Reducing mitochondrial reads in ATAC-seq using CRISPR/Cas9

Abstract ATAC-seq is a high-throughput sequencing technique that identifies open chromatin. Depending on the cell type, ATAC-seq samples may contain ~20–80% of mitochondrial sequencing reads. As the regions of open chromatin of interest are usually located in the nuclear genome, mitochondrial reads...

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Autores principales:	Lindsey Montefiori, Liana Hernandez, Zijie Zhang, Yoav Gilad, Carole Ober, Gregory Crawford, Marcelo Nobrega, Noboru Jo Sakabe
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/5fcc8d791745498ea430b40864f3d932
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spelling	oai:doaj.org-article:5fcc8d791745498ea430b40864f3d9322021-12-02T15:05:32ZReducing mitochondrial reads in ATAC-seq using CRISPR/Cas910.1038/s41598-017-02547-w2045-2322https://doaj.org/article/5fcc8d791745498ea430b40864f3d9322017-05-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-02547-whttps://doaj.org/toc/2045-2322Abstract ATAC-seq is a high-throughput sequencing technique that identifies open chromatin. Depending on the cell type, ATAC-seq samples may contain ~20–80% of mitochondrial sequencing reads. As the regions of open chromatin of interest are usually located in the nuclear genome, mitochondrial reads are typically discarded from the analysis. We tested two approaches to decrease wasted sequencing in ATAC-seq libraries generated from lymphoblastoid cell lines: targeted cleavage of mitochondrial DNA fragments using CRISPR technology and removal of detergent from the cell lysis buffer. We analyzed the effects of these treatments on the number of usable (unique, non-mitochondrial) reads and the number and quality of peaks called, including peaks identified in enhancers and transcription start sites. Both treatments resulted in considerable reduction of mitochondrial reads (1.7 and 3-fold, respectively). The removal of detergent, however, resulted in increased background and fewer peaks. The highest number of peaks and highest quality data was obtained by preparing samples with the original ATAC-seq protocol (using detergent) and treating them with CRISPR. This strategy reduced the amount of sequencing required to call a high number of peaks, which could lead to cost reduction when performing ATAC-seq on large numbers of samples and in cell types that contain a large amount of mitochondria.Lindsey MontefioriLiana HernandezZijie ZhangYoav GiladCarole OberGregory CrawfordMarcelo NobregaNoboru Jo SakabeNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-9 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Lindsey Montefiori Liana Hernandez Zijie Zhang Yoav Gilad Carole Ober Gregory Crawford Marcelo Nobrega Noboru Jo Sakabe Reducing mitochondrial reads in ATAC-seq using CRISPR/Cas9
description	Abstract ATAC-seq is a high-throughput sequencing technique that identifies open chromatin. Depending on the cell type, ATAC-seq samples may contain ~20–80% of mitochondrial sequencing reads. As the regions of open chromatin of interest are usually located in the nuclear genome, mitochondrial reads are typically discarded from the analysis. We tested two approaches to decrease wasted sequencing in ATAC-seq libraries generated from lymphoblastoid cell lines: targeted cleavage of mitochondrial DNA fragments using CRISPR technology and removal of detergent from the cell lysis buffer. We analyzed the effects of these treatments on the number of usable (unique, non-mitochondrial) reads and the number and quality of peaks called, including peaks identified in enhancers and transcription start sites. Both treatments resulted in considerable reduction of mitochondrial reads (1.7 and 3-fold, respectively). The removal of detergent, however, resulted in increased background and fewer peaks. The highest number of peaks and highest quality data was obtained by preparing samples with the original ATAC-seq protocol (using detergent) and treating them with CRISPR. This strategy reduced the amount of sequencing required to call a high number of peaks, which could lead to cost reduction when performing ATAC-seq on large numbers of samples and in cell types that contain a large amount of mitochondria.
format	article
author	Lindsey Montefiori Liana Hernandez Zijie Zhang Yoav Gilad Carole Ober Gregory Crawford Marcelo Nobrega Noboru Jo Sakabe
author_facet	Lindsey Montefiori Liana Hernandez Zijie Zhang Yoav Gilad Carole Ober Gregory Crawford Marcelo Nobrega Noboru Jo Sakabe
author_sort	Lindsey Montefiori
title	Reducing mitochondrial reads in ATAC-seq using CRISPR/Cas9
title_short	Reducing mitochondrial reads in ATAC-seq using CRISPR/Cas9
title_full	Reducing mitochondrial reads in ATAC-seq using CRISPR/Cas9
title_fullStr	Reducing mitochondrial reads in ATAC-seq using CRISPR/Cas9
title_full_unstemmed	Reducing mitochondrial reads in ATAC-seq using CRISPR/Cas9
title_sort	reducing mitochondrial reads in atac-seq using crispr/cas9
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/5fcc8d791745498ea430b40864f3d932
work_keys_str_mv	AT lindseymontefiori reducingmitochondrialreadsinatacsequsingcrisprcas9 AT lianahernandez reducingmitochondrialreadsinatacsequsingcrisprcas9 AT zijiezhang reducingmitochondrialreadsinatacsequsingcrisprcas9 AT yoavgilad reducingmitochondrialreadsinatacsequsingcrisprcas9 AT caroleober reducingmitochondrialreadsinatacsequsingcrisprcas9 AT gregorycrawford reducingmitochondrialreadsinatacsequsingcrisprcas9 AT marcelonobrega reducingmitochondrialreadsinatacsequsingcrisprcas9 AT noborujosakabe reducingmitochondrialreadsinatacsequsingcrisprcas9
_version_	1718388766628904960

Reducing mitochondrial reads in ATAC-seq using CRISPR/Cas9

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