YEAST DISPLAY SCREENING FOR C05 VARIANTS

This README describes the scripts used for the analyses in:
In vitro evolution of an influenza broadly neutralizing antibody is modulated by hemagglutinin receptor specificity

FILES

• All sequencing raw reads, which can be downloaded from NIH SRA database PRJNA326694, should placed in fastq/ folder:
  • Input library: Wu-1_S1_L001_R1_001.fastq and Wu-1_S1_L001_R2_001.fastq
  • H1 Round 1 library: Wu-1_S2_L001_R1_001.fastq and Wu-1_S2_L001_R2_001.fastq
  • H1 Round 2 library: Wu-1_S3_L001_R1_001.fastq and Wu-1_S3_L001_R2_001.fastq
  • H1 Round 3 library: Wu-1_S4_L001_R1_001.fastq and Wu-1_S4_L001_R2_001.fastq
  • H3 Round 1 library: Wu-1_S5_L001_R1_001.fastq and Wu-1_S5_L001_R2_001.fastq
  • H3 Round 2 library: Wu-1_S6_L001_R1_001.fastq and Wu-1_S6_L001_R2_001.fastq
  • H3 Round 3 library: Wu-1_S7_L001_R1_001.fastq and Wu-1_S7_L001_R2_001.fastq
  • H5 Round 1 library: Wu-1_S9_L001_R1_001.fastq and Wu-1_S9_L001_R2_001.fastq
  • H5 Round 2 library: Wu-1_S10_L001_R1_001.fastq and Wu-1_S10_L001_R2_001.fastq
  • H5 Round 3 library: Wu-1_S11_L001_R1_001.fastq and Wu-1_S11_L001_R2_001.fastq
• data/*count: Counting for each variant in each library
• data/VariantFreqTable.tsv: Frequency of each variant in each library. Units = per million.
• data/AAFreqTable.tsv: Frequency of each amino acid at each residue in each library.

ANALYSIS PIPELINE

• script/C05_Read2Count.py
  • Input file: fastq/Wu-1_S*_L001_R1_001.fastq
  • Output file: data/*.count
• script/C05_Count2VarFreq.py
  • Input file: data/*.count
  • Output file: data/VariantFreqTable.tsv
• script/C05_VarFreq2AAFreq.py
  • Input file: data/VariantFreqTable.tsv
  • Output file: data/AAFreqTable.tsv

PLOTTING

• script/C05_plot_aaheatmap.R: Plot the frequency of each amino acid at each residue in each library as heatmaps
  • Input file: data/AAFreqTable.tsv
  • Output file: graph/HM*.png
• script/C05_plot_top10varfreq.R: Plot the frequency of 10 top variants with the highest frequency in round 3 against each HA
  • Input file: data/AAFreqTable.tsv
  • Output file: graph/H?Top10Freq.png
• script/C05_plot_WTfrac.R: Plot the frequency of WT in each library
  • Input file: data/AAFreqTable.tsv
  • Output file: graph/WTFrac.png
• script/C05_plot_H1vsH3.R: Compare the frequency of each variant in H1 round 3 vs H3 round 3
  • Input file: data/AAFreqTable.tsv
  • Output file: graph/H1H3specialists.png