Long-read genomes reveal pangenomic variation underlying yeast phenotypic diversity

Abstract

Understanding the genetic causes of trait variation is a primary goal of genetic research. One way that individuals can vary genetically is through the existence of variable pangenomic genes - genes that are only present in some individuals in a population. The presence or absence of entire genes could have large effects on trait variation. However, variable pangenomic genes can be missed in standard genotyping workflows, due to reliance on aligning short-read sequencing to reference genomes. A popular method for studying the genetic basis of trait variation is linkage mapping, which identifies quantitative trait loci (QTLs), regions of the genome that harbor causative genetic variants. Large-scale linkage mapping in the budding yeast Saccharomyces cerevisiae has found thousands of QTLs affecting myriad yeast phenotypes. To enable the resolution of QTLs caused by variable pangenomic genes, we used long-read sequencing to generate highly complete de novo assemblies of 16 diverse yeast isolates. With these assemblies we resolved growth QTLs to specific genes that are absent from the reference genome but present in the broader yeast population at appreciable frequency. Copies of genes also recombine onto chromosomes where they are absent in the reference genome, and we found that these copies generate additional QTLs whose resolution requires pangenome characterization. Our findings demonstrate the power of long-read sequencing to identify the genetic basis of trait variation.

Comprehensive deletion scan of anti-CRISPR AcrIIA4 reveals essential and dispensable domains for Cas9 inhibition

“…We transformed the repair template library into the acrIIA4-URA3-acrIIA4 yeast strain. The strain also carried two copies of SpCas9, one under a MAL62 promoter and the other under a GALL promoter, allowing SpCas9 to be activated by either maltose or galactose (16)(17)(18). Both promoters are repressed by glucose.…”

“…We named the resulting plasmid MSp500. The sequence of the MAL genes was taken from the genome sequence of YJM145 (MSY28) (18). Amplification of the MAL locus sequence was done with KAPA Hi-Fi Hotstart DNA Polymerase (and primers ANx16.5 and 17.5), and inserted into MSp14 via Gibson assembly (New England Biolabs, Ipswich, MA, USA) following a digestion of the plasmid with SpeI-HF (New England Biolabs).…”

Discovery of a rapidly evolving yeast defense factor, KTD1 , against the secreted killer toxin K28

“…The generation and analysis of these genome assemblies are described in more detail in Weller et al. ( 62 ).…”

Cluster efficient pangenome graph construction with nf-core/pangenome

“…The availability of high-quality collections of population-wide wholegenome assemblies (Liao et al, 2023;Kang et al, 2023;Weller et al, 2023;Zhou et al, 2022;Liu et al, 2020;Leonard et al, 2022) offers new opportunities to study sequence evolution and variation within and between genomic populations. A challenge is simultaneously representing and analyzing hundreds to thousands of genomes at a gigabase scale.…”