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Year:: 2024
Contributor:: Martinez Sandoval, Aleisha (Author); Wang, Peizhe (Author); Wang,Xuezhu (Author); Trembley, Michael (Author); Xiao, Feng (Author); Pu, William (Author)
Subject:: genomics; cardiomyocytes; Congenital Heart Disease; iPSC-CMs; Single Cell Sequencing; non coding de novo variants
DOI: https://doi.org/10.26300/s90h-v172

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Abstract:: Simultaneous High-throughput ATAC and RNA Expression with sequencing (SHARE-seq) is a powerful technique for the measurement of chromatin accessibility and gene expression at a single cell level. Notably, SHARE-seq is amenable to customization and can be further optimized to obtain information on a cell's genotype and gene expression simultaneously. Whole exome sequencing has provided extensive data sets of coding variants that code for congenital heart disease (CHD), yet far less is known about non-coding de novo variants (ncDNVs). The Pu lab aims to leverage this technology to study the function of ncDNVs in CHD using an iPS-CM differentiation (induced pluripotent stem cells - cardiomyocytes) model. In previous work, the Pu lab demonstrated that SHARE-seq can be applied to iPSC-CMs, providing insights into the functional effects of ncDNVs. This project’s goal is to validate the capability of SHARE-seq in facilitating precise single cell genotyping by assigning each cell’s genotype to its gene expression profile, as current methods produce results with low resolution and very poor accuracy in detecting ncDNVs. We customized the SHARE-seq protocol to assess CHD ncDNVs in their native genomic context, by coupling SHARE-seq with sequence capture technology. We simultaneously generated single cell barcoded DNA-seq and RNA-seq libraries. Relevant DNA regions were enriched and sequenced to determine the genotype of each ncDNV. The genotyping results were subsequently projected onto the UMAP generated from the RNA-seq data. In this study, to assess the capability of SHARE-seq based single cell genotyping, we mixed the wildtype fibroblast and iPSC cell line harboring a defined ncDNV with a sequential ratio to compare and validate their actual genetic makeup against their original mixing ratio.

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Martinez Sandoval, Aleisha, Wang, Peizhe, Wang,Xuezhu, et al., "Customized SHARE-seq for Enhanced Single Cell Genotyping" (2024). Summer Research Symposium, The Leadership Alliance Digital Library. Brown Digital Repository. Brown University Library. https://doi.org/10.26300/s90h-v172

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Summer Research Symposium

Each year, Brown University showcases the research of its undergraduates at the Summer Research Symposium. More than half of the student-researchers are UTRA recipients, while others receive funding from a variety of Brown-administered and national programs and fellowships and go …
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The Leadership Alliance Digital Library

A collection of research presentations and posters created by students participating in the Leadership Alliance's Summer Research - Early Identification Program.
When using this resource, please cite the digital object identifier (DOI) for the specific resource as well as the …

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Customized SHARE-seq for Enhanced Single Cell Genotyping