CUT&amp;Tag for efficient epigenomic profiling of small samples and single cells

Kaya-Okur, Hatice S; Wu, Steven J.; Codomo, Christine A.; Pledger, Erica S.; Bryson, Terri D.; Henikoff, Steven; Ahmad, Kami; Henikoff, Steven

doi:10.1038/s41467-019-09982-5

Cited by 1,450 publications

(1,197 citation statements)

References 32 publications

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“…In addition to profiling in single cells, factor binding was profiled in individual early blastocysts (consisting of between 30-50 cells each), an application not previously possible using ChIP-based techniques. More recently, Cleavage Under Targets and Tagmentation, or CUT&Tag, was developed as a modification on CUT&RUN that uses a recombinant Protein A-Tn5 transposase fusion instead of a recombinant pA-MNase fusion protein (Kaya-Okur et al 2019). CUT&Tag has been used to profile histone modifications in single cells, although it has not yet been used to profile transcription factor binding in single cells (Kaya-Okur et al 2019).…”

Section: Cutandrunmentioning

confidence: 99%

“…More recently, Cleavage Under Targets and Tagmentation, or CUT&Tag, was developed as a modification on CUT&RUN that uses a recombinant Protein A-Tn5 transposase fusion instead of a recombinant pA-MNase fusion protein (Kaya-Okur et al 2019). CUT&Tag has been used to profile histone modifications in single cells, although it has not yet been used to profile transcription factor binding in single cells (Kaya-Okur et al 2019). In addition to CUT&Tag, a similar single-cell modification of ChIC, scChIC-seq, which involves tethering of MNase to a specific antibody and cleavage of target sites using the antibody to direct the MNase, then selectively amplifying cleaved fragments by PCR was developed (Ku et al 2019).…”

Section: Cutandrunmentioning

confidence: 99%

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Genomic methods in profiling DNA accessibility and factor localization

Klein

Hainer

2019

Chromosome Res

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Recent advancements in next-generation sequencing technologies and accompanying reductions in cost have led to an explosion of techniques to examine DNA accessibility and protein localization on chromatin genome-wide. Generally, accessible regions of chromatin are permissive for factor binding and are therefore hotspots for regulation of gene expression; conversely, genomic regions that are highly occupied by histone proteins are not permissive for factor binding and are less likely to be active regulatory regions. Identifying regions of differential accessibility can be useful to uncover putative gene regulatory regions, such as enhancers, promoters, and insulators. In addition, DNA-binding proteins, such as transcription factors that preferentially bind certain DNA sequences and histone proteins that form the core of the nucleosome, play essential roles in all DNA-templated processes. Determining the genomic localization of chromatin-bound proteins is therefore e s s e n t i a l i n d e t e r m i n i n g f u n c t i o n a l r o l e s , sequence motifs important for factor binding, and regulatory networks controlling gene expression. In this review, we discuss techniques for determining DNA accessibility and nucleosome positioning (DNase-seq, FAIRE-seq, MNase-seq, and ATACseq) and techniques for detecting and functionally characterizing chromatin-bound proteins (ChIP-seq, DamID, and CUT&RUN). These methods have been optimized to varying degrees of resolution, specificity, and ease of use. Here, we outline some advantages and disadvantages of these techniques, their general protocols, and a brief discussion of their development. Together, these complimentary approaches have provided an unparalleled view of chromatin architecture and functional gene regulation. Keywords Chromatin. DNase. MNase. ATAC. ChIP. CUT&RUN. nucleosome occupancy. transcription factors. genomics Abbreviations DHS DNase I hypersensitive site DNase-seq DNase I coupled with deep sequencing XL-DNaseseq Crosslinking DNase I coupled with deep sequencing scDNaseseq Single-cell DNase I coupled with deep sequencing FAIRE-seq Formaldehyde-assisted isolation of regulatory elements MNase-seq Micrococcal nuclease digestion coupled with deep sequencing MPE-seq Methidiumpropyl-EDTA cleavage coupled with deep sequencing ATAC-seq An assay for transposase accessibility

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Section: Cutandrunmentioning

confidence: 99%

Section: Cutandrunmentioning

confidence: 99%

Genomic methods in profiling DNA accessibility and factor localization

Klein

Hainer

2019

Chromosome Res

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“…Thus, when applied to tissues composed of diverse cell types, these methods can suffer from a low signal to noise ratio. This issue can potentially be overcome by single‐cell approaches, which have been developed for many of the genome‐wide methods described above, including ChIP‐seq (Rotem et al, ), CUT&RUN (Hainer, Bošković, McCannell, Rando, & Fazzio, ), CUT&TAG (Kaya‐okur et al, ), DNase‐seq (W. Jin et al, ), MNase‐seq (Lai et al, ), ATAC‐seq (Buenrostro et al, ; DeWitt et al, ), and Hi‐C (Nagano et al, ). Although highly scalable, at present the single‐cell versions of these methods yield sparse, low‐coverage data that are difficult to analyze using conventional approaches (Ji, Zhou, & Ji, ; Schep, Wu, Buenrostro, & Greenleaf, ).…”

Section: Genome‐wide Methodsologies To Identify Putative Enhancersmentioning

confidence: 99%

Functional genomic approaches to elucidate the role of enhancers during development

Ryan

Farley

2019

WIREs Mechanisms of Disease

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Successful development depends on the precise tissue-specific regulation of genes by enhancers, genetic elements that act as switches to control when and where genes are expressed. Because enhancers are critical for development, and the majority of disease-associated mutations reside within enhancers, it is essential to understand which sequences within enhancers are important for function. Advances in sequencing technology have enabled the rapid generation of genomic data that predict putative active enhancers, but functionally validating these sequences at scale remains a fundamental challenge. Herein, we discuss the power of genome-wide strategies used to identify candidate enhancers, and also highlight limitations and misconceptions that have arisen from these data. We discuss the use of massively parallel reporter assays to test enhancers for function at scale. We also review recent advances in our ability to study gene regulation during development, including CRISPR-based tools to manipulate genomes and single-cell transcriptomics to finely map gene expression. Finally, we look ahead to a synthesis of complementary genomic approaches that will advance our understanding of enhancer function during development.

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“…Cleavage under targets and tagmentation (CUT&Tag) is another technology that can detect DNA-protein interaction in low cellular input samples or even single cells [32]. Instead of Protein A/G-fused MNase, CUT&-Tag applies Protein A/G-fused Tn5 transposase (pA/G-Tn5) to cut DNA, demanding high-quality core enzyme.…”

Section: Lowering Cellular Inputmentioning

confidence: 99%

Profiling chromatin regulatory landscape: insights into the development of ChIP-seq and ATAC-seq

Zhang

2020

Mol Biomed

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Chromatin regulatory landscape plays a critical role in many disease processes and embryo development. Epigenome sequencing technologies such as chromatin immunoprecipitation sequencing (ChIP-seq) and assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) have enabled us to dissect the pan-genomic regulatory landscape of cells and tissues in both time and space dimensions by detecting specific chromatin state and its corresponding transcription factors. Pioneered by the advancement of chromatin immunoprecipitation-chip (ChIP-chip) technology, abundant epigenome profiling technologies have become available such as ChIP-seq, DNase I hypersensitive site sequencing (DNase-seq), ATAC-seq and so on. The advent of single-cell sequencing has revolutionized the next-generation sequencing, applications in single-cell epigenetics are enriched rapidly. Epigenome sequencing technologies have evolved from low-throughput to high-throughput and from bulk sample to the single-cell scope, which unprecedentedly benefits scientists to interpret life from different angles. In this review, after briefly introducing the background knowledge of epigenome biology, we discuss the development of epigenome sequencing technologies, especially ChIP-seq & ATAC-seq and their current applications in scientific research. Finally, we provide insights into future applications and challenges.

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CUT&Tag for efficient epigenomic profiling of small samples and single cells

Cited by 1,450 publications

References 32 publications

Genomic methods in profiling DNA accessibility and factor localization

Genomic methods in profiling DNA accessibility and factor localization

Functional genomic approaches to elucidate the role of enhancers during development

Profiling chromatin regulatory landscape: insights into the development of ChIP-seq and ATAC-seq

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