Can a count matrix made with GRCr8 reference genome be annotated with org.Rn.eg.db? My attempt to add gene name annotations failed, as described below.
UPDATE: the gene ids are not in ensembl format, so perhaps I need to use a different annotation file? Or a different key?
GRCr8 genome: https://www.ncbi.nlm.nih.gov/datasets/genome/GCF_036323735.1/
Files used to make STAR reference genome: https://ftp.ncbi.nlm.nih.gov/genomes/all/GCF/036/323/735/GCF_036323735.1_GRCr8/GCF_036323735.1_GRCr8_genomic.gtf.gz https://ftp.ncbi.nlm.nih.gov/genomes/all/GCF/036/323/735/GCF_036323735.1_GRCr8/GCF_036323735.1_GRCr8_genomic.fna.gz
# Count matrix
> slice_sample(adult_matrix, n=10)
07 09 58 08 59 03 06 12 05 04 01 10 02 11 60 57
LOC120093222 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
Dcaf17 97 91 74 54 94 76 111 105 70 82 103 68 105 97 69 85
LOC134483108 0 1 0 0 0 1 0 0 0 0 0 0 1 0 0 2
Itga2b 17 23 21 26 29 25 27 39 16 30 33 21 24 25 23 12
Naa20 244 335 191 243 279 221 242 380 210 217 286 220 291 242 202 177
LOC134481053 1 1 0 1 1 1 0 0 0 0 0 0 0 0 1 2
Helb 67 72 39 38 56 54 46 69 41 32 58 31 50 50 33 46
LOC100911384 1 0 0 0 1 1 1 1 0 1 0 0 0 0 1 0
Slc5a7 16 9 4 7 9 6 13 8 9 5 12 10 4 7 41 9
LOC120099428 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0 0
# Meta data for count matrix
> slice_sample(adult_meta, n = 10)
subject diet age timepoint
1 09 CON Adult T28
2 10 CON Adult T28
3 58 EtOH Adult T28
4 07 CON Adult T28
5 59 CON Adult T28
6 01 EtOH Adult T28
7 05 EtOH Adult T28
8 08 CON Adult T28
9 60 CON Adult T28
10 57 EtOH Adult T28
# constructing DeSeq object
adult_dds <- DESeqDataSetFromMatrix(countData = adult_matrix,
colData = adult_meta,
design = ~diet)
# prefiltering, only keeping genes with more than XX reads
adult_keep <- rowSums(counts(adult_dds)) >= 10
adult_dds <- adult_dds[adult_keep,]
# running DESeq
adult_dds <- DESeq(adult_dds)
adult_res <- results(adult_dds, contrast = c("diet", "CON", "EtOH"), alpha = 0.05)
adult_res
summary(adult_res)
# genes ranked by p-value
adult_res_ordered <- adult_res[order(adult_res$pvalue),] #padj?
head(adult_res_ordered)
# DF and gene name annotations
adult_res_df <- as.data.frame(adult_res_ordered)
adult_res_df$symbol <- mapIds(org.Rn.eg.db, keys = rownames(adult_res_df), keytype = "ENSEMBL", column = "SYMBOL")
# Error in .testForValidKeys(x, keys, keytype, fks) :
# None of the keys entered are valid keys for 'ENSEMBL'. Please use the keys method to see a listing of valid arguments.
# compare rownames (genes) to ensembl_ids
> rownames(adult_res_df)
[1] "Sncb" "Ptp4a3" "Zmiz2" "Klc4" "Nicol1" "Stard10" "Rpsa" "Clip3"
[9] "Pld3" "Rnasek"
> available_ensembl_ids <- keys(org.Rn.eg.db, keytype = "ENSEMBL")
> available_ensembl_ids[1:10]
[1] "ENSRNOG00000017701" "ENSRNOG00000028896" "ENSRNOG00000032908" "ENSRNOG00000009845" "ENSRNOG00000016924" "ENSRNOG00000005260"
[7] "ENSRNOG00000013594" "ENSRNOG00000029762" "ENSRNOG00000010265" "ENSRNOG00000049882"
# incompatible formats
> sessionInfo( )
R version 4.4.1 (2024-06-14)
Platform: aarch64-apple-darwin20
Running under: macOS Ventura 13.7.4
Matrix products: default
BLAS: /System/Library/Frameworks/Accelerate.framework/Versions/A/Frameworks/vecLib.framework/Versions/A/libBLAS.dylib
LAPACK: /Library/Frameworks/R.framework/Versions/4.4-arm64/Resources/lib/libRlapack.dylib; LAPACK version 3.12.0
locale:
[1] en_US.UTF-8/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8
time zone: America/Chicago
tzcode source: internal
attached base packages:
[1] stats4 stats graphics grDevices utils datasets methods base
other attached packages:
[1] scales_1.3.0 ggprism_1.0.5 EnhancedVolcano_1.24.0 ggrepel_0.9.6
[5] org.Rn.eg.db_3.20.0 AnnotationDbi_1.68.0 DESeq2_1.46.0 SummarizedExperiment_1.36.0
[9] Biobase_2.66.0 MatrixGenerics_1.18.1 matrixStats_1.5.0 GenomicRanges_1.58.0
[13] GenomeInfoDb_1.42.3 IRanges_2.40.1 S4Vectors_0.44.0 BiocGenerics_0.52.0
[17] BiocManager_1.30.25 afex_1.4-1 lme4_1.1-36 Matrix_1.7-3
[21] rstatix_0.7.2 emmeans_1.10.7 lubridate_1.9.4 forcats_1.0.0
[25] stringr_1.5.1 dplyr_1.1.4 purrr_1.0.4 readr_2.1.5
[29] tidyr_1.3.1 tibble_3.2.1 ggplot2_3.5.1 tidyverse_2.0.0
loaded via a namespace (and not attached):
[1] DBI_1.2.3 rlang_1.1.5 magrittr_2.0.3 compiler_4.4.1 RSQLite_2.3.9
[6] png_0.1-8 vctrs_0.6.5 reshape2_1.4.4 pkgconfig_2.0.3 crayon_1.5.3
[11] fastmap_1.2.0 backports_1.5.0 XVector_0.46.0 labeling_0.4.3 utf8_1.2.4
[16] rmarkdown_2.29 tzdb_0.4.0 UCSC.utils_1.2.0 nloptr_2.2.0 bit_4.6.0
[21] xfun_0.51 cachem_1.1.0 zlibbioc_1.52.0 jsonlite_1.9.1 blob_1.2.4
[26] DelayedArray_0.32.0 BiocParallel_1.40.0 broom_1.0.7 parallel_4.4.1 R6_2.6.1
[31] stringi_1.8.4 RColorBrewer_1.1-3 car_3.1-3 boot_1.3-31 numDeriv_2016.8-1.1
[36] estimability_1.5.1 Rcpp_1.0.14 knitr_1.49 splines_4.4.1 timechange_0.3.0
[41] tidyselect_1.2.1 rstudioapi_0.17.1 abind_1.4-8 yaml_2.3.10 codetools_0.2-20
[46] lattice_0.22-6 lmerTest_3.1-3 plyr_1.8.9 KEGGREST_1.46.0 withr_3.0.2
[51] evaluate_1.0.3 Biostrings_2.74.1 pillar_1.10.1 carData_3.0-5 generics_0.1.3
[56] vroom_1.6.5 hms_1.1.3 munsell_0.5.1 minqa_1.2.8 glue_1.8.0
[61] tools_4.4.1 locfit_1.5-9.12 mvtnorm_1.3-3 grid_4.4.1 colorspace_2.1-1
[66] nlme_3.1-167 GenomeInfoDbData_1.2.13 Formula_1.2-5 cli_3.6.4 fansi_1.0.6
[71] S4Arrays_1.6.0 gtable_0.3.6 digest_0.6.37 SparseArray_1.6.2 farver_2.1.2
[76] memoise_2.0.1 htmltools_0.5.8.1 lifecycle_1.0.4 httr_1.4.7 bit64_4.6.0-1
[81] MASS_7.3-65