| 1. |
- Jahangir Tafrechi, Roshan S., et al.
(författare)
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Single-cell A3243G mitochondrial DNA mutation load assays for segregation analysis
- 2007
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Ingår i: Journal of Histochemistry and Cytochemistry. - 0022-1554 .- 1551-5044. ; 55:11, s. 1159-1166
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Tidskriftsartikel (refereegranskat)abstract
- Segregation of mitochondrial DNA (mtDNA) is an important underlying pathogenic factor in mtDNA mutation accumulation in mitochondrial diseases and aging, but the molecular mechanisms of mtDNA segregation are elusive. Lack of high-throughput single-cell mutation load assays lies at the root of the paucity of studies in which, at the single-cell level, mitotic mtDNA segregation patterns have been analyzed. Here we describe development of a novel fluorescence-based, non-gel PCR restriction fragment length polymorphism method for single-cell A3243G mtDNA mutation load measurement. Results correlated very well with a quantitative in situ Padlock/rolling circle amplification–based genotyping method. In view of the throughput and accuracy of both methods for single-cell A3243G mtDNA mutation load determination, we conclude that they are well suited for segregation analysis.
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| 2. |
- Allalou, Amin, et al.
(författare)
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Image Based Measurements of Single Cell mtDNA Mutation Load
- 2007
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Ingår i: Image Analysis, Proceedings. - Berlin, Heidelberg : Springer Berlin Heidelberg. - 9783540730392 ; , s. 631-640
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Konferensbidrag (refereegranskat)abstract
- Cell cultures as well as cells in tissue always display a certain degree of variability, and measurements based on cell averages will miss important information contained in a heterogeneous population. This paper presents automated methods for image based measurements of mitochondiral DNA (mtDNA) mutations in individual cells. The mitochondria are present in the cell’s cytoplasm, and each cytoplasm has to be delineated. Three different methods for segmentation of cytoplasms are compared and it is shown that automated cytoplasmic delineation can be performed 30 times faster than manual delineation, with an accuracy as high as 87%. The final image based measurements of mitochondrial mutation load are also compared to, and show high agreement with, measurements made using biochemical techniques.
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