Diagnosis of mitochondrial disorders applying massive pyrosequencing
- 233 Downloads
Mitochondrial disorders are a frequent cause of neurological disability affecting children and adults. Traditionally, molecular diagnosis of mitochondrial diseases was mostly accomplished by the use of Sanger sequencing and PCR–RFLP. However, there are particular drawbacks associated with the use of these methods. Recent multidisciplinary advances have led to new sequencing methods that may overcome these limitations. Our goal was to explore the use of a next generation sequencing platform in the molecular diagnosis of mitochondrial diseases reporting our findings in adult patients that present with a clinical-pathological diagnosis of a mitochondrial encephalomyopathy. Complete genomic sequences of mitochondrial DNA were obtained by 454 massive pyrosequencing from blood samples. The analysis of these sequences allowed us to identify two diagnostic pathogenic mutations and 74 homoplasmic polymorphisms, useful for obtaining high-resolution mitochondrial haplogroups. In summary, molecular diagnosis of mitochondrial disorders could be efficiently done from readily accessible samples, such as blood, with the use of a new sequencing platform.
KeywordsMitochondrial disorders Genetics Next generation sequencing
This study was supported by a grant from the Argentine National Research Council (CONICET). MAK, MV and SK are members of the research career of CONICET.
Conflict of interest
The authors declare that they have no competing interests.
- 3.Poulton J, Chiaratti MR, Meirelles FV, Kennedy S, Wells D, Holt IJ (2010) Transmission of mitochondrial DNA diseases and ways to prevent them. PLoS Genet 6(8):e1001066. doi: 10.1371/journal.pgen.1001066
- 12.Margulies M, Egholm M, Altman WE, Attiya S, Bader JS, Bemben LA, Berka J, Braverman MS, Chen YJ, Chen Z, Dewell SB, Du L, Fierro JM, Gomes XV, Godwin BC, He W, Helgesen S, Ho CH, Irzyk GP, Jando SC, Alenquer ML, Jarvie TP, Jirage KB, Kim JB, Knight JR, Lanza JR, Leamon JH, Lefkowitz SM, Lei M, Li J, Lohman KL, Lu H, Makhijani VB, McDade KE, McKenna MP, Myers EW, Nickerson E, Nobile JR, Plant R, Puc BP, Ronan MT, Roth GT, Sarkis GJ, Simons JF, Simpson JW, Srinivasan M, Tartaro KR, Tomasz A, Vogt KA, Volkmer GA, Wang SH, Wang Y, Weiner MP, Yu P, Begley RF, Rothberg JM (2005) Genome sequencing in microfabricated high-density picolitre reactors. Nature 437(7057):376–380. doi: 10.1038/nature03959 PubMedGoogle Scholar
- 13.Ramos A, Santos C, Alvarez L, Nogues R, Aluja MP (2009) Human mitochondrial DNA complete amplification and sequencing: a new validated primer set that prevents nuclear DNA sequences of mitochondrial origin co-amplification. Electrophoresis 30(9):1587–1593. doi: 10.1002/elps.200800601 PubMedCrossRefGoogle Scholar
- 19.Remes AM, Karppa M, Moilanen JS, Rusanen H, Hassinen IE, Majamaa K, Uimonen S, Sorri M, Salmela PI, Karvonen SL (2003) Epidemiology of the mitochondrial DNA 8344A > G mutation for the myoclonus epilepsy and ragged red fibres (MERRF) syndrome. J Neurol Neurosurg Psychiatry 74(8):1158–1159PubMedCrossRefGoogle Scholar