Question: Why chip-seq is strand specific?
1
gravatar for bxia
2.4 years ago by
bxia140
bxia140 wrote:

Why chip-seq is strand specific?

The antibody will bind to double strand DNA, right?

chip-seq • 1.6k views
ADD COMMENTlink modified 2.4 years ago by spacemorrissey200 • written 2.4 years ago by bxia140
2
gravatar for spacemorrissey
2.4 years ago by
United States
spacemorrissey200 wrote:

ChIP-seq itself is not strand specific. The reads generated when performing ChIP-seq are strand specific, which is important. The reads do not indicate the location of the TF binding or histone modification, they are at the end of the fragment that includes the TF binding position. The fact that the reads on the two strands should be seperated by ~ the average fragment size of your library is an important quality check for the data.

ADD COMMENTlink written 2.4 years ago by spacemorrissey200

How do you reconcile this answer with Devon's?

ADD REPLYlink written 2.4 years ago by Brice Sarver2.6k

RNA-seq can be "stranded" because one strand is transcribed into an mRNA. This is interesting because there are cases where a protein coding gene can be transcribe from one strand, and a non-coding RNA can be transcribed from the opposite strand. Chip-seq doesn't really work that way. You are just looking for the location of a TF binding event or Histone modification. The TF binds to a fragment of double stranded DNA. The first ~50 bases of both strands are sequenced, leading to reads that are separated by the size of the original double strand fragment. While you will not be able to deduce from these reads weather the TF binds primarily to one strand or both. You can use the distance between the reads as a quality measure. See the work of Anshul Kundaje - https://sites.google.com/site/anshulkundaje/projects/idr - "It is important to check the cross-correlation plot that is produced by SPP to make sure the estimated fragment length is appropriate."

ADD REPLYlink written 2.4 years ago by spacemorrissey200
0
gravatar for Devon Ryan
2.4 years ago by
Devon Ryan89k
Freiburg, Germany
Devon Ryan89k wrote:
  1. It's generally not strand specific.
  2. The antibody doesn't bind DNA, it binds a protein that binds DNA. Of course, the protein is typically binding to both strands of DNA (often one of the grooves), which goes back to point 1.
ADD COMMENTlink written 2.4 years ago by Devon Ryan89k
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