Mitochondrial Diseases
Mitochondrial genome sequencing

Lab method: Next generation sequencing
Heteroplasmy less than 20% is not detectable by sequencing.
TAT: 2-4 weeks
Specimen requirements: 2-4 ml of blood with anticoagulant EDTA

1 µg DNA in TE, AE or pure sterile water at 100-250 ng/µl
The A260/A280 ratio should be 1.8-2.0. DNA sample should be run on an agarose gel as a single band, showing no degradation, alongside with a quantitative DNA marker.

50-75 mg fresh frozen tissue (in case suspected mtDNA mutations may not be detected in DNA extracted from blood)
Tissue should be frozen immediately at collection, stored at -80°C and shipped on dry ice.
Ordering information: Go to online ordering or download sample submission form

Nuclear genes of NGS panel

Genes
(full
coding region):		 AARS2, AASS, ABAT, ABCB6, ABCB7, ABCD1, ABCD3, ACACA, ACAD8, ACAD9, ACADL, ACADM, ACADS, ACADSB, ACADVL, ACAT1, ACO2, ACOX1, ACSF3, ACSL4, AFG3L2, AGK, AGXT, AIFM1, AK2, ALAS2, ALDH18A1, ALDH2, ALDH3A2, ALDH4A1, ALDH5A1, ALDH6A1, ALDH7A1, AMACR, AMT, APTX, ATIC, ATP5F1A, ATP5F1E, ATP7B, ATPAF2, ATXN2, AUH, BAX, BCKDHA, BCKDHB, BCKDK, BCL2, BCS1L, BOLA3, BRIP1, BTD, C12orf65, C19orf12, CA5A, CARS2, CASP8, CAT, CAVIN1, CEL, CHCHD10, CISD2, CLPB, CLPP, COA3, COA5, COA6, COA8 (APOPT1), COASY, COMT, COQ2, COQ4, COQ6, COQ8A, COQ8B, COQ9, COX10, COX14, COX15, COX20, COX4I1, COX4I2, COX6A1, COX6A2, COX6B1, COX7B, COX8A, CPOX, CPS1, CPT1A, CPT1C, CPT2, CRBN, CYB5A, CYB5R3, CYC1, CYCS, CYP11A1, CYP11B1, CYP11B2, CYP24A1, CYP27A1, CYP27B1, D2HGDH, DARS2, DBT, DGUOK, DHCR24, DHODH, DHTKD1, DIABLO, DLAT, DLD, DMGDH, DMPK, DNA2, DNAJC19, DNAJC3, DNM1L, EARS2, ECHS1, ELAC2, EPHX2, ETFA, ETFB, ETFDH, ETHE1, FAH, FARS2, FASTKD2, FBP1, FBXL4, FDX2 (FDX1L), FECH, FH, FKBP10, FLAD1, FOXRED1, FTH1, FXN, G6PC, GAMT, GARS1 (GARS), GATM, GCDH, GDAP1, GFER, GFM1, GFM2, GK, GLDC, GLRX5, GLUD1, GLYCTK, GPI, GPT2, GPX1, GRHPR, GSR, GTPBP3, GYS2, HADH, HADHA, HAMP, HARS2, HAX1, HCCS, HIBCH, HINT1, HK1, HLCS, HMBS, HMGCL, HMGCS2, HOGA1, HSD17B10, HSD17B4, HSD3B2, HSPA9, HSPD1, HTRA2, IARS2, IBA57, IDH2, IDH3B, ISCA2, ISCU, IVD, KARS1, KIF1B, KRT5, L2HGDH, LAMP2, LARS1, LARS2, LIAS, LIPT1, LONP1, LRPPRC, LYRM4, LYRM7, MAOA, MARS2, MCCC1, MCCC2, MCEE, MFF, MFN2, MGME1, MICU1, MIP, MLYCD, MMAA, MMAB, MMACHC, MMADHC, MMUT, MOCS1, MPC1, MPV17, MRPL12, MRPL3, MRPL44, MRPS16, MRPS22, MRPS7, MSRB3, MTFMT, MTO1, MTPAP, MTRR, NADK2, NAGS, NARS2, NBAS,NDUFA1, NDUFA10, NDUFA11, NDUFA12, NDUFA2, NDUFA4, NDUFA9, NDUFAF1, NDUFAF2, NDUFAF3, NDUFAF4, NDUFAF5, NDUFAF6, NDUFAF7, NDUFB11, NDUFB3, NDUFB9, NDUFS1, NDUFS2, NDUFS3, NDUFS4, NDUFS6, NDUFS7, NDUFS8, NDUFV1, NDUFV2, NFS1, NFU1, NNT, NR2F1, NTHL1, NUBPL, OAT, OGDH, OGG1, OPA1, OPA3, OTC, OXCT1, P4HB, PAM16, PANK2, PARK7, PARS2, PC, PCCA, PCCB, PCK2, PDHA1, PDHB, PDHX, PDK3, PDP1, PDSS1, PDSS2, PDX1, PET100, PEX11B, PHYH, PINK1, PKLR, PNPLA8, PNPO, PNPT1, POLG, POLG2, PPOX, PRODH, PTRH2, PTS, PUS1, PYCR1, PYCR2, QDPR, RARS2, RDH11, REEP1, RMND1, RNASEH1, RNASEL, RPIA, RPL35A, RPS14, RRM2B, SARS2, SCO1, SCO2, SDHA, SDHAF1, SDHAF2, SECISBP2, SERAC1, SETX, SFXN4, SLC16A1, SLC19A3, SLC25A12, SLC25A13, SLC25A15, SLC25A19, SLC25A20, SLC25A22, SLC25A26, SLC25A3, SLC25A38, SLC25A4, SLC25A46, SLC37A4, SLC6A8, SLC9A6, SNAP29, SOD1, SOD2, SPAST, SPG7, SPR, SPTLC2, SUCLA2, SUCLG1, SUGCT, SUOX, SURF1, TACO1, TARS2, TAZ, TCIRG1, TFAM, TFR2, TIMM8A, TK2, TMEM126A, TMEM126B, TMEM70, TMLHE, TPI1, TPK1, TRIT1, TRMT10C, TRMU, TRNT1, TSFM, TTC19, TUBB3, TUFM, TWNK, TYMP, UNG, UQCC2, UQCC3, UQCRB, UQCRC2, UQCRQ, VARS2, WDR81, WFS1, XPNPEP3, YARS2

List of diseases covered by the panel
Lab method: NGS panel with CNV analysis
TAT: 6-9 weeks
Specimen requirements: 2-4 ml of blood with anticoagulant EDTA

1 µg DNA in TE, AE or pure sterile water at 100-250 ng/µl
The A260/A280 ratio should be 1.8-2.0. DNA sample should be run on an agarose gel as a single band, showing no degradation, alongside with a quantitative DNA marker.
Ordering information: Go to online ordering or download sample submission form

Single gene sequencing

Genes: ACADS, ACADVL
Lab method: Sanger sequencing, next generation sequencing
TAT: 2-4 weeks
Specimen requirements: 2-4 ml of blood with anticoagulant EDTA

2 µg DNA in TE, AE or pure sterile water at 100-250 ng/µl
The A260/A280 ratio should be 1.8-2.0. DNA sample should be run on an agarose gel as a single band, showing no degradation, alongside with a quantitative DNA marker.
Ordering information: Go to online ordering or download sample submission form

MELAS Syndrome targeted mutation analysis

Genes: MT-TL1
No of
detectable
markers:		 1 (m.3243A>G)
Lab method: Sanger sequencing
TAT: 1-2 weeks
Specimen requirements: 2-4 ml of blood with anticoagulant EDTA

120 ng DNA in TE, AE or pure sterile water at 100-250 ng/µl
The A260/A280 ratio should be 1.8-2.0. DNA sample should be run on an agarose gel as a single band, showing no degradation, alongside with a quantitative DNA marker.
Ordering information: Go to online ordering or download sample submission form

Deletion/duplication analysis

Genes: DGUOK, MPV17, POLG, POLG2, RRM2B, SLC25A4, SUCLA2, SUCLG1, TK2, TWNK
Lab method: MLPA
TAT: 4-6 weeks
Specimen requirements: 2-4 ml of blood with anticoagulant EDTA

2 µg DNA in TE, AE or pure sterile water at 100-250 ng/µl
The A260/A280 ratio should be 1.8-2.0. DNA sample should be run on an agarose gel as a single band, showing no degradation, alongside with a quantitative DNA marker.
Ordering information: Go to online ordering or download sample submission form

Indications for genetic testing:

1. Diagnosis of patients with phenotype characteristic for mitochondrial disease
2. Diagnosis of patients with family history suggestive for mitochondrial disease
3. Genetic counseling of individuals with mitochondrial disease and affected family members

Mitochondrial diseases are a genetically and clinically heterogeneous group of disorders that arise as a consequence of dysfunction of the mitochondrial respiratory chain. The estimate for the prevalence of all mitochondrial disorders 1:8500, but they are thought to be greatly under-diagnosed. Mitochondrial disorders can be caused by mutations of nuclear or mitochondrial DNA (mtDNA). If nuclear gene defects may be inherited in an autosomal recessive or autosomal dominant manner, mtDNA defects are transmitted only maternally. As the female could have heteroplasmic mtDNA mutations, which could be transmitted unequally to her offspring, the sibs could exhibit considerable clinical variability.

Symptoms of the mitochondrial disease can begin at any age. Mitochondrial disorders may affect a single organ (e.g. Leber hereditary optic neuropathy, LHON) or involve multiple organ systems (e.g. Myoclonic epilepsy with ragged-red fibers, MERRF). Common clinical features of mitochondrial disorder include, for example muscle weakness, exercise intolerance, trouble with balance and coordination, sensorineural deafness, impaired vision, seizures and learning deficits, cardiomyopathy, diabetes mellitus, stunted growth, and a high incidence of mid- and late pregnancy loss.

References:

Wallace DC. Mitochondrial diseases in man and mouse. Science. 1999;283:1482–8.
Chinnery PF. Mitochondrial Disorders Overview. Pagon RA, Adam MP, Bird TD, et al., editors. Seattle (WA): University of Washington, Seattle; 1993-2013.
DiMauro S, Schon EA. Nuclear power and mitochondrial disease. Nat Genet. 1998;19:214–5.
Leonard JV, Schapira AVH. Mitochondrial respiratory chain disorders I: mitochondrial DNA defects. Lancet. 2000a;355:299–304.
Leonard JV, Schapira AVH. Mitochondrial respiratory chain disorders II: neurodegenerative disorders and nuclear gene defects. Lancet. 2000b;355:389–94.