title: Detection of heteroplasmic mitochondrial DNA in single mitochondria.
abstract: BACKGROUND
  Mitochondrial DNA (mtDNA) genome mutations can lead to energy and respiratory-related disorders like myoclonic epilepsy with ragged red fiber disease (MERRF), mitochondrial myopathy, encephalopathy, lactic acidosis and stroke (MELAS) syndrome, and Leber's hereditary optic neuropathy (LHON). It is not well understood what effect the distribution of mutated mtDNA throughout the mitochondrial matrix has on the development of mitochondrial-based disorders. Insight into this complex sub-cellular heterogeneity may further our understanding of the development of mitochondria-related diseases.
  
  
  METHODS
  This work describes a method for isolating individual mitochondria from single cells and performing molecular analysis on that single mitochondrion's DNA. An optical tweezer extracts a single mitochondrion from a lysed human HL-60 cell. Then a micron-sized femtopipette tip captures the mitochondrion for subsequent analysis. Multiple rounds of conventional DNA amplification and standard sequencing methods enable the detection of a heteroplasmic mixture in the mtDNA from a single mitochondrion.
  
  
  CONCLUSIONS
  Molecular analysis of mtDNA from the individually extracted mitochondrion demonstrates that a heteroplasmy is present in single mitochondria at various ratios consistent with the 50/50 heteroplasmy ratio found in single cells that contain multiple mitochondria.
authors.affiliation: Physical Measurement Laboratory, National Institute of Standards and Technology, Gaithersburg, Maryland, United States of America. joseph.reiner@nist.gov
authors.forename: Joseph E
authors.initials: JE
authors.lastname: Reiner
authors.forename: Rani B
authors.initials: RB
authors.lastname: Kishore
authors.forename: Barbara C
authors.initials: BC
authors.lastname: Levin
authors.forename: Thomas
authors.initials: T
authors.lastname: Albanetti
authors.forename: Nicholas
authors.initials: N
authors.lastname: Boire
authors.forename: Ashley
authors.initials: A
authors.lastname: Knipe
authors.forename: Kristian
authors.initials: K
authors.lastname: Helmerson
authors.forename: Koren Holland
authors.initials: KH
authors.lastname: Deckman
mesh_terms: D003584:Cytological Techniques; D004247:DNA / Q000235:genetics; D004272:DNA, Mitochondrial / Q000235:genetics*; D018922:HL-60 Cells; D006801:Humans; D008928:Mitochondria / Q000378:metabolism*; D028361:Mitochondrial Diseases / Q000235:genetics; D017240:Mitochondrial Myopathies / Q000235:genetics; D008957:Models, Genetic; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA; D014466:Ultraviolet Rays
chemical_list: D004272:DNA, Mitochondrial; D004247:DNA