# Mouse Whole Cortex and Hippocampus SMART-seq intron counts not in hdf5 expression matrix file? **URL:** \ **Category:** General\ **Tags:** atlas-mouse-brain-adult, transcriptomics, rna-seq\ **Created:** [February 28, 2023, 7:51pm UTC](https://community.brain-map.org/t/mouse-whole-cortex-and-hippocampus-smart-seq-intron-counts-not-in-hdf5-expression-matrix-file/2185 "2023-02-28T19:51:53Z")\ **Posts on this page:** 2\ **Page:** 1
**Author:** ![MatiNemera](https://avatars.discourse-cdn.com/v4/letter/m/65b543/32.png) [@MatiNemera](https://community.brain-map.org/u/MatiNemera)\ **Post date:** [February 28, 2023, 7:51pm UTC](https://community.brain-map.org/t/mouse-whole-cortex-and-hippocampus-smart-seq-intron-counts-not-in-hdf5-expression-matrix-file/2185/1 "2023-02-28T19:51:53Z")
Hello, I am working with the Allen smart-seq data posted here ([Mouse Whole Cortex and Hippocampus SMART-seq - brain-map.org](https://portal.brain-map.org/atlases-and-data/rnaseq/mouse-whole-cortex-and-hippocampus-smart-seq)). I’m hoping to use Seurat to eventually integrate this smartseq dataset with the 10x dataset here ([Mouse Whole Cortex and Hippocampus 10x - brain-map.org](https://portal.brain-map.org/atlases-and-data/rnaseq/mouse-whole-cortex-and-hippocampus-10x)) to use as a reference dataset, and the 10x HDF5 expression matrix file does seem to have both introns and exons combined. However, while the smartseq HDF5 expression matrix file is described as having both introns and exons combined, the labels of the expression matrix file suggest only exon counts are included. Is this a mistake or does the HDF5 file really only contain the exon counts? What I see in R for the smartseq dataset: > library(rhdf5) > h5ls(“expression\_matrix.hdf5”) > group name otype dclass dim > 0 / data H5I\_GROUP > 1 /data exon H5I\_GROUP > 2 /data/exon dims H5I\_DATASET INTEGER 2 > 3 /data/exon i H5I\_DATASET FLOAT 703397415 > 4 /data/exon p H5I\_DATASET INTEGER 45769 > 5 /data/exon x H5I\_DATASET FLOAT 703397415 > 6 /data t\_exon H5I\_GROUP > 7 /data/t\_exon dims H5I\_DATASET INTEGER 2 > 8 /data/t\_exon i H5I\_DATASET FLOAT 703397415 > 9 /data/t\_exon p H5I\_DATASET INTEGER 73364 > 10 /data/t\_exon x H5I\_DATASET FLOAT 703397415 > 11 /data total\_exon\_counts H5I\_DATASET FLOAT 73363 > 12 / gene\_names H5I\_DATASET STRING 45768 > 13 / sample\_names H5I\_DATASET STRING 73363 Thank you for any help! ---
**Author:** ![Susan.Sunkin](https://avatars.discourse-cdn.com/v4/letter/s/6a8cbe/32.png) [@Susan.Sunkin](https://community.brain-map.org/u/Susan.Sunkin)\ **Post date:** [March 3, 2023, 11:42am UTC](https://community.brain-map.org/t/mouse-whole-cortex-and-hippocampus-smart-seq-intron-counts-not-in-hdf5-expression-matrix-file/2185/2 "2023-03-03T11:42:50Z")
Hi, I asked one of our bioinformatics staff members to address your question and here is his response: The Mouse SMARTerV4 reference is rsem, which means it only has exonic reads (we haven’t used rsem alignments since 2017). None of the genome reference files are current and I don’t think any contain intron/exon counts. • Mouse rsem\_GRCm38.p3.gtf.zip is an rsem reference, which means alignment only to transcripts (no genomic or intronic alignments). Again we haven’t used any rsem since Q4 2017. • Human rsem\_GRCh38.p2.gtf.zip is the same as mouse. This would have to be an incomplete release of Human SMARTerV4 MTG data. • Mouse mouse\_10x\_gtf.zip is likely our 10X Cell Ranger v3 reference. This reference has all introns reclassified to exons (premRNA reference). This was done, so both intron and exon alignments are included in the count matrix (Cell Ranger v3 did not have an option for this). For any SMARTerV4 data anything aligned with STAR will have separate intron/exon count matrices. For 10X only Cell Ranger 6 aligned data will have intron/exon counts. Additionally, since the data sets use different references integrating them will be difficult. Thank you