Hi you all,

I am trying to perform featureCount over miRNA reads from CHO cells for differential expression analysis purposes. I have created my own annotation file by mapping hairpin miRNA sequences downloaded from mirBase (http://www.mirbase.org/cgi-bin/mirna_summary.pl?org=cgr) against ENSEMBL reference genome for CHO cell. For that purpose, I have used gmap -D:

gmap -D ~/miRNA/crigri_gmap -d crigri_gmap -f 2 -n 0 -t 16 --gff3-cds=genomic hairpin_crigri_dna_mod.fa > trial_1.gff3

I retrieve a .gff file with 9 columns containing the 9 expected fields (seqid,source,type,start,end,score,strand,phase and attributes), but for some scaffolds, I get a "-1" value in the "phase" field. According to ENSEMBLE (https://www.ensembl.org/info/website/upload/gff3.html) 'One of '0', '1' or '2'. '0' indicates that the first base of the feature is the first base of a codon, '1' that the second base is the first base of a codon, and so on.' So I don't know how to interpret this value.

This is how the head of my .gff3 file looks:

scaffold_6      crigri_gmap     gene    62246878        62246977        .       +       .       ID=cgr_let_7a_MI0020368.path1;Name=cgr_let_7a_MI0020368

scaffold_6      crigri_gmap     mRNA    62246878        62246977        .       +       .       ID=cgr_let_7a_MI0020368.mrna1;Name=cgr_let_7a_MI0020368;Parent=cgr_let_7a_MI0020368.path1;coverage=100.0;identity=100.0;matches=100;mismatches=0;indels=0;unknowns=0

scaffold_6      crigri_gmap     exon    1       62246878        100     +       .       ID=cgr_let_7a_MI0020368.mrna1.exon1;Name=cgr_let_7a_MI0020368;Parent=cgr_let_7a_MI0020368.mrna1;Target=cgr_let_7a_MI0020368 1 1 +

scaffold_6      crigri_gmap     CDS     62246878        62246975        100     +       -1      ID=cgr_let_7a_MI0020368.mrna1.cds1;Name=cgr_let_7a_MI0020368;Parent=cgr_let_7a_MI0020368.mrna1;Target=cgr_let_7a_MI0020368 1 98 +

Any help with this will be much appreciated. Thank you very much!!