Random Base Editing for Genome Evolution in Saccharomyces cerevisiae

2021 ◽  
Author(s):  
Yingjia Pan ◽  
Siyang Xia ◽  
Chang Dong ◽  
Haojie Pan ◽  
Jin Cai ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Genome Evolution ◽  
Base Editing

scholarly journals Rapid Colorimetric Detection of Genome Evolution in SCRaMbLEd Synthetic Saccharomyces cerevisiae Strains

2020 ◽  
Vol 8 (12) ◽  
pp. 1914
Author(s):  
Elizabeth L. I. Wightman ◽  
Heinrich Kroukamp ◽  
Isak S. Pretorius ◽  
Ian T. Paulsen ◽  
Helena K. M. Nevalainen
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Genome Evolution ◽  
Colorimetric Detection ◽  
Cre Recombinase ◽  
Control Measures ◽  
Genomic Rearrangements ◽  
Industrial Yeast ◽  
Poor Growth ◽  
Yeast Strains ◽  
Industrial Strains

Genome-scale engineering and custom synthetic genomes are reshaping the next generation of industrial yeast strains. The Cre-recombinase-mediated chromosomal rearrangement mechanism of designer synthetic Saccharomyces cerevisiae chromosomes, known as SCRaMbLE, is a powerful tool which allows rapid genome evolution upon command. This system is able to generate millions of novel genomes with potential valuable phenotypes, but the excessive loss of essential genes often results in poor growth or even the death of cells with useful phenotypes. In this study we expanded the versatility of SCRaMbLE to industrial strains, and evaluated different control measures to optimize genomic rearrangement, whilst limiting cell death. To achieve this, we have developed RED (rapid evolution detection), a simple colorimetric plate-assay procedure to rapidly quantify the degree of genomic rearrangements within a post-SCRaMbLE yeast population. RED-enabled semi-synthetic strains were mated with the haploid progeny of industrial yeast strains to produce stress-tolerant heterozygous diploid strains. Analysis of these heterozygous strains with the RED-assay, genome sequencing and custom bioinformatics scripts demonstrated a correlation between RED-assay frequencies and physical genomic rearrangements. Here we show that RED is a fast and effective method to evaluate the optimal SCRaMbLE induction times of different Cre-recombinase expression systems for the development of industrial strains.

scholarly journals Stress tolerance enhancement via SPT15 base editing in Saccharomyces cerevisiae

2021 ◽  
Author(s):  
Yanfang Liu ◽  
Yuping Lin ◽  
Yufeng Guo ◽  
Fengli Wu ◽  
Yuanyuan Zhang ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Stress Tolerance ◽  
Rna Polymerase Ii ◽  
Convex Surface ◽  
Cytidine Deaminase ◽  
Structure Alignment ◽  
Protein Structure Alignment ◽  
Base Editing ◽  
And Performance

Abstract Background Saccharomyces cerevisiae is widely used in traditional brewing and modern fermentation industries to produce biofuels, chemicals and other bioproducts, but challenged by various harsh industrial conditions, such as hyperosmotic, thermal and ethanol stresses. Thus, its stress tolerance enhancement has been attracting broad interests. Recently, CRISPR/Cas-based genome editing technology offers unprecedented tools to explore genetic modifications and performance improvement of S. cerevisiae. Results Here, we presented that the Target-AID (activation-induced cytidine deaminase) base editor of enabling C-to-T substitutions could be harnessed to generate in situ nucleotide changes on the S. cerevisiae genome, thereby introducing protein point mutation in cells. The general transcription factor gene SPT15 was targeted, and total 36 mutants with diversified stress tolerances were obtained. Among them, the 18 tolerant mutants against hyperosmotic, thermal and ethanol stresses showed more than 1.5-fold increases of fermentation capacities. These mutations were mainly enriched at the N-terminal region and the convex surface of the saddle-shaped structure of Spt15. Comparative transcriptome analysis of three most stress-tolerant (A140G, P169A and R238K) and two most stress-sensitive (S118L and L214V) mutants revealed common and distinctive impacted global transcription reprogramming and transcriptional regulatory hubs in response to stresses, and these five amino acid changes had different effects on the interactions of Spt15 with DNA and other proteins in the RNA Polymerase II transcription machinery according to protein structure alignment analysis. Conclusions Taken together, our results demonstrated that the Target-AID base editor provided a powerful tool for targeted in situ mutagenesis in S. cerevisiae and more potential targets of Spt15 residues for enhancing yeast stress tolerance.

Construction of lactic acid-tolerant Saccharomyces cerevisiae by using CRISPR-Cas-mediated genome evolution for efficient d-lactic acid production

2020 ◽  
Vol 104 (21) ◽  
pp. 9147-9158
Author(s):  
Ryosuke Mitsui ◽  
Ryosuke Yamada ◽  
Takuya Matsumoto ◽  
Shizue Yoshihara ◽  
Hayato Tokumoto ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Lactic Acid ◽  
Genome Evolution ◽  
Acid Production ◽  
Lactic Acid Production ◽  
Acid Tolerant

scholarly journals Genome evolution across 1,011 Saccharomyces cerevisiae isolates

Nature ◽  
2018 ◽  
Vol 556 (7701) ◽  
pp. 339-344 ◽  
Author(s):  
Jackson Peter ◽  
Matteo De Chiara ◽  
Anne Friedrich ◽  
Jia-Xing Yue ◽  
David Pflieger ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Genome Evolution

RNAi-Assisted Genome Evolution in Saccharomyces cerevisiae for Complex Phenotype Engineering

2014 ◽  
Vol 4 (3) ◽  
pp. 283-291 ◽  
Author(s):  
Tong Si ◽  
Yunzi Luo ◽  
Zehua Bao ◽  
Huimin Zhao
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Genome Evolution ◽  
Complex Phenotype

Improved Stress Tolerance of Saccharomyces cerevisiae by CRISPR-Cas-Mediated Genome Evolution

2019 ◽  
Vol 189 (3) ◽  
pp. 810-821 ◽  
Author(s):  
Ryosuke Mitsui ◽  
Ryosuke Yamada ◽  
Hiroyasu Ogino
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Stress Tolerance ◽  
Genome Evolution
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