Supplemental Data Information ---------- File S1: TS collection strain and phenotype information --------- ***** strain.information ***** strain: The strain name used in this collection set: The set this strain comes from lab.origin: The original lab that generated the strain original.strain.name: The original name given to this strain allele: The allele assigned by the origin lab ***** set1.phenotype.data ***** ts.strain: The strain name used in this collection bowerman.strain: The original name given to this strain allele: The allele assigned by the Bowerman lab Average Sequencing coverage: The average sequencing coverage generated by Waterston Lab TS.status confirmed: Whether or not the temperature-sensitive status of the strain was confirmed by the Bowerman or Moerman lab Defect: The general defect observed Phenotype Notes: Description of any phenotypes observed during analysis or handling ***** set2.phenotype.data ***** ts.strain: The strain name used in this collection schnabel.strain: The original name given to this strain allele: The allele assigned by the Schnabel lab phenotype.summary: A summary of the adjacent columns to indicate which categories were observed during strain phenotyping. A description of categories is included in "set2.phenotype.categories" ---------- File S2: TS collection sequencing variants identified --------- A series of headerless xlsx file containing all of the single-nucleotide variants identified in our MMP pipeline. There are 10 spreadsheets divided into the following categories: 1.all.strain.SNV: a listing of all the SNVs identified for all 173 TS strains, and the parental strain from set 1 (VC50151) 2.combined.unique.SNV: a listing of all the unique SNVs identified in analysis from the combined 173 TS strains, and the parental strain from set 1 (VC50151) 3.set1.unique.SNV: a listing of all the unique SNVs identified in analysis of only the 72 TS strains from set 1 and its parental strain (VC50151) 4.set2.unique.SNV: a listing of all the unique SNVs identified in analysis of only the 67 TS strains from set 2 5.set3.unique.SNV: a listing of all the unique SNVs identified in analysis of only the 34 TS strains from set 3 6.all.strain.INDEL: a listing of all the INDELs identified for all 173 TS strains, and the parental strain from set 1 (VC50151) 7.combined.unique.INDEL: a listing of all the unique INDELs identified in analysis from the combined 173 TS strains, and the parental strain from set 1 (VC50151) 8.set1.unique.INDEL: a listing of all the unique INDELs identified in analysis of only the 72 TS strains from set 1 and its parental strain (VC50151) 9.set2.unique.INDEL: a listing of all the unique INDELs identified in analysis of only the 67 TS strains from set 2 10.set3.unique.INDEL: a listing of all the unique INDELs identified in analysis of only the 34 TS strains from set 3 * Note that overlapping unique SNVs/INDELs exist between sets 1, 2, and 3. These are not present on "2.combined.unique.SNV" The following column format applies to sheets 1-5. column 1: strain name column 2: SNV chromosome column 3: SNV chromosomal location column 4: SNV reference allele column 5: SNV strain allele column 6: SNV chromosome column 7: SNV region category 1 (Link, Coding_transcript, curated) column 8: SNV region category 2 (Link[region], Coding_transcript[intron,exon], curated[intron, coding_exon]) column 9: region start position column 10: region end position column 11: "." separator column 12: strand column 13: "." separator column 14: Gene information if applicable column 15-28: Repetition of information IF the given SNV region has multiple classifications The following column format applies to sheets 7-10. column 1: strain name column 2: INDEL chromosome column 3: INDEL chromosomal location column 4: INDEL reference allele column 5: INDEL strain allele ---------- File S3: MIP primer information and VC20019 MIP sequence information --------- ***** MIP primer info ***** Listed are the primer sequences used for the PCR amplication steps during library preparation AND the custom primers spiked in for sequencing on Illumina kits. "MIP Rev Generic PCR Primer" is included for designing additional library barcode primers beyond the 96 provided on this spreadsheet. "MIP Seq Read 2 primer" is included to help readers if they should attempt to design experiments for paired-end sequencing. It is not necessary for single-end sequencing. "MIP Common Backbone sequence" is included to help readers to understand the design of the MIP oligo sequences themselves Sequencing notes: Use "MIP Seq Read 1 primer" and "MIP Seq Index primer" for Illumina platform sequencing, with a working concentration of 100 µM. These primers are spiked into the default wells holding standard Illumina Read 1 and Index primers (respectively). Typically 5 µl each are spiked into Illumina MiSeq kits while 15 µl are spiked into Illumina NextSeq kits. ***** VC20019 MIP Information ***** MIP.name: Name of the MIP oligo in the format of [STRAIN]-MIPS-[chromosome]-[megabase position] snv.chr: Chromosome of target SNV snv.loc: Location of the target SNV based on WS230 coordinates allele: Allele name of SNV target wt.allele: The reference sequence of the allele snv.allele: The mutant sequence of the allele for the STRAIN strand: The strand used to generate the wt.allele and snv.allele information lig.gap: The number of bases between the end of the ligation annealing arm and the SNV target lig.length: The total length of the ligation annealing arm lig.seq: The sequence of the ligation annealing arm as part of the MIP lig.GC: The percent GC content of the ligation annealing arm lig.gap.seq: The sequence information located 3' of the ligation annealing arm and 5' of the SNV target ext.gap: The number of bases between the SNV target and the beginning of the extension annealing arm ext.length: The total length of the extension annealing arm ext.seq: The sequence of the extension annealing arm ext.GC: The percent GC content of the extension annealing arm ext.gap.seq: The sequence information located 5' of the SNV target through to the expected read size of 50 basepairs. A typical single-end read includes a UMI (12 bp) + lig.seq(20 bp) + lig.gap.seq(x bp) + SNV (1 bp) + ext.gap.seq (y bp) lig.seq.read: The nucleotide sequence used to identify reads originating from this MIP wt.gap.fill.read: The expected gap-fill sequence information for a reference allele snv.gap.fill.read: The expected gap-fill sequence information for the strain-specific allele ext.seq.read: The initial nucleotide sequence expected from a the second read in a paired-end run (not usually utilized) gap.size: The overall distance between the 5' and 3' annealing sequences of the MIP PCR.size: The expected size of the linearized PCR product for this MIP (includes adapters, barcode primers etc.) MIP.seq: The 5'-->3' sequence of the MIP to ORDER BLAT.seq: The sequence used for UCSC BLAT of the MIP arms to check for target specificity primer.BLAST: The sequenced used to check NCBI Primer BLAST for target specificity ---------- File S4: VC20019 Normalization parameters --------- List of parameters used in the post-processing normalization of MIP data. The Mode of these parameters was used in the normalization process. Each MIP target has a height (h), mean, offset (off), and variance (sigma) parameter associated with it.