Normalizing Bam Files
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11.1 years ago
ChIP ▴ 600

Hi!

I am back again with one more naive question.

What according to you is the best way to Normalise a BAM file (read density) and why it is the best available method?

Kindly, name the tool(s) you use to normalise the BAM files.

More Details: Say for instance I have ChIP-seq of Histone marks from patient sample (H3K36me3,H3K4me3,H3K27ac,H3K4me1), and is mapped using BWA.

Thank you

chip-seq • 7.2k views
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The normalization methods will depend on the methods to generate and analyze the data and the biases that exist and accumulate in the generation of the data. It would be good if you tell people what sort of data is contained in the BAM file, and if the BAM files! have a relation to each other.

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I made an edit, I hope it helps in answering the question.

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Do you mean, read density normalization??

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yes, I mean read density normalization.

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I am not sure about bam file but had you already thought of MA plot?

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11.1 years ago
Ido Tamir 5.2k

What do you want to do?

  1. peak calling: unless the read numbers are not too different, the peak caller will normalize input and IP track. manual downsampling is also an option
  2. generation of bigwig tracks to visualize: read count normalization to RPM, optionally extend reads in read direction by fragment length (MACS can do this for you)
  3. there are other normalization methods to generate more "even" tracks that take into account GC content or similar, but they are not in widespread use.
  4. Quantitative differences e.g. diffbind have their own normalization methods.

.... I guess there are other use cases and then you would do something else entirely

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