Required reagents included in kit
Kit components are stable for 6 months upon date of receipt. Store as outlined below.
Store at room temperature (RT) upon receipt:
| Item | Catalog Number | Notes |
|---|---|---|
| 8-strip Tubes | 10-0009a | Enables use of multi-channel pipettors. |
| DNA Cleanup Columns *for kit Versions 1-3 | 10-0010 | Use with the DNA Collection Tubes. |
| DNA Collection Tubes *for kit Versions 1-3 | 10-0011 | Use with the DNA Cleanup Columns. |
| DNA Binding Buffer *for kit Versions 1-3 | 21-1008 | Before first use, add 6.9 mL isopropanol. WARNING: Contains toxic ingredients (see Safety Data Sheet). |
| DNA Wash Buffer *for kit Versions 1-3 | 21-1009 | Before first use, add 20 mL of ≥95% ethanol. |
| DNA Elution Buffer *for kit Versions 1-3 | 21-1010 | Use to elute final CUT&RUN DNA. |
| SPRIselect Reagent Manufactured by Beckman Coulter **for kit Version 4 | 21-1405 | DO NOT FREEZE. Reagent is slightly viscous. Thoroughly mix prior to use and pipette carefully to ensure correct volume is transferred. Used to purify CUT&RUN-enriched DNA from assay supernatant. |
| 0.1X TE Buffer **for kit Version 4 | 21-1025 | Used to elute CUT&RUN-enriched DNA. |
| 0.5 M EDTA | 21-1006 | 250X concentration. Use to prepare Antibody Buffer FRESH for each experiment. |
| 100 mM Calcium Chloride | 21-1007 | Activates chromatin-tethered pAG-MNase to cleave DNA. |
Store at 4°C upon receipt:
| Item | Catalog Number | Notes |
|---|---|---|
| ConA Beads | 21-1401 | DO NOT FREEZE. Concanavalin A (ConA) beads are used for immobilizing nuclei or cells. Because ConA can cause immune cell activation, it is recommended to use nuclei for immune cell studies (see this article). |
| E. coli spike-in DNA | 18-1401 | 100 ng lyophilized E. coli DNA. Before first use, quick spin and reconstitute in 200 μL DNase-free water (0.5 ng/μL). Add to reactions for sequencing normalization. NOTE: After reconstitution, store at -20°C. |
| Bead Activation Buffer | 21-1001 | Use to prepare ConA beads prior to sample immobilization. |
| Pre-Wash Buffer | 21-1002 | Use to prepare Wash, Cell Permeabilization, and Antibody Buffers FRESH for each experiment. |
| Stop Buffer | 21-1003 | 3X concentration. Use to terminate pAG-MNase cleavage activity. |
Store at -20°C upon receipt:
| Item | Catalog Number | Notes |
|---|---|---|
| 5% Digitonin | 21-1004k | Thaw at RT. Use to prepare Cell Permeabilization and Antibody Buffers FRESH for each experiment. Final Digitonin concentration should be optimized for each sample type, see this article. |
| 1 M Spermidine | 21-1005 | 2,000X concentration. Use to prepare Wash Buffer FRESH for each experiment. |
| SNAP-CUTANA™ K-MetStat Panel | 19-1002k | SMALL VOLUME: quick spin before use. Pipette to resuspend - DO NOT VORTEX. Panel of biotinylated nucleosomes coupled to streptavidin-coated magnetic beads. Pair with IgG and H3K4me3 control antibodies. Sufficient for 20 reactions. See this section for more information. |
| Rabbit IgG Negative Control Antibody | 13-0041k | SMALL VOLUME: quick spin before use. 0.5 mg/mL stock. Add 1μL to negative control reactions. Sufficient volume for 10 reactions. |
| H3K4me3 Positive Control Antibody | 13-0042k | SMALL VOLUME: quick spin before use. 0.5 mg/mL rabbit mixed monoclonal antibody. Add 1 μL to positive control reactions. Sufficient volume for 10 reactions. |
| pAG-MNase | 15-1016 | 20X concentration. Proteins A and G (pAG) bind antibodies of various isotypes and host species including total IgG for rabbit, mouse, goat, donkey, rat, guinea pig, horse, and cow. |
SPRIselect Reagent
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