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RNA-seq: identifying pericentromeric region

Hi everyone, I'm new in bioinformatics and I'm currently trying to identify the pericentromeric region (satellites, families and sub-families) using RNA-seq (long non-coding and pair-end reads) data from samples from cancer patients. I'm not quite sure what the pipeline should be as this is my first time working with RNA-seq, what I'm currently trying to do is: I know that there are repeats with the sequence "TTGGA" that belong to this specific region, so I'm trying to do a manual search of the reads that contain at least 5 of these repeats to then check with IGV if they fall within the pericentromere. Has anybody worked with RNA-seq data and could help me out what the best pipeline to achieve this task should be? Thanks a lot in advance.

genomics transcriptomics rna-seq assembly aligning
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updated by William • William • written by Polly • Polly

When you say "RNA-seq", what sort of RNA-seq is it - illumina?

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written by i.sudbery i.sudbery

1 answer

If this is standard RNA-seq data, then

  1. Reads coming from the pericentromeric satellites repeats, I think its very unlikely that you will be able to align the reads, thus you are unlikely to see them in IGV. Mostly likely these will be unaligned genes.
  2. Do we expect perimcentromeric satelites to be transcribed? It was my understanding that transcription from these regions is extremely low, and therefore these sequences are not likely to be present in RNA-seq data, although I could be wrong.
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written by i.sudbery i.sudbery

@i.sudbery Yes, pericentromeric satellites are apparently expressed in certain contexts, for example :

"At the moment of fertilization the organization of pericentromeric maternal and paternal heterochromatin differs considerably,Citation19 although both are equivalent by the 8-cell stage.Citation20,21 During this early stage of development in mice a particular heterochromatin state is acquired that requires transcription of a pericentromeric repetitive sequence, known as major satellite RNA, that is initiated by a burst in synthesis of the forward major satellite strand. At the 4-cell stage, transcription of both strands is downregulated. Disruption of this transcriptional heterochromatin regulation using locked-nucleic acid (LNA)-DNA gapmers that target major transcripts leads to developmental arrest at the 2-cell stage.Citation22 These repetitive transcripts are detected again in advanced stages of mouse development and are ubiquitously expressed in several tissues. However, the expression pattern in the adult is more restricted and tissue-specific.Citation23" - from Cebrià-Costa, J. P., Millanes-Romero, A., de Herreros, A. G., & Peiró, S. (2014). The Epithelial-to-Mesenchymal Transition (EMT), a Particular Case. Molecular & Cellular Oncology, 1(2). https://doi.org/10.4161/23723548.2014.960770

I'm also curious if there would be any way to quantify total satellite repeat expression with standard RNA seq. It would be tough to get reads to map to any specific satellite sequence locus, but maybe add a major satellite repeat as a custom chromosome in a custom reference, and then just count the number of reads that align to it for total satellite sequence expression?

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written by William • William

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