Single lymphocytes from two healthy individuals with mitochondrial point heteroplasmy are mainly homoplasmic

Lutz-Bonengel, Sabine; Sänger, Timo; Parson, Walther; Müller, Helena; Ellwart, Joachim W.; Follo, Marie; Bonengel, Bernhard; Niederstätter, Harald; Heinrich, Marielle; Schmidt, Ulrike

doi:10.1007/s00414-007-0190-6

Cited by 16 publications

(13 citation statements)

References 46 publications

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“…Amplification and subsequent sequencing of mtDNA were performed as described previously [28] but with primers L15908/H16432, L16268/H159, L16450/H429 and L314/H611 for PCR and sequencing [29]. Also, restriction fragment length analysis of nucleotide (nt) positions 303-310 was performed as described earlier [27].…”

Section: Samplesmentioning

confidence: 99%

Analysis of mitochondrial length heteroplasmy in monozygous and non-monozygous siblings

et al. 2008

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The segregation of mitochondrial genomes and the inheritance of mitochondrial DNA are constant matters of debate. To obtain more information about this issue and to answer the question whether or not it is possible to distinguish mitochondrial DNA (mtDNA) samples from monozygous individuals by analysing heteroplasmic length variants, 290 monozygous and 121 dizygous twin pairs and 34 sets of multiples were studied by RFLP and partly by direct sequencing. A factor D describing the respective pattern of length variants in a given sample was also calculated. The results show that monozygous individuals exhibit a significantly lower median and closer distribution of D than non-monozygous siblings. Thus, a differentiation of mtDNA samples from monozygous twins by this trait is not possible. The high percentage of heteroplasmic individuals, the low median of the D values and the unexpectedly very similar distribution of length variants in monozygotic individuals support the existence of a relatively wide bottleneck or the assumption of a regeneration of length heteroplasmy following a tight bottleneck and agree with a random segregation of mtDNA genomes in dividing oocytes.

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Section: Samplesmentioning

confidence: 99%

Analysis of mitochondrial length heteroplasmy in monozygous and non-monozygous siblings

et al. 2008

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“…For example, Brandstätter and Parson demonstrated that the amplification strategy itself has a substantial impact on the appearance of heteroplasmy [26]. We also observed an interrelationship between the measured nucleotide ratios and the sequencing primers used [15], but other factors also have to be considered.…”

Section: Introductionmentioning

confidence: 68%

“…Moreover, low ratios of 1%, 2%, 3%, 4%, 7%, and 9% were created from 16093 C and 16093 T T lymphocytes to test the sensitivity of heteroplasmy detection. Instrument settings and the cell selection process were described previously [15], but selection of T lymphocytes was facilitated by fluorescent labeling. One row per plate was omitted to allow for positive and negative PCR and sequencing controls, respectively.…”

Section: Generation Of Artificial Heteroplasmy Using Cell Mixturesmentioning

confidence: 99%

“…However, their discrimination from sequence background, from phantom mutations [13], and from sample mixtures often is difficult, and the interpretation of mtDNA results might be complicated. The frequency of heteroplasmy in humans is still under debate [14][15][16][17]. Compounding the problem, the heteroplasmy level, i.e., the ratio of nucleotides, seems to vary within ill or healthy individuals [7,8,[16][17][18], as well as with age [19][20][21].…”

Section: Introductionmentioning

confidence: 99%

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Factors affecting the detection and quantification of mitochondrial point heteroplasmy using Sanger sequencing and SNaPshot minisequencing

et al. 2011

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Mitochondrial DNA analysis plays an important role in forensic science as well as in the diagnosis of mitochondrial diseases. The occurrence of two different nucleotides at the same sequence position can be caused either by heteroplasmy or by a mix of samples. The detection of superimposed positions in forensic samples and their quantification can provide additional information and might also be useful to identify a mixed sample. Therefore, the detection and visualization of heteroplasmy has to be robust and sensitive at the same time to allow for reliable interpretation of results and to avoid a loss of information. In this study, different factors influencing the analysis of mitochondrial heteroplasmy (DNA polymerases, PCR and sequencing primers, nucleotide incorporation, and sequence context) were examined. BigDye Sanger sequencing and the SNaPshot minisequencing were compared as to the accuracy of detection using artificially created mitochondrial DNA mixtures. Both sequencing strategies showed to be robust, and the parameters tested showed to have a variable impact on the display of nucleotide ratios. However, experiments revealed a high correlation between the expected and the measured nucleotide ratios in cell mixtures. Compared to the SNaPshot minisequencing, Sanger sequencing proved to be the more robust and reliable method for quantification of nucleotide ratios but showed a lower detection sensitivity of minor cytosine components.

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“…Two types of heteroplasmy are recognized in mitochondrial genome: point and length heteroplasmy (Bendall and Sykes, 1995;Bendall et al, 1996). Point (sequence or site) heteroplasmy is defined by sequences presenting different bases at the same nucleotide position, which are usually observed in a sequence electropherogram as a superimposition of two different bases at this nucleotide position (Lutz-Bonengel et al, 2008). Length heteroplasmy is identified by repetitive tracts of consecutive cytosines, and has been found in four regions within the mtDNA control region (Bendall and Sykes, 1995;Lee HY et al, 2004;Forster et al, 2010).…”

Section: Accepted M Manuscriptmentioning

confidence: 99%

The link between mitochondrial DNA hypervariable segment I heteroplasmy and ageing among genetically unrelated Latvians

Pliss

Brakmanis

Ranka

et al. 2011

Experimental Gerontology

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To cite this version:This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. A C C E P T E D M A N U S C R I P T ACCEPTED MANUSCRIPT A C C E P T E D M A N U S C R I P T ACCEPTED MANUSCRIPT AbstractVarious studies have demonstrated that mitochondrial DNA (mtDNA) heteroplasmy tends to increase with age and that the observed frequency of heteroplasmy among populations mostly depends on the way it is measured. Therefore, we investigated age-related association on the presence of mtDNA heteroplasmy within the hypervariable segment 1 (HVS-I) in a selected study group. The study group consisted of 300 maternally unrelated Latvians ranging in age from 18 to over 90 years. To determine the optimal method for mtDNA heteroplasmy detection, three The results indicate that heteroplasmy in HVS-I is relatively common and occurs in a broad spectrum of sites. The above is supported by evidence to eventual increase of the probability of heteroplasmy with age due to specific mitochondrial haplogroup background. Research HighlightsThe results indicate that heteroplasmy in HVS-I is relatively common and occurs in a broad spectrum of sites. Consequently, a strong association is found to exist between the mtDNA heteroplasmy and ageing in the studied population.We found that the Denaturing Gradient Gel Electrophoresis (DGGE) and the SURVEYOR TM Mutation Detection Kit assay exhibited a lower limit of detection (LOD) of mtDNA heteroplasmy and revealed the presence of heteroplasmy at the already existing 1% and 5% occurrence in the sample.

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Single lymphocytes from two healthy individuals with mitochondrial point heteroplasmy are mainly homoplasmic

Cited by 16 publications

References 46 publications

Analysis of mitochondrial length heteroplasmy in monozygous and non-monozygous siblings

Analysis of mitochondrial length heteroplasmy in monozygous and non-monozygous siblings

Factors affecting the detection and quantification of mitochondrial point heteroplasmy using Sanger sequencing and SNaPshot minisequencing

The link between mitochondrial DNA hypervariable segment I heteroplasmy and ageing among genetically unrelated Latvians

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