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