Reducing mitochondrial reads in ATAC-seq using CRISPR/Cas9
| العنوان: | Reducing mitochondrial reads in ATAC-seq using CRISPR/Cas9 |
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| المؤلفون: | Gregory E. Crawford, Yoav Gilad, Zijie Zhang, Lindsey E. Montefiori, Noboru J. Sakabe, Carole Ober, Marcelo A. Nobrega, Liana Gonzales |
| المصدر: | Scientific Reports Scientific Reports, Vol 7, Iss 1, Pp 1-9 (2017) |
| سنة النشر: | 2017 |
| مصطلحات موضوعية: | 0301 basic medicine, Mitochondrial DNA, Nuclear gene, Science, 0206 medical engineering, Genomics, ATAC-seq, 02 engineering and technology, Computational biology, Biology, Mitochondrion, DNA, Mitochondrial, DNA sequencing, Article, Cell Line, 03 medical and health sciences, chemistry.chemical_compound, CRISPR, Humans, Guide RNA, Lymphocytes, Enhancer, 030304 developmental biology, Genetics, Cell Nucleus, 0303 health sciences, Multidisciplinary, Cas9, High-Throughput Nucleotide Sequencing, Chromatin, Mitochondria, 030104 developmental biology, chemistry, Medicine, CRISPR-Cas Systems, Peak calling, 020602 bioinformatics, DNA |
| الوصف: | 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. |
| تدمد: | 2045-2322 |
| الوصول الحر: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::a04f72e38fc1e5bfd28e394be220b109Test https://pubmed.ncbi.nlm.nih.gov/28550296Test |
| حقوق: | OPEN |
| رقم الانضمام: | edsair.doi.dedup.....a04f72e38fc1e5bfd28e394be220b109 |
| قاعدة البيانات: | OpenAIRE |
| تدمد: | 20452322 |
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