Hello everybody,
I am PhD student and i am working with a nonmodel tree under drought stress. I want to know if i can use GSEA in my experimental design with my nonomodel plant. One of my experiments consist in three control plants and three stressed plants. I took total RNA and performed a RNA Sequencing, "de novo" assembly and DE analysis, thus I have about 190.000 genes with its normalized counts (TMM). I could create the files: data set (.gct) and phenotypes labels (.cls) .
1) But I can´t or I don´t know how to create a Gene sets file (.gmt) matched with GO terms because my IDs data set file comes from Illumina, they are like: c0001_g1. And there is more,
2) I do not quite understand if it is necessary to have a chip necessarily to run the analysis.
I would be very grateful in an answer adapted for biologists not specialists in bioinformatics.
Thnaks
Edgardo,
Hi Edgardo,
Technically, you still can perform GSEA even though you are using non-model plant. Some of the necessary steps/files are as follows:
1. a gmt file. I don't reckon that there is a readily available gmt file for your plant. So, you may be able to use the gmt file of the closest plant relative to your species. This publication (https://pubmed.ncbi.nlm.nih.gov/23632162/) and this database (http://systemsbiology.cau.edu.cn/PlantGSEA/download.php) may help.
2. You will need to map your gene ID to match that of the gmt file. To do so, you will need to annotate the transcriptome which you have assembled. I am not familiar with the annotation process, but this manuscript may help (Article De-novo transcriptome assembly for gene identification, anal...
). After the transcriptome annotation, you can map your transcripts to genes. If the gene ID matches that of the gmt file, the files can be used directly for GSEA. If not, you will need to remap the gene ID before GSEA. BiomaRt (https://plants.ensembl.org/biomart/martview/76f17c602cf7f4cb9adc75e9a97c87df) may be helpful in this case.I hope this helps.
Regards
Hong Sheng
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