IntegrativeProjectWT2015
This repo contains the experiment that I like to call “IntegrativeProjectWT2015” because it is an integrative analysis of behavior, electrophysiology, and RNA-seq data collected from wild type mice in 2015.
Organization
Three are four main directories for the R scripts, UNIX workflow, figures, and data.
The file names were chosen to indicated their order of execution in the workflow and provide a brief description of the contents.
Workflow
- Overview
- 00_methods: a description of the behavior, electrophysiology, and RNA-seq methods
- Part 1: Behavior
- 01a_behavior_create_dfs: wrangling the behavior data
- 01b_behavior_figures: statistical analysis and behavior data visualization
- Part 2.1: UNIX for RNA-seq
- UNIX workflow Initial processing of RNA sequencing was performed on the Stampede cluster at the Texas Advanced Computing Facility. My workflow for this is described in the ‘UNIXworkflow’ directory.
- ../UNIXworkflow/00_rawdata: Download the data to scratch on Stampede with
00_gsaf_download.sh. - ../UNIXworkflow/01_fastqc: Evaluate the quality of the reads using the program FastQC.
- ../UNIXworkflow/02_filtrimreads: Filter low quality reads and trim adapters using the program cutadapt.
- ../UNIXworkflow/03_fastqc: Evaluate the quality of the processed reads
- ../UNIXworkflow/04_kallisto: Quantify transcript-level expression using Kallisto
- Part 2.2: R for RNA-seq
- 02a_rnaseq_makedfs: converting the Kallisto transcript counts to gene counts and wrangling the categorical data about the samples
- 02b_rnaseqALL: analyzing all the RNA-seq data together
- 02c_rnaseqSubfield: analyzing the data for each hippocampal subfield separately
- 02d_rnaseqAvoidance: combining the two yoked group and the two training groups before analyzing and then analyzing each subfield separately
- 02e_GO_MWU an analysis of gene ontology. Note: this directory contains scripts and data
- Part 3: Electrophysiology
- 03_ephys: analysis of electrophysiology data
- 03_ephys: analysis of electrophysiology data
- Part 4: Integrative analysis
- 04_integration: correlations across levels, mostly using PCA data
- Extras
- 99_colorblindr: a simulation of how some of my figures will look to colorblind readers
- 99_forDeep: a targeted PC analysis of a colleague
Results
Quick view of the overall numbers and patterns of differentially expressed beahvioras and genes in response to beahvioral manipulatlions.
Figure 1: Experimental overview
Figure 2: Cognitive training induces avoidance behavior in conflict and consistently trained animals. See 01b_behavior_figures:Rmd for the corresponding analysis
Figure 3: RNA sequencing confirms large differences in DG, CA3, and CA1 hippocampal subfields
See 02b_rnaseqALL:Rmd for the corresponding analysis
Table 1: Numbers of differentially expressed genes by two-way contrast when all data is included in the normalization and quantificaiton. See 02b_rnaseqALL:Rmd for the corresponding analysis
| contrast | up | down | total |
|---|---|---|---|
| CA3 vs. DG | 1625 | 1361 | 2986 |
| CA1 vs. DG | 968 | 956 | 1924 |
| CA1 vs. CA3 | 651 | 791 | 1442 |
| consistent vs. yoked consistent | 113 | 7 | 120 |
| yoked conflict vs. yoked consistent | 30 | 1 | 31 |
| conflict vs. yoked conflict | 15 | 24 | 39 |
| conflict vs. consistent | 0 | 0 | 0 |
Table 2: Sample sizes for tissue specific analysis of the resopnse to behavior manipulations in DG, CA3, and CA1 hippocampal subfields. See 02c_rnaseqSubfield.Rmd for the corresponding analysis
| Training Group | DG | CA3 | CA1 |
|---|---|---|---|
| consistent yoked | 4 | 3 | 2 |
| consistent trained | 3 | 2 | 4 |
| conflict yoked | 4 | 3 | 5 |
| conflict trained | 5 | 5 | 4 |
Figure 4: Cognitive training alters gene expression in DG and CA1 but not CA3. See 02c_rnaseqSubfield.Rmd for the corresponding analysis
Table 1: Numbers of differentially expressed genes by two-way contrast the data is subset by hippocampal tissue before normalization and quantification. See 02c_rnaseqSubfield.Rmd for the corresponding analysis
| Two-way contrasts between groups | DG | CA3 | CA1 |
|---|---|---|---|
| consistent vs. yoked-consisten | 95 | 0 | 196 |
| conflict vs. consistent | 0 | 1 | 0 |
| yoked-conflict vs. yoked-consistent | 2 | 0 | 201 |
| conflict vs. yoked-conflict | 5 | 0 | 2 |
Figure 5: Training has little effect on electrophysiology. See 003_ephys.Rmd for the corresponding analysis
Figure 6: Correlations across levels of biological organization. See 04_integration.Rmd for the corresponding analysis
Data
This directory contains both raw and intermediate data files. - Intermediate data files have alphanumeric prefixes that correspond to the R script that created them. - Raw files have only numeric prefix (aka lack an alphabetical character) that indicates whether it is for behavior (01), RNA-seq (02), or ephys (03). - Files with more general names were created for public repositories