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A rapid and robust method for Mitochondrial DNA profiling

ABSTRACT: A rapid and robust method for Mitochondrial DNA profiling

PROVIDER: PRJNA1108319 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR28919629_1.fastq.gz	Fastqsanger.gz
	SRR28919629_2.fastq.gz	Fastqsanger.gz
	SRR28919630_1.fastq.gz	Fastqsanger.gz
	SRR28919630_2.fastq.gz	Fastqsanger.gz
	SRR28919631_1.fastq.gz	Fastqsanger.gz

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A rapid and robust method for Mitochondrial DNA profiling

Project description:Human mitochondrial DNA (mtDNA) variations, particularly the presence of both mutated and wild type mtDNA known as mtDNA heteroplasmies, have garnered increasing attention due to their clinical relevance to numerous diseases. Nonetheless, the absence of a feasible and economical approach has hampered large-scale population studies on mtDNA variations. Here, we present a novel human mtDNA sequencing method called 123-seq, which is based on the 123-multiplex PCR and completed in consecutive three-step biochemical reactions, with experimental validation of its reliability. The entire library construction process takes only about 90 minutes, enabling efficient capture, amplification, and library construction of mtDNA at the bulk genomic DNA level or single-cell level in a streamlined workflow. We then applied our method to assess single-cell mtDNA mutations in a human sample, profiling over 286 T lymphocytes from a 79-year-old female. We discovered that over 90% of cells carried at least one mtDNA mutation with variant allele frequencies (VAFs) exceeding 20%. Furthermore, 83.78% of these mutation sites were enriched in the protein-coding regions of mtDNA. Our findings suggest that mtDNA mutations with functional significance may be widespread in advanced age, indicating a potential link to T cell aging. This emphasizes the need for further investigation into the dynamics of age-related mtDNA mutations at the single-cell level.

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Super cChIP-seq, a robust and universal epigenetic profiling method using 10 cells

Project description:ChIP-seq is the standard technique to characterize the genome-wide epigenomic modifications, which requires millions of cells as starting material. Although some methods have recently reported to be suitable for epigenomic profiling with a small number of cells, there are still no methods capable of profiling both histone modifications and DNA methylation with extremely low input. Here, we developed a new method by supplementing carrier materials of chemically modified epigenetic markers and dUPT-containing DNA fragments during ChIP procedures. Using this strategy, we generated high-quality epigenomic profiles of both histone modifications (super cChIP-seq) and DNA methylation (super cMeDIP-seq) with as few as 10 cells. Thus, we provide a robust and straightforward universal approach for epigenomic profiling starting with extremely low input.

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