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A method for DNA methylation analysis in single cells using Next Generation Sequencing
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
2018 (English)Independent thesis Advanced level (degree of Master (Two Years)), 30 credits / 45 HE creditsStudent thesis
Abstract [en]

A cell’s fate is predominantly decided by the epigenetic regulations especially DNA methylation. The methylome of a cell is dynamic and varies between cell types. Analysis of cells in bulk masks the presence of rare subtypes of cells that may be important in executing functions of a tissue or tumor progression. Thus, it is important to analyse methylomes of single cells from a tissue or tumor to understand key aspects of cellular functions. There are very few methods for whole genome DNA methylation analysis in single cells. The current available methods lack the sensitivity of capturing the entire methylome and thus have low CpG detection rates and low mappability rates. In this project, we aim to adapt and modify Splinted Ligation Adapter Tagging (SPLAT) method for methylation analysis in single cells. We have thus far been able to successfully construct genomic libraries from low cell concentration as10 cells by SPLAT and sequenced them to observe satisfactory base call quality. We achieved a reasonable alignment rate of 40-46%. The average methylation rate was 56% and bisulfite conversion rate was a sub-optimal 93%. The protocol further needs to be shaped to apply it on single cells. DNA methylome decoding from single cells not only has immense scope in diagnosis and prognosis but also in stratification of cancer sub-groups and staging.

 

Place, publisher, year, edition, pages
2018. , p. 26
Keywords [en]
DNA methylation, single cells, CpG detection, WGBS, SPLAT, cell subtypes
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:uu:diva-359424OAI: oai:DiVA.org:uu-359424DiVA, id: diva2:1265541
Educational program
Master Programme in Medical Research
Presentation
2018-05-30, A11:220b, Husargatan 3, 751 23 Uppsala, Sweden, Uppsala, 11:51 (English)
Supervisors
Examiners
Available from: 2018-11-26 Created: 2018-11-24 Last updated: 2018-11-26Bibliographically approved

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The full text will be freely available from 2025-01-01 12:01
Available from 2025-01-01 12:01

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