Chromatin shearing for ChIP-Seq using Covaris S2

menkelab · 03-24-2010, 11:10 AM

Hi,

I'm testing shearing conditions using a covaris S2 to shear chromatin to 150~300 bp for ChIP-Seq. My sample consists of mouse embryonic limbs in 500~600 ul buffer (20 mM Tris, pH 8.0; 0.5 mM EGTA, 1mM EDTA) in a TC12 tube. My settings were 20% DC, 5 In, 200 C/B, 30 sec cycle time, varying numbers of cycle (10, 20, 30, 40, 50 cycles). I did not include treatment 2 following treatment 1 for a pause between cycles. I had a size range of 200~1,500 bp with a peak around 500~700 bp at highest cycle number. I do not know why I could not get a smaller size range. However, I noticed a substantial amount of foam formation. Is there anybody who can help me on this matter? Any advice or suggestions would be appreciated.

Thanks,

Sung D

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03-24-2010, 11:10 AM	#1
menkelab Junior Member Location: Athens Join Date: Mar 2010 Posts: 2	Chromatin shearing for ChIP-Seq using Covaris S2 Hi, I'm testing shearing conditions using a covaris S2 to shear chromatin to 150~300 bp for ChIP-Seq. My sample consists of mouse embryonic limbs in 500~600 ul buffer (20 mM Tris, pH 8.0; 0.5 mM EGTA, 1mM EDTA) in a TC12 tube. My settings were 20% DC, 5 In, 200 C/B, 30 sec cycle time, varying numbers of cycle (10, 20, 30, 40, 50 cycles). I did not include treatment 2 following treatment 1 for a pause between cycles. I had a size range of 200~1,500 bp with a peak around 500~700 bp at highest cycle number. I do not know why I could not get a smaller size range. However, I noticed a substantial amount of foam formation. Is there anybody who can help me on this matter? Any advice or suggestions would be appreciated. Thanks, Sung D