Dataset DOI: 10.5061/dryad.34tmpg4x4

Description of the data and file structure

In this study, we investigate genetic and regulatory architecture associated with late-life survival in a genetically diverse yeast population. Using a GWAS-inspired experimental framework, we combine pooled-population genomic and transcriptome sequencing of paired aged and young cohorts to identify variants and expression patterns linked to age-associated phenotypes.

Files and variables

File: Dryad_datasets.zip

Description: This zip file contains the raw and filtered SNP tables and raw and normalized gene count matrices. Additionally, it contains complete results data and significant result tables. This folder contains three subdirectories: raw_data , clean_data , complete_results, and significant_results.

Subdirectory: raw_data:

1) filtered_snps.txt contains the "raw" SNP table that was generated using the Burke Lab pipeline. Columns are described below: 

• Nmiss - number of missing values
• CHROM - chromosome number
• POS - SNP position on the chromosome
• REF - the reference allele at that genomic position
• ALT - the alternate allele at that genomic position
• For each of the twelve paired replicates (01-12) there are four additional columns: 
  • alt_old_XX - Gives the count of the alternate allele for old replicate #XX at each genomic position
  • alt_young_XX - Give the count of the alternate allele for young replicate #XX at each genomic position
  • N_old_XX - Gives total coverage for old replicate #XX at each genomic position
  • N_young_XX - Gives total coverage for young replicate #XX at each genomic position

2) filtered_indels.txt contains the "raw" indel table that was generated using the Burke Lab pipeline. Follows the same naming conventions as filtered_snps.txt.

3) annotated_snps.txt contains the SNPeff annotations for the generated SNP table. Columns are as follows: 

• CHROM - chromosome number
• POS - SNP position on the chromosome
• REF - the reference allele at that genomic position
• ALT - the alternate allele at that genomic position
• HOM.VAR - count of homozygous variant genotypes
• HET - count of heterozygous genotypes
• HOM.REF - count of homozygous reference genotypes
• NO.CALL - count of no-call genotypes
• EVENTLENGTH - length of event
• TYPE - variant type (SNP/indel)
• MULTI.ALLELIC - true or false
• ANN - SNPeff annotation (described in SNPeff documentation)
• For each replicate, there are also two additional columns for each old and young fraction
  • {age}_XX.GT - genotypes
  • {age}_XX.AD - allele depth

4) annotated_indels.txt contains the SNPeff annotations for the generated indel table, and follows the same conventions as annotated_snps.txt.

5) founder_states.txt contains the haplotype frequencies for the data. Columns are described below: 

• CHROM - chromosome number
• Pos - position
• REF - reference allele
• ALT - alternate allele
• The next four columns are binary data for each haplotype. A 1 indicates that haplotype has the reference allele at this position, a 0 indicates it has the alternate allele at this position.
  • A1 - data for SGRP strain DBVPG6765
  • A2 - data for SGRP strain DBVPG6044
  • B3 - data for SGRP strain YPS128
  • B4 - data for SGRP strain Y12
• anc - frequency of the reference allele in the ancestor population at each position

6) gene_count_matrix.txt contains the gene count matrix from the transcriptomic sequencing, generated using FeatureCounts.

• The first several columns are general output from FeatureCounts:
  • Geneid - Gives the gene ID from the GTF file
  • Chr - chromosome number
  • Start - start position of gene
  • End - end position of gene
  • Strand - orientation of the transcript
  • Length - length of feature
• For each of the twelve paired replicates, there are two additional columns: 
  • RNAseq_OLD_repXX.bam - transcript counts for the old replicate #XX
  • RNAseq_YOUNG_repXX.bam - transcript counts for the young replicate #XX

7) all_founders.txt contains whole genome haplotype data for all founders (including some not used in this study).

• The first five columns follow the same convention as filtered_snps.txt
• The remaining columns are count data for alternate alleles (alt) and total coverage (N) for different haplotype strains. For this study, the columns of interest are for haplotypes A1 (DBVPG6765), A2 (DBVPG6044), B3 (YPS128), and B4 (Y12): 
  • alt_YEE_hap_A1_00
  • N_YEE_hap_A1_00
  • alt_YEE_hap_A2_00
  • N_YEE_hap_A2_00
  • alt_YEE_hap_B3_00
  • N_YEE_hap_B3_00
  • alt_YEE_hap_B4_00
  • N_YEE_hap_B4_00

8) budscar_counts (folder) contains the raw bud scar count data for each replicate. Each file corresponds to a single replicate, with two tabs (old and young). The columns for each file have the following convention: 

• Image - image ID that the cell was counted from
• Cell - count of identified cell from that image
• Bud scars - number of bud scars identified on cell
• buds - number of buds identified on that cell
• dead - Number of cells (including buds) that appeared to be dead

Subdirectory: clean_data

1) scarsdata.txt contains the compiled bud scar count data. Columns are as follows: 

• replicate - replicate number (matching files in the budscar_counts folder in the raw_data subdirectory. Strains are as follows: 1= 4S, 2=YPS128, 3=Y12, 4=DBVPG6044, 5=DBVPG6765).
• date - date of sort
• strain - Strain name (4x should read 4S)
• age - sorted fraction (aged or young)
• images_counted - the number of images that were counted for that fraction
• cells_nobuds - the total number of cells (excluding buds that were still attached to mother cell)
• cells_budsincluded - the total number of cells (including buds that were still attached to mother cell)
• max_scars - the maximum number of bud scars found on a single cell in that fraction
• avg_scars_mothers - the average number of bud scars found on a single cell in that fraction, excluding buds
• avg_scars_all - the average number of bud scars found on a cell in that fraction, including buds as individual cells
• dead - the number of dead cells found in that fraction
• nobuds_living - the number of live cells found in that fraction (excluding attached buds).

2) GWAS_SNPS_cov20_maf05.csv contains the SNP table after filtering for a coverage of 20 and a minor allele frequency of 0.05. This table uses the same column structure as filtered_snps.txt

3) indels_cov20_maf05.csv contains the indel table after filtering for a coverage of 20 and a minor allele frequency of 0.05. This table uses the same column structure as filtered_snps.txt

4) normalized_counts_deseq.csv contains the gene count matrix, normalized for visualization. This table uses the same column structure as gene_count_matrix.txt.

Subdirectory: complete_results

1) WG_CMHtest_results.csv contains results of the genomic CMH test for all detected SNPs, This table uses the same column structure as gene_count_matrix.txt, plus the following additional columns: 

• mincov - the minimum coverage found for any sample at that genomic position
• maxcov - the maximum coverage for any sample at that genomic position
• sum_mac - the sum of minor allele counts for all samples at that position
• sum_cov - the sum of coverage for all samples at that position
• maf- minor allele frequency at that position
• logp -log10 transformed p-values from CMH test
• untransformed p-values for CMH test

2) indels_CMHtest_results.csv contains results of the genomic CMH test for all detected indels. The columns follow the same convention as WG_CMHtest_results.csv.

3) rnaseq_results_batch_adjusted.csv contains the results of the DGE analysis in DESeq2 (adjusted for the batch effect, treating "young" as the reference condition). The format follows a standard DESeq2 result file, including the Gene_ID as the first column (unlabelled), baseMean expression, log2FoldChange, lfcsE (standard error estimate of log2fold change), stat (test statistic value), pvalue, and padj.

Subdirectory: significant_results

1) SuppTable3_GenomicVariantAnn.csv contains all significant SNPs and indels with a cutoff of p<0.05. The columns are as follows: 

• CHROM - chromosome number
• POS - genomic position
• TYPE - indel or SNP
• pval - P-value from CMH test
• logp -log10 transformed p-value
• Gene_Name - Gene name
• Annotation - annotation assignment taken from SNPeff

2) RNA_genes_p<0.1.csv contains all DEGs with a significance cutoff of 0.1 (with young set as reference condition).

• Convention shared with rnaseq_results_batch_adjusted.csv, except the Gene_Name assigned by Saccharomyces Genome Database has been appended as the last column. 

3) RNA_genes_p<0.05.csv contains all DEGs with a significance cutoff of 0.05 (with young set as reference condition). Columns shared with RNA_genes_p<0.1.csv.

Code/software

R and R studio were used for analysis unless otherwise noted. All libraries and packages are listed within the scripts they are used. The order in which scripts should be run is indicated by the numbering scheme (e.g. any script beginning 00_ should be run first, 01_ second, etc). More information about each script can be found in nested README files.This zip file contains the data and code used for analysis and figure generation within the manuscript, including more in-depth README files. This .zip serves as a backup for the data and code found on Github at: https://github.com/Katie-McHugh/regulation_of_aging/tree/Dryad_v3.

Fiji (is just ImageJ) was used to open image files and count bud scars.