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Question: MACE (Massive analysis of cDNA ends)
1
gravatar for Philippe
2.1 years ago by
Philippe • 10
Germany / Freiburg / Albert-Ludwigs-Universität
Philippe • 10 wrote:

Hi,

does anybody have experience in doing downstream analysis on data created by the MACE-technology (Massive Analysis of cDNA ends)?

What are important differences compared to a regular analysis of mRNA or miRNA?

I know, it´s kind of a wide, non-precise question, but I just started working on that kind of data and I´m kind of lost.

Any kind of helpful advise would be much appreciated.

Cheers,

Philippe
rna-seq galaxy • 783 views
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modified 2.1 years ago by Mo Heydarian ♦ 830 • written 2.1 years ago by Philippe • 10
1
gravatar for Mo Heydarian
2.1 years ago by
Mo Heydarian ♦ 830
United States
Mo Heydarian ♦ 830 wrote:

Hi Philippe,

I'm not super familiar with this RNA sequencing strategy, but from looking at this paper (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3928179/) it seems like MACE is a strategy for specifically sequencing the 3' ends of RNA transcripts. The libraries will have a large polyA stretches within the reads (oligo-dT is used to prime the reverse transcriptase), so you would need to perform a clipping/trimming step prior to alignment and subsequent abundance estimation.

For trimming the polyA stretches, have a look at the tools: Clip, Trimmomatic, Manipulate FASTQ, and Trim Galore, these are under the NGS: QC and manipulation category. For alignment you should use Tophat or HISAT, these tools can deal with mapping spliced reads if you capture splice junctions in your MACE library. To quantify abundance estimates you could use Cuffdiff or HTseq-count with DEseq2. These tools are all available on useGalaxy.org.

Hope this helps!

Cheers, Mo Heydarian, Galaxy Team
ADD COMMENT • link written 2.1 years ago by Mo Heydarian ♦ 830

Cheers for your response Mo!

This is the paper I also found while checking pubmed. I´ll try the procedures you mentioned.

Have a nice one.

Regards, Philippe

ADD REPLY • link written 2.1 years ago by Philippe • 10
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