Saccharomyces cerevisiae Genome Shuffling through Recursive Population Mating Leads to Improved Tolerance to Spent Sulfite Liquor

Pinel, Dominic; D'Aoust, Frédéric; Cardayré, Stephen B. del; Bajwa, Paramjit K.; Lee, Hung; Martin, Vincent J. J.

doi:10.1128/aem.02769-10

Cited by 51 publications

(41 citation statements)

References 42 publications

(54 reference statements)

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“…We estimated population SNP allele frequencies, identified genomic regions with genetic variation responsible for the phenotypic variation, and determined the population-favorable QTL allele at those regions using a combination of published analysis methods 16,31,35 (see also Methods). In total, we identified 17 QTLs (Figure 2A, Table 1 and Table S1), confirming that hydrolysate tolerance is a complex genetic trait 12,36,37 . We observed favorable QTL alleles of both parental ancestries (eight JAY291 and nine S288C alleles) (Figure 2A and Table 1).…”

Section: High-resolution Qtl Mapping Of Hydrolysate Tolerance Identifsupporting

confidence: 54%

QTL-guided metabolic engineering of a complex trait

Maurer

Sutardja

Pinel

et al. 2016

Preprint

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Engineering complex phenotypes for industrial and synthetic biology applications is difficult and often confounds rational design. Bioethanol production from lignocellulosic feedstocks is a complex trait that requires multiple host systems to utilize, detoxify, and metabolize a mixture of sugars and inhibitors present in plant hydrolysates. Here, we demonstrate an integrated approach to discovering and optimizing host factors that impact fitness of Saccharomyces cerevisiae during fermentation of a Miscanthus x giganteus plant hydrolysate. We first used high-resolution Quantitative Trait Loci (QTL) mapping and systematic Bulk Reciprocal Hemizygosity analysis (bRHA) to discover 17 loci that differentiate hydrolysate tolerance between an industrially related (JAY291) and a laboratory (S288C) strain. We then used this data to identify a subset of favorable allelic loci that were most amenable for strain engineering. Guided by this "genetic blueprint", and using a dual-guide Cas9-based method to efficiently perform multi-kilobase locus replacements, we engineered an S288C strain with superior hydrolysate tolerance than JAY291. Our methods should be generalizable to engineering any complex trait in S. cerevisiae, as well as other organisms.

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Section: High-resolution Qtl Mapping Of Hydrolysate Tolerance Identifsupporting

confidence: 54%

QTL-guided metabolic engineering of a complex trait

Maurer

Sutardja

Pinel

et al. 2016

Preprint

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“…The conditions of the inactivation processes reported in previous studies varied significantly (Ge et al ., ; Pinel et al ., ; Bajwa et al ., ; Hou, ). In this study, we observed that the harsh conditions of inactivation process led to significant decreases in survival rate, making the cells more difficult to resuscitate.…”

Section: Resultsmentioning

confidence: 99%

Enhanced deacidification activity in Schizosaccharomyces pombe by genome shuffling

Ding

Zhang

et al. 2014

Yeast

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A problem frequently occurring in making some kinds of wines, particularly Vitis quinquangularis Rehd wine, is the presence of malic acid at high concentrations, which is detrimental to the quality of wines. Thus, there is a need of the ways for effectively reducing the malic acid levels in wine. This study aimed to generate shuffled fusants of Schizosaccharomyces pombe with enhanced deacidification activity for reducing the excessive malic acid content in wine. Sz. pombe CGMCC 2.1628 was used as the original strain. The starting mutant population was generated by UV treatment. The mutants with higher deacidification activity were selected and subjected to recursive protoplast fusion. The resulting fusants were screened by using the indicator of malic acid concentration of fermentation supernatants on 96-well microtitre plates, measured with bromocresol green. After three rounds of genome shuffling, the best-performing fusant, named GS3-1, was obtained. Its deacidification activity (consumed 4.78 g/l malic acid within 10 days) was increased by 225.2% as compared to that of original strain. In the Vitis quinquangularis Rehd wine fermentation test, GS3-1 consumed 4.0 g/l malic acid during the whole cycle of fermentation, providing up to 185.7% improvement in malic acid consumption compared with that of the original strain. This study shows that GS3-1 has great potential for improving the quality of Vitis quinquangularis Rehd wine.

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“…1A; Pinel et al 2011;Suzuki et al 2011Suzuki et al , 2012. With a green fluorescent protein (GFP) reporter gene inserted in each deletion locus, the enrichment of higher-order yeast deletion strains in the meiotic population can be accomplished using flow cytometry.…”

Section: ó 2015 Suzuki Et Al This Article Is Distributed Exclusivelymentioning

confidence: 99%

Bacterial genome reduction using the progressive clustering of deletions via yeast sexual cycling

et al. 2015

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The availability of genetically tractable organisms with simple genomes is critical for the rapid, systems-level understanding of basic biological processes. Mycoplasma bacteria, with the smallest known genomes among free-living cellular organisms, are ideal models for this purpose, but the natural versions of these cells have genome complexities still too great to offer a comprehensive view of a fundamental life form. Here we describe an efficient method for reducing genomes from these organisms by identifying individually deletable regions using transposon mutagenesis and progressively clustering deleted genomic segments using meiotic recombination between the bacterial genomes harbored in yeast. Mycoplasmal genomes subjected to this process and transplanted into recipient cells yielded two mycoplasma strains. The first simultaneously lacked eight singly deletable regions of the genome, representing a total of 91 genes and~10% of the original genome. The second strain lacked seven of the eight regions, representing 84 genes. Growth assay data revealed an absence of genetic interactions among the 91 genes under tested conditions. Despite predicted effects of the deletions on sugar metabolism and the proteome, growth rates were unaffected by the gene deletions in the seven-deletion strain. These results support the feasibility of using single-gene disruption data to design and construct viable genomes lacking multiple genes, paving the way toward genome minimization. The progressive clustering method is expected to be effective for the reorganization of any mega-sized DNA molecules cloned in yeast, facilitating the construction of designer genomes in microbes as well as genomic fragments for genetic engineering of higher eukaryotes.

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Saccharomyces cerevisiae Genome Shuffling through Recursive Population Mating Leads to Improved Tolerance to Spent Sulfite Liquor

Cited by 51 publications

References 42 publications

QTL-guided metabolic engineering of a complex trait

QTL-guided metabolic engineering of a complex trait

Enhanced deacidification activity in Schizosaccharomyces pombe by genome shuffling

Bacterial genome reduction using the progressive clustering of deletions via yeast sexual cycling

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