Data postprocessing
Transcriptome location to genome location
In modification detection tasks that utilize Direct RNA Sequencing (DRS) data, where sequences are mapped to transcripts, it is often necessary to convert transcriptome locations to genome locations for comparison with other tools or experimental results. The script scripts/transcriptome_location_to_genome_location.py will take TandemMod prediction file as input and convert the transcriptome locations to genome locations.
usage: transcriptome_location_to_genome_location.py [-h] -i INPUT -o OUTPUT -g GFF
Convert transcriptome location to genome location.
optional arguments:
-h, --help show this help message and exit
-i INPUT, --input INPUT Input file,transcriptome location.
-o OUTPUT, --output OUTPUT Output file, genome location.
-g GFF, --gff GFF Annotation file.
python scripts/transcriptome_location_to_genome_location.py \
--input predictions.tsv \
--output predictions_genome_loc.tsv \
--gff /path/to/your/genome_annotation.gtf
In the above command, replace “predictions.tsv” with the actual TandemMod prediction file you have, and specify the desired output file name using --output. The script will read the input prediction file, convert the transcriptome locations to genome locations, and save the converted predictions in the specified output file. Additionally, for the conversion, the script requires a genome annotation file in GFF (General Feature Format) format. The GFF file contains genomic coordinates and annotations for various features such as genes, exons, and transcripts.
Here is an example to illustrate the transcriptome locations format:
transcript_id site motif read_id prediction probability
LOC_Os08g44480.1 218 GACCA b32995bf-9975-4ff6-86c6-29c900f321d6 C 0.11261273
LOC_Os08g44480.1 219 ACCAG b32995bf-9975-4ff6-86c6-29c900f321d6 m5C 0.95669556
LOC_Os08g44480.1 223 GGCCA b32995bf-9975-4ff6-86c6-29c900f321d6 C 4.225098e-14
LOC_Os08g44480.1 224 GCCAC b32995bf-9975-4ff6-86c6-29c900f321d6 C 6.3074664e-11
LOC_Os08g44480.1 226 CACCT b32995bf-9975-4ff6-86c6-29c900f321d6 C 5.365696e-05
LOC_Os08g44480.1 227 ACCTA b32995bf-9975-4ff6-86c6-29c900f321d6 C 1.9726056e-06
LOC_Os08g44480.1 230 TACGA b32995bf-9975-4ff6-86c6-29c900f321d6 C 0.00010113342
LOC_Os08g44480.1 233 GACAA b32995bf-9975-4ff6-86c6-29c900f321d6 C 4.4892527e-11
LOC_Os08g44480.1 237 AGCTG b32995bf-9975-4ff6-86c6-29c900f321d6 C 5.053165e-10
LOC_Os08g44480.1 240 TGCTC b32995bf-9975-4ff6-86c6-29c900f321d6 C 3.4177435e-16
LOC_Os08g44480.1 242 CTCTC b32995bf-9975-4ff6-86c6-29c900f321d6 C 0.031539176
LOC_Os08g44480.1 244 CTCCG b32995bf-9975-4ff6-86c6-29c900f321d6 C 0.08256312
LOC_Os08g44480.1 245 TCCGA b32995bf-9975-4ff6-86c6-29c900f321d6 C 0.0004769674
LOC_Os08g44480.1 252 TGCCC b32995bf-9975-4ff6-86c6-29c900f321d6 C 2.2711574e-30
LOC_Os08g44480.1 253 GCCCA b32995bf-9975-4ff6-86c6-29c900f321d6 C 7.629347e-21
Here are the converted results in genome locations.
transcript_id site chr site motif prediction probability
LOC_Os06g36590.1 826 Chr6 21495252 ATCGG C 4.780567e-05
LOC_Os06g36590.1 831 Chr6 21495257 ATCTT C 3.1403553e-09
LOC_Os06g36590.1 836 Chr6 21495262 GTCGT m5C 0.998418
LOC_Os06g36590.1 850 Chr6 21495276 GTCTG C 2.6889346e-08
LOC_Os06g36590.1 853 Chr6 21495279 TGCTA C 7.5095416e-11
LOC_Os06g36590.1 856 Chr6 21495282 TACAT C 0.013431426
LOC_Os06g36590.1 871 Chr6 21495297 TGCCT C 2.3783017e-05
LOC_Os06g36590.1 872 Chr6 21495298 GCCTT C 8.105786e-09
LOC_Os06g36590.1 877 Chr6 21495303 TGCTT C 1.7306347e-12
LOC_Os06g36590.1 881 Chr6 21495307 TGCCT C 1.315495e-09
LOC_Os06g36590.1 882 Chr6 21495308 GCCTT C 6.700999e-10
LOC_Os06g36590.1 888 Chr6 21495314 GGCAA C 4.613933e-09
LOC_Os06g36590.1 891 Chr6 21495317 AACCT C 6.412733e-07
LOC_Os06g36590.1 892 Chr6 21495318 ACCTG C 2.1830398e-05
LOC_Os06g36590.1 901 Chr6 21495445 TACAG C 0.009539205
LOC_Os06g36590.1 912 Chr6 21495456 TACTT C 0.3794193
LOC_Os06g36590.1 916 Chr6 21495460 TGCAG C 0.08652508
LOC_Os06g36590.1 922 Chr6 21495466 ATCTA C 5.3866494e-05
Read-level predictions to site-level predictions
TandemMod provided single-read predictions as well as their modification probabilities. By aggregating all the predictions for each site, we can derive a consensus or summary prediction for that specific genomic location. The script is located under scripts/read_level_prediction_to_site_level_prediction.py
usage: read_level_prediction_to_site_level_prediction.py [-h] -i INPUT -o OUTPUT
Convert read-level predictions to site-level predictions.
optional arguments:
-h, --help show this help message and exit
-i INPUT, --input INPUT Input file,transcriptome location.
-o OUTPUT, --output OUTPUT Output file, genome location.
python scripts/read_level_prediction_to_site_level_prediction.py \
--input read_level_prediction.tsv \
--output site_level_prediction.tsv
In the given command, please replace “read_level_prediction.tsv” with the converted results obtained from the transcriptome_location_to_genome_location.py script. Specify the desired output file name using the --output option. The script will then aggregate the read-level predictions to derive site-level predictions. The resulting predictions will include the count of modified bases, considering the predictions with probability values ranging from 0.5 to 0.95 as the cutoff range. The total base count is located in the last column.
transcriptome_id site chr site motif p_0.5 p_0.6 p_0.7 p_0.8 p_0.9 p_0.95 total
LOC_Os05g41060.1 445 Chr5 24059817 TGCGC 2 2 2 2 2 1 63
LOC_Os05g41060.1 447 Chr5 24059815 CGCCA 1 0 0 0 0 0 63
LOC_Os05g41060.1 448 Chr5 24059814 GCCAG 4 4 4 4 2 1 63
LOC_Os05g41060.1 451 Chr5 24059811 AGCGG 10 8 7 6 3 2 63
LOC_Os05g41060.1 454 Chr5 24059808 GGCAC 15 14 14 11 10 8 63
LOC_Os05g41060.1 456 Chr5 24059806 CACTG 2 2 2 2 2 1 63
LOC_Os05g41060.1 462 Chr5 24059800 TACAT 0 0 0 0 0 0 63
LOC_Os05g41060.1 465 Chr5 24059797 ATCCA 6 6 6 5 5 3 63
LOC_Os05g41060.1 466 Chr5 24059796 TCCAG 2 2 2 1 1 0 63
LOC_Os05g41060.1 471 Chr5 24059791 AGCAC 11 8 7 5 3 2 63
LOC_Os05g41060.1 473 Chr5 24059789 CACAT 1 1 1 0 0 0 63
LOC_Os05g41060.1 479 Chr5 24059783 TGCTA 1 1 1 1 0 0 63
LOC_Os05g41060.1 482 Chr5 24059780 TACCT 5 5 5 4 2 2 63
LOC_Os05g41060.1 483 Chr5 24059779 ACCTC 5 5 5 5 4 4 63
LOC_Os05g41060.1 485 Chr5 24059777 CTCTG 4 4 4 3 2 1 63
LOC_Os05g41060.1 508 Chr5 24059754 GGCTT 2 2 2 1 1 1 63
Futher analysis
The other data processing and plot scripts are located under the plot directory.