{
  "protocol": {
    "uri": "https://www.protocols.io/api/v3/protocols/9451",
    "source": "protocolsio/v3",
    "doi": "dx.doi.org/10.17504/protocols.io.mgjc3un",
    "published_on": 1516132805,
    "version": 0,
    "source_id": 9451,
    "name": "CUT&amp;RUN: Targeted in situ genome-wide profiling with high efficiency for low cell numbers",
    "description": {
      "body": "<div class = \"text-blocks\"><div class = \"text-block\">Cleavage Under Targets and Release Using Nuclease (CUT&RUN) is an epigenomic profiling strategy in which antibody-targeted controlled cleavage by micrococcal nuclease releases specific protein-DNA complexes into the supernatant for paired-end DNA sequencing. As only the targeted fragments enter into solution, and the vast majority of DNA is left behind, CUT&RUN has exceptionally low background levels. CUT&RUN outperforms the most widely used Chromatin Immunoprecipitation (ChIP) protocols in resolution, signal-to-noise, and depth of sequencing required. In contrast to ChIP, CUT&RUN is free of solubility and DNA accessibility artifacts and can be used to profile insoluble chromatin and to detect long-range 3D contacts without cross-linking. Here we present an improved CUT&RUN protocol that does not require isolation of nuclei and provides high-quality data starting with only 100 cells for a histone modification and 1000 cells for a transcription factor. From cells to purified DNA CUT&RUN requires less than a day at the lab bench.</div><div class = \"text-block\">In summary, CUT&RUN has several advantages over ChIP-seq: (1) The method is performed in situ in non-crosslinked cells and does not require chromatin fragmentation or solubilization; (2) The intrinsically low background allows low sequence depth and identification of low signal genomic features invisible to ChIP; (3) The simple procedure can be completed within a day and is suitable for robotic automation; (4) The method can be used with low cell numbers compared to existing methodologies; (5) A simple spike-in strategy can be used for accurate quantitation of protein-DNA interactions. As such, CUT&RUN represents an attractive replacement for ChIPseq, which is one of the most popular methods in biological research.</div></div>",
      "image": "https://s3.amazonaws.com/pr-journal/rzxfw26.png",
      "extra_content": []
    },
    "authors": "Peter J. Skene, Steven Henikoff",
    "steps": [
      {
        "source_id": 601564,
        "name": "Binding cells to beads",
        "attachments": [
          {
            "url": "https://pbs.twimg.com/media/Cwu3zrZWQAA7axs.jpg",
            "name": "First file"
          },
          {
            "url": "http://something.com/wp-content/uploads/2014/11/14506718045_5b3e71dacd_o.jpg",
            "name": "Second file"
          },
          {
            "url": "http://something.com/wp-content/uploads/2014/11/14506718045_5b3e71dacd_o.jpg",
            "name": "14506718045_5b3e71dacd_o.jpg"
          }
        ],
        "description": {
          "body": "<div class = \"text-blocks\"><div class = \"text-block\"><span style = \":justify;\">Harvest fresh culture(s) at room temperature and count cells. The same protocol can be used for 100 to 250,000 mammalian cells per sample and/or digestion time point.</span></div></div>",
          "components": [
            {
              "type": "amount",
              "value": 123,
              "unit": "µl",
              "name": "of MGH"
            },
            {
              "type": "duration",
              "value": 11,
              "name": "boil"
            },
            {
              "type": "link",
              "source": "www.google.com/protocols/123"
            },
            {
              "type": "software",
              "name": "RubyMine",
              "source": "www.rubymine.com",
              "details": {
                "repository_link": "www.github.com/rubymine/",
                "developer": "JozaHans",
                "os_name": "OSX and Linux."
              }
            },
            {
              "type": "dataset",
              "name": "WiFi passwords dictionary",
              "source": "www.passwords.com/wifi"
            },
            {
              "type": "command",
              "software_name": "Terminal",
              "command": "curl",
              "details": {
                "os_name": "OSX and Linux"
              }
            },
            {
              "type": "result",
              "body": "<div class = \"text-blocks\"><div class = \"text-block\">Centrifuge 3 min... This is expected result with HTML.</div></div>"
            },
            {
              "type": "warning",
              "body": "<div class = \"text-blocks\"><div class = \"text-block\">Centrifuge 3 min... This is WARNING with HTML.</div></div>",
              "details": {
                "link": "www.warnings.com/be_careful"
              }
            },
            {
              "type": "reagent",
              "name": "2 mg Gastrin I, human",
              "link": "https://www.biorbyt.com/gastrin-i-human-orb321073",
              "details": {
                "catalog_number": "",
                "linear_formula": "C130H204N38O31S",
                "mol_weight": 0.1
              }
            },
            {
              "type": "gotostep",
              "value": "In case of someting, go to step",
              "step_id": 601565
            },
            {
              "type": "temperature",
              "value": 55,
              "unit": "°C",
              "name": "boil"
            },
            {
              "type": "concentration",
              "value": 12,
              "unit": "g/L",
              "name": "Name of concentration"
            },
            {
              "type": "note",
              "author": "Frank Jones",
              "body": "<div class = \"text-blocks\"><div class = \"text-block\">Centrifuge 3 min... This is expected result with HTML.</div></div>"
            }
          ]
        },
        "position": 0
      },
      {
        "source_id": 601565,
        "name": "Binding cells to beads 2",
        "attachments": [],
        "description": {
          "body": "<div class = \"text-blocks\"><div class = \"text-block\">Centrifuge 3 min 600 x g at room temperature and withdraw liquid.</div></div>",
          "components": []
        },
        "position": 1
      }
    ]
  }
}
