Chromatin Shearing

Covaris workflows enable standardized chromatin shearing from versatile samples such as primary cells, cultured cells, tissues, and FFPE samples. Covaris Adaptive Focused Acoustics® (AFA®) technology guarantees efficient and random chromatin fragmentation and supports nuclei extraction from hard to lyse specimens.

Covaris truChIP®: A Different Approach to Chromatin Shearing

Epigenetic characterization provides a powerful tool to better understand cell-fate changes during development and disease. However, standardizing the sample preparation steps in chromatin assays is crucial to retrieve meaningful genome-wide epigenomic data that can be compared against different experimental set-ups and different laboratories.
Covaris technology has enabled the standardization of chromatin preparation and shearing from virtually any sample input with consistent results. AFA technology in combination with optimized reagents provide researchers an easy, fast, reproducible and scalable workflow to retrieve high-quality sheared chromatin to analyze the epigenetic make-up of their sample on genome-wide scale.
Chromatin shearing with truChIP and AFA has many benefits including:
  • Lower cell number requirement​
    • Reduced cell culturing​, enable analysis of scarce clinical samples
  • Controlled shearing size range​
    • Reproducible ChIP-seq results
  • Isothermal processing​
    • Maintaining epitope integrity and no reversal of crosslinks during shearing​
  • Low detergent requirement (reduced SDS 0.1%)​
    • Suitable for all IP protocols​ and less chromatin dilutions
  • Gentle shearing​
    • Lower fixation time to avoid epitope masking
  • Universal protocol guaranteed to work with all mammalian cells​


Covaris truChIP Chromatin Shearing Benefits

Watch the video case study, Standardization of the ChIP Workflow Using Covaris Adaptive Focused Acoustics (AFA) to Study the Transcriptional Regulation of SOD1 in ALS Pathogenesis, presented by Byung Woo Kim, PhD Candidate, Johns Hopkins University and Hamid Khoja, PhD, Covaris
Download the Chromatin Preparation and Shearing Brochure, a presentation of Chromatin Applications enabled by AFA-energetics®

Downstream Epigenomic Applications: Translational Discoveries Powered by AFA-energetics®

Covaris instruments powered by Adaptive Focused Acoustics® (AFA®) are the well established and trusted high-throughput chromatin platforms available that allow chromatin shearing in a 96 well format. Sample types supported include cultured cell lines, primary cells, fresh/ frozen tissues, and FFPE Fixed tissues.
AFA is widely used by scientists to decipher the regulatory potential of chromatin landscape at several layers including histone modifications, DNA methylation patterns as well as interactions with biomolecules like transcription factors, epigenetic regulators and ncRNAs. The Covaris technology has further been proven beneficial to restore multi-subunit complexes from chromatin and to study 3D chromatin organization.
AFA is the method of choice in NuGEN’s Ovation® platform and Agilent’s SureSelectXT Methyl-Seq protocols for profiling 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC). It is also cited as the preferred sample prep method for meDIP-Seq, ChIP-Seq, HI-C, HI-ChIP, ChIRP-Seq, and other epigenomics applications.



Click here to download our Comprehensive Epigenomic Sample Prep Flyer

“In the past we struggled to get reproducible chromatin shearing to a size range suitable for ChIPseq experiments. However, we were relieved that using the Covaris E220 Focused-ultrasonicator enabled us to optimize a chromatin sample preparation protocol in less than 2 weeks. The fast optimization and high degree of reproducibility in shearing, without day to day or sample to sample variation, has made our scientific life much easier.”
Stefanie Dichtl, PhD

Max-Planck-Institut für Biochemie

Resources

References

  1. Caliskan M, Manduchi E, Rao S et.al. Genetic and Epigenetic Fine Mapping of Complex Trait Associated Loci in the Human Liver. AJHG. Vol 105, Issue1, p89-107, July 03, 2019. DOI: 10.1016/j.ajhg.2019.05.010
  2. Michaelson JJ, Shin MK, Koh JY, et al. Neuronal PAS Domain Proteins 1 and 3 Are Master Regulators of Neuropsychiatric Risk Genes. Biol Psychiatry. 2017;82(3):213-223. DOI: 10.1016/j.biopsych.2017.03.021
  3. Kuznetsov VI, Haws SA, Fox CA, Denu JM. General method for rapid purification of native chromatin fragments. J Biol Chem. 2018. DOI: 10.1074/jbc.RA118.002984
  4. Tharp KM, Kang MS, Timblin GA, et al. Actomyosin-Mediated Tension Orchestrates Uncoupled Respiration in Adipose Tissues. Cell Metab. 2018;27(3):602-615.e4. DOI: 10.1038/s41590-018-0056-8
  5. Manni M, Gupta S, Ricker E, et al. Regulation of age-associated B cells by IRF5 in systemic autoimmunity. Nat Immunol. 2018;19(4):407-419. DOI:10.1038/s41590-018-0056-8
  6. Yao H, Hill SF, Skidmore JM, et al. CHD7 represses the retinoic acid synthesis enzyme ALDH1A3 during inner ear development. JCI Insight. 2018;3(4). DOI: 10.1172/jci.insight.97440
  7. Donnard E, Vangala P, Afik S, et al. Comparative Analysis of Immune Cells Reveals a Conserved Regulatory Lexicon. Cell Syst. 2018;6(3):381-394.e7. DOI: 10.1016/j.cels.2018.01.002
  8. Liang C, Wang S, Qin C, et al. TRIM36, a novel androgen-responsive gene, enhances anti-androgen efficacy against prostate cancer by inhibiting MAPK/ERK signaling pathways. Cell Death Dis. 2018;9(2):155. DOI: 10.1038/s41419-017-0197-y
  9. Park SM, Choi EY, Bae DH, Sohn HA, Kim SY, Kim YJ. The LncRNA EPEL Promotes Lung Cancer Cell Proliferation Through E2F Target Activation. Cell Physiol Biochem. 2018;45(3):1270-1283. DOI: 10.1159/000487460
  10. Josipovic I, Pflüger B, Fork C, et al. Long noncoding RNA LISPR1 is required for S1P signaling and endothelial cell function. J Mol Cell Cardiol. 2018;116:57-68. DOI: 10.1016/j.yjmcc.2018.01.015
  11. Kim M, Astapova II, Flier SN, et al. Intestinal, but not hepatic, ChREBP is required for fructose tolerance. JCI Insight. 2017;2(24). DOI: 10.1172/jci.insight.96703
  12. Ramaswamy K, Forbes L, Minuesa G, et al. Peptidomimetic blockade of MYB in acute myeloid leukemia. Nat Commun. 2018;9(1):110. DOI: 10.1038/s41467-017-02618-6
  13. Humblin E, Thibaudin M, Chalmin F, et al. IRF8-dependent molecular complexes control the Th9 transcriptional program. Nat Commun. 2017;8(1):2085. DOI: 10.1038/s41467-017-01070-w
  14. Bu Y, Yoshida A, Chitnis N, et al. A PERK-miR-211 axis suppresses circadian regulators and protein synthesis to promote cancer cell survival. Nat Cell Biol. 2018;20(1):104-115. DOI: 10.1038/s41556-017-0006-y
  15. Li L, Fan CM. A CREB-MPP7-AMOT Regulatory Axis Controls Muscle Stem Cell Expansion and Self-Renewal Competence. Cell Rep. 2017;21(5):1253-1266. DOI: 10.1016/j.celrep.2017.10.031
  16. Forsyth CB, Shaikh M, Bishehsari F, et al. Alcohol Feeding in Mice Promotes Colonic Hyperpermeability and Changes in Colonic Organoid Stem Cell Fate. Alcohol Clin Exp Res. 2017;41(12):2100-2113. DOI: 10.1111/acer.13519
  17. Isobe K, Jung HJ, Yang CR, et al. Systems-level identification of PKA-dependent signaling in epithelial cells. Proc Natl Acad Sci USA. 2017;114(42):E8875-E8884. DOI: 10.1073/pnas.1709123114
  18. Zheng X, Han H, Liu GP, et al. LncRNA wires up Hippo and Hedgehog signaling to reprogramme glucose metabolism. EMBO J. 2017;36(22):3325-3335. DOI: 10.15252/embj.201797609
  19. Osabe M, Tajika T, Tohkin M. Allopurinol suppresses expression of the regulatory T-cell migration factors TARC/CCL17 and MDC/CCL22 in HaCaT keratinocytes via restriction of nuclear factor-κB activation. J Appl Toxicol. 2018;38(2):274-283. DOI: 10.1002/jat.3522
  20. Hatanaka Y, Tsusaka T, Shimizu N, et al. Histone H3 Methylated at Arginine 17 Is Essential for Reprogramming the Paternal Genome in Zygotes. Cell Rep. 2017;20(12):2756-2765. DOI: 10.1016/j.celrep.2017.08.088
  21. Nathan S, Ma Y, Tomita YA, De oliveira E, Brown ML, Rosen EM. BRCA1-mimetic compound NSC35446.HCl inhibits IKKB expression by reducing estrogen receptor-α occupancy in the IKKB promoter and inhibits NF-κB activity in antiestrogen-resistant human breast cancer cells. Breast Cancer Res Treat. 2017;166(3):681-693. DOI: 10.1007/s10549-017-4442-y
  22. Nathan S, Ma Y, Tomita YA, De oliveira E, Brown ML, Rosen EM. BRCA1-mimetic compound NSC35446.HCl inhibits IKKB expression by reducing estrogen receptor-α occupancy in the IKKB promoter and inhibits NF-κB activity in antiestrogen-resistant human breast cancer cells. Breast Cancer Res Treat. 2017;166(3):681-693. DOI: 10.1007/s10549-017-4442-y
  23. Mehta S, Cronkite DA, Basavappa M, et al. Maintenance of macrophage transcriptional programs and intestinal homeostasis by epigenetic reader SP140. Sci Immunol. 2017;2(9). DOI: 10.1126/sciimmunol.aag3160
  24. Bandyopadhaya A, Tsurumi A, Rahme LG. NF-κBp50 and HDAC1 Interaction Is Implicated in the Host Tolerance to Infection Mediated by the Bacterial Quorum Sensing Signal 2-Aminoacetophenone. Front Microbiol. 2017;8:1211. DOI: 10.3389/fmicb.2017.01211
  25. Gullicksrud JA, Li F, Xing S, et al. Differential Requirements for Tcf1 Long Isoforms in CD8 and CD4 T Cell Responses to Acute Viral Infection. J Immunol. 2017;199(3):911-919. DOI: 10.4049/jimmunol.1700595
  26. Gulchina Y, Xu SJ, Snyder MA, Elefant F, Gao WJ. Epigenetic mechanisms underlying NMDA receptor hypofunction in the prefrontal cortex of juvenile animals in the MAM model for schizophrenia. J Neurochem. 2017;143(3):320-333. DOI: 10.1111/jnc.14101
  27. Shan Q, Zeng Z, Xing S, et al. The transcription factor Runx3 guards cytotoxic CD8 effector T cells against deviation towards follicular helper T cell lineage. Nat Immunol. 2017;18(8):931-939. DOI: 10.1038/ni.3773
  28. Michaelson JJ, Shin MK, Koh JY, et al. Neuronal PAS Domain Proteins 1 and 3 Are Master Regulators of Neuropsychiatric Risk Genes. Biol Psychiatry. 2017;82(3):213-223. DOI: 10.1016/j.biopsych.2017.03.021
  29. Aldiri I, Xu B, Wang L, et al. The Dynamic Epigenetic Landscape of the Retina During Development, Reprogramming, and Tumorigenesis. Neuron. 2017;94(3):550-568.e10. DOI: 10.1016/j.neuron.2017.04.022
  30. Hwang JR, Chou CL, Medvar B, Knepper MA, Jung HJ. Identification of β-catenin-interacting proteins in nuclear fractions of native rat collecting duct cells. Am J Physiol Renal Physiol. 2017;313(1):F30-F46. DOI: 10.1152/ajprenal.00054.2017
  31. Hsieh LT, Nastase MV, Roedig H, et al. Biglycan- and Sphingosine Kinase-1 Signaling Crosstalk Regulates the Synthesis of Macrophage Chemoattractants. Int J Mol Sci. 2017;18(3). DOI: 10.3390/ijms18030595
  32. Hirose M, Hasegawa A, Mochida K, et al. CRISPR/Cas9-mediated genome editing in wild-derived mice: generation of tamed wild-derived strains by mutation of the a (nonagouti) gene. Sci Rep. 2017;7:42476. DOI: 10.1038/srep42476
  33. Li C, Wang S, Xing Z, et al. A ROR1-HER3-lncRNA signalling axis modulates the Hippo-YAP pathway to regulate bone metastasis. Nat Cell Biol. 2017;19(2):106-119. DOI: 10.1038/ncb3464
  34. Zhang J, Vlasevska S, Wells VA, et al. The CREBBP Acetyltransferase Is a Haploinsufficient Tumor Suppressor in B-cell Lymphoma. Cancer Discov. 2017;7(3):322-337. DOI: 10.1158/2159-8290.CD-16-1417
  35. Grajkowska LT, Ceribelli M, Lau CM, et al. Isoform-Specific Expression and Feedback Regulation of E Protein TCF4 Control Dendritic Cell Lineage Specification. Immunity. 2017;46(1):65-77. DOI: 10.1016/j.immuni.2016.11.006
  36. Fork C, Vasconez AE, Janetzko P, et al. Epigenetic control of microsomal prostaglandin E synthase-1 by HDAC-mediated recruitment of p300. J Lipid Res. 2017;58(2):386-392. DOI: 10.1194/jlr.M072280
  37. Bersaas A, Arnoldussen YJ, Sjøberg M, Haugen A, Mollerup S. Epithelial-mesenchymal transition and FOXA genes during tobacco smoke carcinogen induced transformation of human bronchial epithelial cells. Toxicol In Vitro. 2016;35:55-65. DOI: 10.1016/j.tiv.2016.04.012
  38. Song I, Kim K, Kim JH, et al. GATA4 negatively regulates osteoblast differentiation by downregulation of Runx2. BMB Rep. 2014;47(8):463-8. DOI: 10.1002/jbm4.10027
  39. Foxn1 regulates key target genes essential for T cell development in postnatal thymic epithelial cells. Zuklys S, et al.Nature Immunology, 2016. DOI: 10.1038/ni.3537
  40. Tcf1 and Lef1 transcription factors establish CD8+ T cell identity through intrinsic HDAC activity. Xing S, et al. Nature Immunology, 2016. DOI: 10.1038/ni.3456
  41. miR-216b regulation of c-Jun mediates GADD153/CHOP-dependent apoptosis. Xu Z, et al. Nature Communications, 2016. DOI: 10.1038/ncomms11422
  42. cChIP-seq: a robust small-scale method for investigation of histone modifications. Valensisi C, et al. BMC Genomics, 2015. DOI: 10.1186/s12864-015-2285-7
  43. Nrf1 and Nrf2 Transcription Factors Regulate Androgen Receptor Transactivation in Prostate Cancer Cells. Schultz MA, et al. PLoS ONE, 2014. DOI: 10.1371/journal.pone.0087204
  44. Bivalent chromatin marks developmental regulatory genes in the mouse embryonic germline in vivo. Sachs et al. Cell Reports, 2013. DOI: 10.1016/j.celrep.2013.04.032