Hello, I am planning a ChIPseq of Arabidopsis looking at a TF binding. My current plan is to do an induced inflorescence, uninduced inflorescence, and leaf tissue ChIP. I am generally following the standard plant ChIPseq protocol in “Chromatin techniques for plant cells” Bowler 2004. The problem is I have limited amounts of tissue for the induced and uninduced set. Around 0.15g each. Whereas I usually see tissue amounts used around 1g-0.5g. I was wondering if I can just continue on as usual or if there was any modification I should do to the methods or subsequent library prep step. Should I add more controls like input even with the limited tissue?
Hi,Ad!
About library preparation, there is a lot of kits for wide range of input concentration. For example, Nebnext Ultra II (starts from 500 pg).
I'm working with Arabidopsis, but I do RIP experiment. In my case I use 0.1 g of leaf tissue. But I have transgenic plant with high level of protein.
If you use non-transgenic plants and planning to use antibody for the original TF, 150 mg could be insufficient.
Anyway, I dare you to start from this amount and look at result.
Hi,
I think you can take a look this protocol from Wagner Lab. They also use very low amount of starting material (300mg-600mg).
Article PROTOCOL: Chromatin Immunoprecipitation from Arabidopsis Tissues
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