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Table 1 Comparison between our protocol and some other available N-ChIP protocols

From: A native chromatin immunoprecipitation (ChIP) protocol for studying histone modifications in strawberry fruits

Procedures Others [13, 19,20,21, 43,44,45,46,47] Our optimized N-ChIP protocol
Starting material Non-fruit tissues or suspension cells [13, 19,20,21, 43,44,45,46,47] Fleshy fruit tissues
Nuclei isolation No particular steps for homogenization [20, 21, 47]. Homogenize in an all-glass homogenizer using a tight pestle [13, 45, 46]. Homogenize in a roller stirrer [19] or through syringe [44]. Homogenize in an undescribed way [43] The powder was washed in PBS-PSV twice firstly. Homogenize in a glass homogenizer followed by lysis on ice with gentle agitation for 10 min
Chromatin extraction Extraction buffer contains 0.4–0.8% NP40 [45, 46], 0.1% Triton X-100 [43, 47], 0.5% Triton X-100 [21], 1% Triton X-100 [44], 0.25% Tween 40 [19], 1% Tween 40 [13] as the detergent, or no detergent [20] Extraction buffer contains 0.5–1% Triton X-100
Wash at least four times
Fragmentation Digestion at 37 °C for 10–12 min [19,20,21, 43, 46] or 4–7 min [19, 44, 45, 47] Digestion for 10 min at 37 °C (30U/400 μl). Recommend optimizing the enzyme dosage and digestion time for each preparation
Immunoprecipitation
Antibody
Protease inhibitors
Reducing reagents
Protein A–Sepharose [13, 19, 20, 45], Dynabeads Protein G [46], Dynabeads Protein A [21, 47] or Dynabeads Protein A/G beads [43, 44]
ChIP-grade or only tested by western blot [13, 19,20,21, 43,44,45,46,47]
Buffers contain protease inhibitor cocktail [20, 43,44,45,46,47], 0.1–0.2 mM PMSF [13, 19, 20, 43,44,45]
Buffers contain 0.5 μM–0.5 mM DTT [45, 46], 10 mM β-mercaptoethanol [21, 43, 47]
Dynabeads Protein A beads
Passed ChIP-grade antibodies validated by the ENCODE project
Buffer contains 1 × protease inhibitor cocktail tablets, 1 mM PMSF
Buffers contain 1 mM DTT and 11 mM β-mercaptoethanol