Why should ATAC-seq mapped reads be shifted +4 and -5 for +strand and -strand, respectively
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6 months ago
progistar • 0

Hello,

I have started to analyze ATAC-seq data and I have a quick question about tag-shift process in data processing. In my best knowledge, there is 9-bp duplication created by DNA repair of the nick by Tn5 transposase.

To achieve base-pair resolution of TF footprint, all researchers do tag-shift processing (shift mapped read position by +4 and -5 for +strand and -strand, respectively); however, I do not understand why the +strand and -strand should be shifted by different size.

Is anyone answers my question?

Thanks for your effort!

ATAC-seq • 442 views
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6 months ago
benformatics ★ 2.1k

The reason is the Tn5 binds as a dimer and includes a 9 bp spacer in between the two cut sites.

My guess is the decision is arbitrary, you are probably free to shift -4 and +5. Your peaks would still be in the same regions and I don't think anyone is doing single-nt resolution ATAC-seq analysis.

See the following (Figure 4): https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3046479/

Here is another mention: "Previous descriptions of the Tn5 transposase show that the transposon binds as a dimer and inserts two adapters separated by 9 bps. " https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3959825/

I'm pretty sure the following is wrong but I included it anyway: "Lastly, reads should be shifted + 4 bp and − 5 bp for positive and negative strand respectively, to account for the 9-bp duplication created by DNA repair of the nick by Tn5 transposase and achieve base-pair resolution of TF footprint and motif-related analyses..."] https://genomebiology.biomedcentral.com/articles/10.1186/s13059-020-1929-3

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To clarify, the goal of the shifting is to identify the center of the Tn5 dimer complex binding event.

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Thanks for giving references. I agree with that single nt resolution does not matter of finding TF binding event.

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