[miso-users] Questions from an inexperienced user re: GFF3 files for hg38, GRCh38

[Ackerman, William]

Dear MISO Development Team:

I have some RNA-Seq data from human tissues (Illumina, 100-bp paired reads) that were mapped to the lastest Ensembl and UCSC genome builds (GRCh38 and hg38, respectively) using Tophat2.  I am interested in performing an initial analysis of alternative splicing using MISO, in preparation for Iso-Seq on similar samples using the PacBio platform.

I have sucessfully loaded the MISO package and its dependencies (scipy was holding me up for several days), and the installation appears to be successful according to the diagnostics recommended in the helpful MISO documentation.

However, I have much confusion over the "annotations (typically specified in GFF format) of the isoforms of alternative events" that must be provided.  I assume that the only human annotations available are those available on https://miso.readthedocs.org/en/fastmiso/annotation.html, as I have not been able to find other sources of MISO-compatible GFF files online.  I wondered if there was a way to produce similar files for the more recent assemblies (either UCSC or Ensembl), or if it would be best to map using one of the assemblies for which the GFF3 files are provided?  (As a side note, I have not been successful with installing the rnaseqlib-chip software to my virtual Python environment in the cluster I am using).  Also, I notice that there are two versions of these annotation files (1 and 2), and each has multiple GFF3 files.  If one were to use these files, should one concatenate the GFF3s and run once for each concatenated BAM file (I have three phenotypical groups), or run these in parallel?

Thanks in advance for any help you could kindly provide,

William E. Ackerman IV, M.D.
Dept. of Obstetrics & Gynecology
Div. of Maternal-Fetal Medicine
N521 Doan Hall
410 West 10th Avenue
Columbus, OH  43210

Tel: 614/293-4438
E-mail: William.Ackerman at osumc.edu; ackerman.72 at osu.edu

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