High-quality and high-throughput massively parallel sequencing of the human mitochondrial genome using the Illumina MiSeq

King, Jonathan L.; LaRue, Bobby; Novroski, Nicole M.M.; Stoljarova, Monika; Seo, Seung Beom; Zeng, Xiangpei; Warshauer, David H.; Davis, Carey; Parson, Walther; Sajantila, Antti; Budowle, Bruce

doi:10.1016/j.fsigen.2014.06.001

Cited by 153 publications

(155 citation statements)

References 41 publications

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“…Of these mtDNA profiles, 100 (87.7 %) were unique within the data set, and 12.3 % of sequenced mtGenomes were observed twice. Compared to the previous whole mtGenome population studies [6,14], the number of shared haplotypes in the current study is relatively high. This might be a reflection of lower mtGenome diversity in Estonia or just may arise from sampling variance.…”

Section: Resultsmentioning

confidence: 69%

“…The quantity of extracted DNA was determined using the Qubit® dsDNA HS (High Sensitivity) Quantification Kit and a Qubit® 2.0 Fluorometer (Life Technologies, Foster City, CA, USA). Amplification of the mtGenome was accomplished as described by King et al [6].…”

Section: Sample Preparation and Target Amplificationmentioning

confidence: 99%

“…The MiSeq re-sequencing protocol for small genome sequencing was followed according to the manufacturer's recommendations. Criteria used for variant calling (quality threshold, heteroplasmy threshold, coverage threshold) were as described by King et al [6]. On-board software (i.e., Real-TimeAnalysis and MiSeq Reporter) converted raw data to Binary Alignment/Map (BAM) [20] and Variant Call Format (VCF) v4.1 files [21] using Genome Analysis Toolkit (GATK) [22].…”

Section: Sequencing and Data Generationmentioning

confidence: 99%

“…However, it has been reported that 75 % of the total variation within the mtGenome resides outside the control region, and therefore sequencing of the entire mtGenome increases the discrimination power and the value of generated data [6]. Sanger-type sequencing (STS) is still the main mtDNA typing technique in case work laboratories, and sequencing beyond the control region is attempted rarely as the well-established methodology is labor intensive, costly, and time consuming.…”

Section: Introductionmentioning

confidence: 99%

“…STS. With the utility of bench-top sequencers like Illumina MiSeq and Ion Torrent PGM (Personal Genome Machine), generation of whole mtGenome data is feasible for the application-orientated laboratory [6][7][8][9][10].…”

Section: Introductionmentioning

confidence: 99%

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Whole mitochondrial genome genetic diversity in an Estonian population sample

et al. 2015

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Mitochondrial DNA is a useful marker for population studies, human identification, and forensic analysis. Commonly used hypervariable regions I and II (HVI/HVII) were reported to contain as little as 25% of mitochondrial DNA variants and therefore the majority of power of discrimination of mitochondrial DNA resides in the coding region. Massively parallel sequencing technology enables entire mitochondrial genome sequencing. In this study, buccal swabs were collected from 114 unrelated Estonians and whole mitochondrial genome sequences were generated using the Illumina MiSeq system. The results are concordant with previous mtDNA control region reports of high haplogroup HV and U frequencies (47.4 and 23.7% in this study, respectively) in the Estonian population. One sample with the Northern Asian haplogroup D was detected. The genetic diversity of the Estonian population sample was estimated to be 99.67 and 95.85%, for mtGenome and HVI/HVII data, respectively. The random match probability for mtGenome data was 1.20 versus 4.99% for HVI/HVII. The nucleotide mean pairwise difference was 27 ± 11 for mtGenome and 7 ± 3 for HVI/HVII data. These data describe the genetic diversity of the Estonian population sample and emphasize the power of discrimination of the entire mitochondrial genome over the hypervariable regions.

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Section: Resultsmentioning

confidence: 69%

Section: Sample Preparation and Target Amplificationmentioning

confidence: 99%

Section: Sequencing and Data Generationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Whole mitochondrial genome genetic diversity in an Estonian population sample

et al. 2015

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Validation of Expressmarker mtDNA‐SNP60: A mitochondrial SNP kit for forensic application

et al. 2016

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Expressmarker mtDNA-SNP60 kit, a 3-dye fluorescence labeling forensic mitochondrial-SNP kit, was developed and modified based on previous work. In this study, it was validated through sensitivity, precision, accuracy, reproducibility, species specificity, mixture samples, PCR-based conditions, and inhibitor studies, according to the Scientific Working Group on DNA Analysis Methods guidelines. In the sensitivity study, full profile was obtained from template DNA with the quantity ≥ 4 pg. Consistent profiles were obtained at three different laboratories, which demonstrated the reproducibility of the kit. No peak was observed in the profiles of common animal species and microorganisms demonstrating high species specificity of the kit. In the mixture study, some loci (minor contributors) were missing at 19:1 and 9:1 ratios, whereas all loci could be profiled at other ratios (4:1, 2:1, and 1:1). PCR-based studies demonstrated that there recommended PCR conditions were optimized for the kit, and there was acceptable tolerance of error. The inhibitor study indicated that the kit has varying degrees of resistance to the presence of common inhibitors. Twelve random individuals were profiled by next-generation sequencing to confirm the allele-specific PCR assay, and a consistent result was obtained. Furthermore, 520 individuals from 260 families were profiled and the results showed that all loci of the confirmed mother and child were matched. In addition, forensic population genetics parameters, including random match probability, discrimination power, and haplotype diversity, were calculated (0.0474, 0.9526, and 0.9563, respectively). Our study demonstrates that the Expressmarker mtDNA-SNP60 kit can be a useful supplementary tool for forensic application.

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Nanopore sequencing: An enrichment‐free alternative to mitochondrial DNA sequencing

2018

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Mitochondrial DNA sequence data are often utilized in disease studies, conservation genetics and forensic identification. The current approaches for sequencing the full mtGenome typically require several rounds of PCR enrichment during Sanger or MPS protocols followed by fairly tedious assembly and analysis. Here we describe an efficient approach to sequencing directly from genomic DNA samples without prior enrichment or extensive library preparation steps. A comparison is made between libraries sequenced directly from native DNA and the same samples sequenced from libraries generated with nine overlapping mtDNA amplicons on the Oxford Nanopore MinION™ device. The native and amplicon library preparation methods and alternative base calling strategies were assessed to establish error rates and identify trends of discordance between the two library preparation approaches. For the complete mtGenome, 16 569 nucleotides, an overall error rate of approximately 1.00% was observed. As expected with mtDNA, the majority of error was detected in homopolymeric regions. The use of a modified basecaller that corrects for ambiguous signal in homopolymeric stretches reduced the error rate for both library preparation methods to approximately 0.30%. Our study indicates that direct mtDNA sequencing from native DNA on the MinION™ device provides comparable results to those obtained from common mtDNA sequencing methods and is a reliable alternative to approaches using PCR‐enriched libraries.

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High-quality and high-throughput massively parallel sequencing of the human mitochondrial genome using the Illumina MiSeq

Cited by 153 publications

References 41 publications

Whole mitochondrial genome genetic diversity in an Estonian population sample

Whole mitochondrial genome genetic diversity in an Estonian population sample

Validation of Expressmarker mtDNA‐SNP60: A mitochondrial SNP kit for forensic application

Nanopore sequencing: An enrichment‐free alternative to mitochondrial DNA sequencing

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