Dear all,
'd appreciate please having your advise on the process of adding a transgene (it is called "mCherry_4R)" to the mouse reference sequence mm10 in order to be fully analyzed with Seurat ('ve been using Seurat 2.3.4) :
the initial steps work well, however, at the end there is a bit of issue ...
I've followed the steps :
1 . 've updated the GTF file of mm10 genes
2. 've updated the FASTA file of mm10
3. 've made a new reference genome sequence : it is called " mm10_with_mCherry_4R"
4. 've aligned the scRNA-seq FASTQ to the new reference genome sequence
and in the file features.tsv (genes.tsv), I do see the TRANS-GENE:
ENSMUSG00000096730 Vmn2r122 Gene Expression
ENSMUSG00000095742 CAAA01147332.1 Gene Expression
mCherry_4R mCherry_4R Gene Expression
However, after I reading the matrix/ feature /barcodes in Seurat 2.3.4, the transgene "mCherry_4R" is not in the matrix data of Seurat (we know it is expressed)..
Is there a particular way we can fix it ? could it be a problem with the name of the trans-gene ? or is there something else that we do potentially miss ?
thank you,
-- bogdan
How many cells is it expressed in? How many total counts does it have? You should check this in the original matrix before you import it with Seurat.