine TAT.

Blood: 4 mL EDTA is optimal (Minimum: 1 mL EDTA)
DNA: NOT ACCEPTED

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth and ship to the address below. Samples should be shipped at room temperature with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays). 

Prenatal Specimens
Prenatal testing REQUIRES LABORATORY CONSULTATION PRIOR TO THE PROCEDURE and can only be ordered by a Medical Geneticist. Contact the laboratory at 604-875-2852 and choose the appropriate option for the Molecular Geneticist on service.
Chorionic Villi: 20 mg.
Direct Amniotic fluid: 25 mL collected in two separate tubes of equal volume.
Cultured Amniocytes: Two (2) 100% confluent T-25 flasks.

Label each sample with three patient identifiers; preferably patient name, PHN, and date of birth. Ship samples by overnight courier with a completed MGL Requisition to arrive Monday to Friday (not on Canadian statutory holidays) as follows:

• Villi – on wet ice or in media at room temperature
• Amniocytes, Amniotic fluid, DNA – at room temperature

Testing is only available to residents of Canada, except in very specific circumstances where testing is urgent or emergent.  Payment is not required when requests are made for individuals who are insured by Health Insurance BC (administered through the BC Medical Services Plan (MSP)) AND eligible for testing according to the test utilization guidelines / policy. If the individual undergoing testing is not insured by these providers or does not meet utilization guidelines or policy, please complete a billing form; testing will only commence after receipt of billing informationTest prices can be found here.

Molecular genetic testing is limited by the current understanding of the genome and the genetics of a particular disease, as well as by the method of detection used. This method will not detect all mutations (e.g., point mutations in the coding region, promoter mutations, and regulatory element mutations). In rare cases, a point mutation could be detected.

For carrier/predictive testing due to family history, it is generally important to first document the gene mutation in an affected or carrier family member. This information should be provided to the laboratory for assessment of whether the assay is appropriate for detection of the familial mutation, and to aid in the interpretation of data.

In some cases, DNA alterations of undetermined or unclear clinical significance may be identified.

Rare single nucleotide variants or polymorphisms could lead to false-negative results. If results obtained do not match the clinical findings, consult the on-service Molecular Geneticist.

A previous bone marrow transplant from an allogenic donor will result in molecular data that reflects the donor genotype rather than the recipient (patient) genotype. Consult the on-service Molecular Geneticist for approach to testing in such individuals.

Transfusions performed with packed red blood cells will generally not affect the outcome of molecular genetic testing. However, if there is no clinical urgency, the cautious approach is to wait one week post packed red cell transfusion before collecting a sample for genetic testing. Consult the on-service Molecular Geneticist as needed.

Test results should be interpreted in the context of clinical findings, family history, and other laboratory data. Errors in our interpretation of results may occur if information given is inaccurate or incomplete.