Hello, everyone.
Recently, I did chip-seq (H3K4me1、H3K4me3、H3K27ac、H3K27me3、Pol II、P300) from 1×107 insect cells. But, I got poor yield (about 4ng ) as standard Illumina library preparation needs 10ng and I tried many times. The cell I used is hard to culture, So I want to try some new modified chip-seq protocol in order to get high output. There are a lot of methods about doing chip-seq from small number cells.
For example:
nano-ChIP-seq: Genome-wide chromatin maps derived from limited numbers of hematopoietic progenitors.
LinDA: Single-tube linear DNA amplification (LinDA) for robust ChIP-seq.
Small-scale ChIP: In vivo epigenomic profiling of germ cells reveals germ cell molecular signatures.
cChIP-seq: cChIP-seq: a robust small-scale method for investigation of histone modifications.
These mothods will introduce potential PCR bias due to preamplificationI and I don't know to choose them? cChIP-seq seems better, but it is not clear how does it work about carrier?
Whether do I use more cells to get high yield or select one modified chip-seq protocol from above? Is there any other better methods? Anyone can give some suggestions?
Thank you very much!
ChengDong
Hello, we have optimized the True MicroChIP kit for ChIP-seq on 10.000 cells for histone marks. In combination with our MicroPlex library preparation kit v2 that allows to start libraries from 50pg.
https://www.diagenode.com/p/true-microchip-kit-x16-16-rxns
Article Automating ChIP-seq Experiments to Generate Epigenetic Profi...
Hello, we have optimized the True MicroChIP kit for ChIP-seq on 10.000 cells for histone marks. In combination with our MicroPlex library preparation kit v2 that allows to start libraries from 50pg.
https://www.diagenode.com/p/true-microchip-kit-x16-16-rxns
Article Automating ChIP-seq Experiments to Generate Epigenetic Profi...
We often make libraries from less than 10ng...in some cases from undetectable amounts of DNA.
Try ChIPmentation or Tagmentation. 4 ng DNA (measured by Qubit or Pico Green, not by nanodrop) is more than enough for making a high quality library.
Dear Crosby, did you use standard Illumina library preparation? or other library method? As I will send my sample to sequencing facility and they demand at least 10ng ChIPed DNA.
You can make your own libraries from 50 pg with teh MicroPlex kit. only 3 incubation steps in one tube with no intermediate purification. This protocol has been optimized specifically for ChIP-seq.
https://www.diagenode.com/p/microplex-library-preparation-kit-v2-x12-12-indices-12-rxns
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