Troubleshooting for Deseq2 annotation

Hi, I am doing RNA-seq analysis, the pipeline is as follows: upload data > FastQC > Trimmomatic > HISAT2 > Feature counts > Deseq 2 > Annotate Deseq 2 output tables. The reference I used for annotation is “mm10. refgene.gtf.” However, the output shows just NA, like picture showed below, what is the problem?

Thanks for help!

Dear @Xiangdong,
Have you used just one sample (replicate) for DESeq 2? DESeq2 does not calculate p-values (signifiance) if you just use one replicate.

Kind regards,
Florian

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Dear Florian,

Thank you for your response, I have 4 replicates in each group. I just figure out the problem. When I did feature count, I used built-in mm10. When I annotate Deseq2 output, I used mm10 from other website. If I use the same mm10 when I did “feature count” and “annotate Deseq 2 output”. It works.

Thanks for your help

Xiangdong

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