2022-2023_RRP6-NAB3

Table of contents

1. Project description
2. Dependencies
3. Directory structure
4. What to make available, what not to make available
   1. 2023-0111
      1. Make available
         1. Directories
         2. Files
      2. Don't make available
         1. Directories
         2. Files
   2. 2023-0115
      1. Make available
         1. Directories
         2. Files
      2. Don't make available
         1. Directories
         2. Files
      3. General
   3. 2023-0215
      1. Make available
         1. Directories
         2. Files
      2. Don't make available
         1. Directories
         2. Files
      3. Maybe
         1. Directories
         2. Files
5. Copyright

Project description

Project description

Using yeast as a model organism, this project aims to understand how the transcriptome changes during the entry into quiescence, a reversible non-replicative state. In comparison to the transcriptomes of cycling yeast, a significant portion of the quiescent transcriptome is dedicated to noncoding transcription. To explore this noncoding transcription, we used data from 4tU-seq, an NGS assay that enables the isolation of quick-decaying nascent transcription, for transcript assembly and annotation in quiescent yeast.

In addition to nascent transcription, 4tU-seq allows us to isolate steady-state transcripts. During our transcriptome analyses, we found distinct differences between the steady-state and nascent transcriptomes in quiescence, particularly in their compositions. These observation led us to generate and analyze 4tU-seq data from deletion and depletion models for post-transcriptional regulators. The scripts and notebooks in this repository detail the analysis portion of that work, including:

1. Pre-processing, aligning, and post-processing sequenced paired-end reads
2. Drafting nascent transcriptome assemblies for cells in the quiescent (Q) and G1 states
3. Filtering and annotating the draft assemblies
4. Conducting various bioinformatics and statistical analyses and visualizing the results

Dependencies

Dependencies

#TODO

Directory structure

Directory structure

2022-2023_RRP6-NAB3
├── bin
├── data
└── results
      ├── 2023-0111  # Transcriptome assembly
      │     ├── tutorial_troubleshooting
      │     └── work_initial
      ├── 2023-0115  # Pre-processing, aligning, and post-processing sequenced paired-end reads
      │     ├── etc_QC
      │     ├── etc_cleaning
      │     ├── etc_initial
      │     ├── notebook
      │     └── test_tutorial
      └── 2023-0215  # Everything else
            ├── GEO
            ├── bws
            ├── infiles_gtf-gff3
            │     ├── Trinity-GG
            │     │     ├── G_N
            │     │     └── Q_N
            │     ├── already
            │     ├── comprehensive
            │     │     └── S288C_reference_genome_R64-1-1_20110203
            │     └── representation
            │         ├── CUTs-HMM_CUTs-4X
            │         ├── CUTs_SUTs
            │         ├── NUTs
            │         ├── SRATs
            │         ├── XUTs
            │         └── ncRNAs
            ├── notebook
            ├── outfiles_gtf-gff3
            │     ├── Trinity-GG
            │     │     ├── G_N
            │     │     │     ├── err_out
            │     │     │     └── filtered
            │     │     │         ├── CDS
            │     │     │         ├── exon
            │     │     │         ├── introns_filtered
            │     │     │         ├── locus
            │     │     │         └── mRNA
            │     │     └── Q_N
            │     │         ├── err_out
            │     │         └── filtered
            │     │             ├── CDS
            │     │             ├── exon
            │     │             ├── introns_filtered
            │     │             ├── locus
            │     │             └── mRNA
            │     ├── already
            │     │     └── sgd-related
            │     ├── comprehensive
            │     │     └── S288C_reference_genome_R64-1-1_20110203
            │     └── representation
            └── outfiles_htseq-count
                ├── Trinity-GG
                │     ├── G_N
                │     │     ├── err_out
                │     │     ├── filtered
                │     │     │     └── locus
                │     │     │         └── err_out
                │     │     ├── list
                │     │     └── sh
                │     └── Q_N
                │         ├── err_out
                │         ├── filtered
                │         │     └── locus
                │         │         └── err_out
                │         ├── list
                │         └── sh
                ├── already
                │     ├── combined-AG
                │     │     ├── CUT
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── CUT_2016
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── CUT_4X
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── NUTs
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── SRAT
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── SUT
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── XUT
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── antisense_transcript
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── mRNA
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── ncRNA
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── rRNA
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── snRNA
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     ├── snoRNA
                │     │     │     ├── UTK_prim_UMI
                │     │     │     │     └── err_out
                │     │     │     └── UT_prim_UMI
                │     │     │         └── err_out
                │     │     └── tRNA
                │     │         ├── UTK_prim_UMI
                │     │         │     └── err_out
                │     │         └── UT_prim_UMI
                │     │             └── err_out
                │     └── combined-SC-KL-20S
                │         ├── UTK_prim_UMI
                │         │     └── err_out
                │         ├── UTK_prim_no
                │         │     └── err_out
                │         ├── UTK_prim_pos
                │         │     └── err_out
                │         ├── UT_prim_UMI
                │         │     └── err_out
                │         ├── UT_prim_no
                │         │     └── err_out
                │         └── UT_prim_pos
                │             └── err_out
                └── representation
                    └── UT_prim_UMI
                        └── err_out

For clean up

2023-0111

2023-0111

Make available

Directories

Files

• work_GMAP_rough-draft.md
  • Has initial description
  • Makes and works with paths within directory 2023-0111
• work_Trinity-GF-GG-optimization_submit-jobs.md
  • Has initial description
  • Has absolute paths in Code
  • Has absolute paths in Printed
• README.md #TODO Needs to be rewritten to explain what is run with what and in what order

Don't make available

Directories

• tutorial_troubleshooting
• work_initial #TODO #MAYBE Delete contents and subdirectory

Files

• check_Trinity-jobs.md

2023-0115

2023-0115

Make available

Directories

Files

• README.md #TODO Needs to be rewritten to explain what is run with what and in what order
• work_MultiQC.md
  • Has absolute and relative paths in Code and Printed
• work_process-data_4tU-seq_fastqs-UMI.md and work_process-data_4tU-seq_fastqs-UMI-dedup_new-experiments-March.md
  • #TODO Rename the scripts
  • #TODO Add variables from below to top initialization, but don't bother refactoring the parallel calls to HEREDOC submissions

Don't make available

Directories

• etc_cleaning
• etc_initial
• etc_QC
• notebook

Files

• work_env-building.md #TODO Include info in here in new notebook and/or yamls assoc. w/Dependencies

General

• #TODO Add top-of-file descriptions to all notebooks that are made available

2023-0215

2023-0215

Make available

Directories

• infiles_gtf-gff3 #TODO Some, not all—determine what to make available and what to not
• outfiles_gtf-gff3 #TODO Some, not all—determine what to make available and what to not
• outfiles_htseq-count #TODO Some, not all—determine what to make available and what to not

Files

• README.md #TODO Needs to be rewritten to explain what is run with what and in what order
• rough-draft_coverage-tracks_timecourse_size-effect.sh
• rough-draft_coverage-tracks.sh
• rough-draft_draw_scatter-plots.R
• rough-draft_evaluate-categories_expression_initial.Rmd
• rough-draft_evaluate-categories_expression.R
• rough-draft_new-approach-to-analyses.R
• rough-draft_plot-distributions_expression.R
• rough-draft_plot-distributions_length.R
• rough-draft_plot-TPM_N-varies-on-SS_LFC-varies-on-TPM.R
• rough-draft_run-analyses_Fig-5B-5C.R
• rough-draft_run-analyses_rlog-PCA_write-rds.R
• rough-draft_write-gtf-blacklist.R #MAYBE
• run_chi-sq_quantile-filtered-coding-assignments.R #MAYBE Need to check on this—associated with data-for-chi-sq.xlsx below?
• work_assess-process_R64-1-1-gff3_categorize-Trinity-transfrags_part-0.Rmd #COMBINE?
• work_assess-process_R64-1-1-gff3_categorize-Trinity-transfrags_part-1.Rmd #COMBINE?
• work_assess-process_R64-1-1-gff3_categorize-Trinity-transfrags_part-2_legend.txt #COMBINE?
• work_assess-process_R64-1-1-gff3_categorize-Trinity-transfrags_part-2.R #COMBINE?
• work_assess-process_R64-1-1-gff3_categorize-Trinity-transfrags_part-3.R #COMBINE?
• work_assessment-processing_gtfs_part-0_Trinity-etc.md #COMBINE?
• work_assessment-processing_gtfs_part-0.5_non-Trinity.Rmd #COMBINE?
• work_assessment-processing_gtfs_part-1_Trinity.Rmd #COMBINE?
• work_assessment-processing_gtfs_part-1.5_Trinity.R #COMBINE?
• work_assessment-processing_gtfs_part-2_Trinity.md #COMBINE?
• work_calculate_uni-multimappers-etc.md #COMBINE?
• work_combine-gtfs_processed-ncRNA_part-0.Rmd #COMBINE?
• work_combine-gtfs_processed-ncRNA_part-1.md #COMBINE?
• work_combine-gtfs_processed-non-pa-ncRNA_part-0.Rmd #COMBINE?
• work_combine-gtfs_processed-non-pa-ncRNA_part-1.md #COMBINE?
• work_combine-gtfs_processed-pa-ncRNA_part-0.Rmd #COMBINE?
• work_combine-gtfs_processed-pa-ncRNA_part-1.md #COMBINE?
• work_count-features_assessed-processed-R64-1-1-gff3s.md #MAYBE
• work_gff3_include-20S.md
• work_make-blacklist-etc.py #TODO This needs to be rewritten for just the NotFeature file—can remain in Python, or may switch to R (whatever is faster)
• work_prepare-data_GEO_matrices.R
• work_prepare-data_GEO.md
• work_preprocess-prepare_htseq-counts-matrices_gtf-gff3_etc.Rmd #MAYBE Create a directory for initial work and just keep it there w/o making it public? Not sure...
• work_representative-non-coding-transcriptome_part-0.md #COMBINE?
• work_representative-non-coding-transcriptome_part-1.md #COMBINE?
• work_representative-non-coding-transcriptome_part-2.Rmd #COMBINE?
• work_representative-non-coding-transcriptome_part-3.md #COMBINE?
• work_representative-non-coding-transcriptome_part-4.Rmd #COMBINE?
• work_representative-non-coding-transcriptome_part-5.md #COMBINE?

Don't make available

Directories

• bws
• GEO
• notebook

Files

• data_timecourse_counts-raw.tsv #MAYBE Delete?
• data_timecourse_counts-rlog.tsv #TODO Delete
• data-for-chi-sq.xlsx #TODO Delete
• rough-draft_estimate-RNA-degredation.R
• rough-draft_evaluate-categories_expression_scraps_initial.Rmd #MAYBE Delete?
• rough-draft_new-approach-to-analyses_tests-scraps.R #TODO Delete
• rough-draft_plot-TPM_N-varies-on-SS.scraps.R #TODO Delete
• rough-draft_run-analyses_rlog-PCA_write-rds.notes-2.txt #TODO Delete
• rough-draft_run-analyses_rlog-PCA_write-rds.notes.txt #TODO Delete
• rough-draft_run-analyses_rlog-PCA_write-rds.scraps.R #TODO Delete
• tutorial_collapse-intersecting-regions.R
• tutorial_extract-non-overlapping-regions.R
• work_env-building.md #TODO Include info in here in new notebook and/or yamls assoc. w/Dependencies
• test_count_features.md #MAYBE Delete it? Or maybe create a directory for initial work and just keep it there w/o making it public?
• work_count_features_featureCounts.md #MAYBE Delete it? Or create a directory for initial work and just keep it there w/o making it public?
• work_count_features_htseq-count.md #TODO Create a directory for initial work and just keep it there w/o making it public? (This is the work in prep for AG's FHCC seminar.)
• work_evaluation-etc_rough-draft_Rrp6-WT_SS_timecourse_groupwise.Rmd #TODO Create a directory for initial work and just keep it there w/o making it public? (This is the work in prep for AG's FHCC seminar.)
• work_evaluation-etc_variables_pairwise-groupwise.Rmd #TODO Delete
• work_evaluation-etc_variables_pairwise-groupwise.tmp-gw.R #TODO Delete
• work_evaluation-etc_variables_pairwise-groupwise.tmp-pw.R #TODO Delete
• work_evaluation-etc_variables_pairwise-groupwise.TODOs-scraps-etc.txt
• work_examine-snRNA-snoRNA-annotations_part-1.Rmd #TODO Create a directory for initial work and just keep it there w/o making it public? #ORKEEP?
• work_examine-snRNA-snoRNA-annotations_part-2.md #TODO Create a directory for initial work and just keep it there w/o making it public? #ORKEEP?
• work_gff3_convert-strand-designations.Rmd #TODO Create a directory for initial work and just keep it there w/o making it public?
• work_model-variables.md #NOTE Don't need to actually make this file available, but #TODO should include the information contained in this file in either the main or a sub README.md
• work_normalization-etc_rough-draft_NNS_vary-on-transcription.Rmd #TODO Create a directory for initial work and just keep it there w/o making it public
• work_normalization-etc_rough-draft_OsTIR-NNS_vary-on-strain.Rmd #TODO Create a directory for initial work and just keep it there w/o making it public
• work_normalization-etc_rough-draft_wild-type_vary-on-state_antisense.Rmd #TODO Create a directory for initial work and just keep it there w/o making it public
• work_normalization-etc_rough-draft_wild-type_vary-on-state.Rmd #TODO Create a directory for initial work and just keep it there w/o making it public

Maybe

Directories

Files

• collate_sea-tsv.sh #NOTE AG used this for her bootstrapping thing
• rough-draft_handle-matrices-gtfs.R #NOTE Do we use this for anything in the figures?
• rough-draft_handle-tables_establish-study-design.R #NOTE Do we use this for anything in the figures?
• rough-draft_run-analyses_GO.R #NOTE Not sure if we actually use this
• rough-draft_timecourse-samples_processing_part-1a.R #TODO Need to check on this...
• rough-draft_timecourse-samples_processing_part-1b.R #TODO Need to check on this...
• rough-draft_timecourse-samples_processing_part-1c.R #TODO Need to check on this...
• rough-draft_timecourse-samples_processing_part-2a.R #TODO Need to check on this...

Copyright

Copyright © 2022-2023 Kris Alavattam

Permission is hereby granted, free of charge, to any person obtaining a copy of this software and associated files (the “Software”), to deal in the Software without restriction, including without limitation the rights to use, copy, modify, merge, publish, distribute, sublicense, and/or sell copies of the Software, and to permit persons to whom the Software is furnished to do so, subject to the following conditions:

The above copyright notice and this permission notice shall be included in all copies or substantial portions of the Software.

THE SOFTWARE IS PROVIDED "AS IS", WITHOUT WARRANTY OF ANY KIND, EXPRESS OR IMPLIED, INCLUDING BUT NOT LIMITED TO THE WARRANTIES OF MERCHANTABILITY, FITNESS FOR A PARTICULAR PURPOSE AND NONINFRINGEMENT. IN NO EVENT SHALL THE AUTHORS OR COPYRIGHT HOLDERS BE LIABLE FOR ANY CLAIM, DAMAGES OR OTHER LIABILITY, WHETHER IN AN ACTION OF CONTRACT, TORT OR OTHERWISE, ARISING FROM, OUT OF OR IN CONNECTION WITH THE SOFTWARE OR THE USE OR OTHER DEALINGS IN THE SOFTWARE.