Improve Research Reproducibility A Bio-protocol resource
Atac-Seq
Sequeencing

Why no nucleosomal ladder pattern in my single-cell ATAC-seq libraries?

SH
Shuai Han May 23, 2022

I tried several protocols to do single-cell ATAC-seq. But the profiles of my libraries were as follows, with no prominent nucleosomal ladder pattern that a successful library should have. What would be the possible reasons and how to solve the problem? Any suggestions would be helpful. img Read more Read less more less

answer Answer
recommend recommend recommend recommend Recommend
follow follow follow follow Follow
share share Share
1 answer
Sort by: most helpful / most recent

liu lei Answered Jun 24, 2022

china agriculture university

  1. The lower right of the results is better. The two fragments on the left are too small, which should be non-specific lysis caused by excessive lysis and excessive chromosome opening. The fragments on the upper right are too large and insufficient cut, which may be related to incomplete cell lysis or low efficiency of transposase (it is uncertain whether the transposase and buffer used in the four protocols are consistent. If they are inconsistent, it may be related). 
  2. According to the results at the bottom right, the PCR products may not have been sorted. It is recommended to sort when there are a large number of cells, and a better nucleosomal ladder will be obtained. However, the output of scATAC is low, so sorting is not recommended. Therefore, the results at the bottom right are more in line with the peak shape of ATAC products that are not sorted. You can try sequencing, but you may need a larger amount of sequencing to filter out useful information.
1 helpful
0 unhelpful
comment 0 comments down up
share Share

My answer

Write your answer...

References (optional)

add Add more

post Post an Answer