scholarly journals A new strategy for seamless gene editing and marker recycling in Saccharomyces cerevisiae using lethal effect of Cwp1

2018 ◽  
Vol 8 (6) ◽  
Author(s):  
Yuxiao Hu ◽  
Yanrong Jia ◽  
Xiangdong Zhao ◽  
Zihao Yang ◽  
Zhimin Hao ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Editing ◽  
Lethal Effect ◽  
Marker Recycling ◽  
New Strategy

Strain-specific lethal effect of the adenovirus E1a protein on Saccharomyces cerevisiae

1990 ◽  
Vol 170 (2) ◽  
pp. 470-476 ◽  
Author(s):  
Tadashi Wada ◽  
Yasuhisa Nogi ◽  
Hiroshi Handa ◽  
Toshio Fukasawa
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Lethal Effect ◽  
Adenovirus E1a ◽  
E1a Protein

scholarly journals Aberrant protein phosphorylation at tyrosine is responsible for the growth-inhibitory action of pp60v-src expressed in the yeast Saccharomyces cerevisiae.

1994 ◽  
Vol 5 (3) ◽  
pp. 283-296 ◽  
Author(s):  
M Florio ◽  
L K Wilson ◽  
J B Trager ◽  
J Thorner ◽  
G S Martin
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Growth Inhibition ◽  
Protein Phosphorylation ◽  
Kinase Activity ◽  
Catalytic Domain ◽  
Lethal Effect ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Growth Inhibitory ◽  
Type V

Expression of pp60v-src, the transforming protein of Rous sarcoma virus, arrests the growth of the yeast Saccharomyces cerevisiae. To determine the basis of this growth arrest, yeast strains were constructed that expressed either wild-type v-src or various mutant v-src genes under the control of the galactose-inducible, glucose repressible GAL1 promoter. When shifted to galactose medium, cells expressing wild-type v-src ceased growth immediately and lost viability, whereas cells expressing a catalytically inactive mutant (K295M) continued to grow normally, indicating that the kinase activity of pp60v-src is required for its growth inhibitory effect. Mutants of v-src altered in the SH2/SH3 domain (XD4, XD6, SPX1, and SHX13) and a mutant lacking a functional N-terminal myristoylation signal (MM4) caused only a partial inhibition of growth, indicating that complete growth inhibition requires either targeting of the active kinase or binding of the kinase to phosphorylated substrates, or both. Cells arrested by v-src expression displayed aberrant microtubule structures, alterations in DNA content and elevated p34CDC28 kinase activity. Immunoblotting with antiphosphotyrosine antibody showed that many yeast proteins, including the p34CDC28 kinase, became phosphorylated at tyrosine in cells expressing v-src. Both the growth inhibition and the tyrosine-specific protein phosphorylation observed following v-src expression were reversed by co-expression of a mammalian phosphotyrosine-specific phosphoprotein phosphatase (PTP1B). However a v-src mutant with a small insertion in the catalytic domain (SRX5) had the same lethal effect as wild-type v-src, yet induced only very low levels of protein-tyrosine phosphorylation. These results indicate that inappropriate phosphorylation at tyrosine is the primary cause of the lethal effect of pp60v-src expression but suggest that only a limited subset of the phosphorylated proteins are involved in this effect.

scholarly journals Lethal effect of the expression of a killer gene SMK1 in Saccharomyces cerevisiae

2000 ◽  
Vol 13 (2) ◽  
pp. 73-76 ◽  
Author(s):  
C. Suzuki ◽  
M. Kawano ◽  
T. Kashiwagi ◽  
Y. Arata ◽  
T. Kawasumi ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Lethal Effect ◽  
Killer Gene

scholarly journals Construction of CRISPR/CAS9 system for industrial Saccharomyces cerevisiae strain and genetic manipulation effect on 2-phenylethanol pathway

2021 ◽  
Author(s):  
Zhiwei Xu ◽  
Zhe Chen ◽  
Lucheng Lin ◽  
Kun Wang ◽  
Jie Sun ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Editing ◽  
High Efficiency ◽  
Genetic Manipulation ◽  
Market Demand ◽  
Dna Homology ◽  
Ehrlich Pathway ◽  
Key Factor ◽  
Industrial Strains ◽  
Manipulation Strategy

Abstract Background The market demand for natural 2-phenylethanol (2-PE) continues to increase. Saccharomyces cerevisiae can synthesize 2-PE through the Ehrlich pathway. There are few studies on the improvement of the diploid industrial strains of S. cerevisiae by gene editing technology. There is no report on the comparison of genetic manipulation effect among S.cerevisiae strains with different 2-PE yield background, and the study on knockout of 2-PE downstream product synthesis gene and its effect on the yield of 2-PE have not been found. Results The CRISPR/CAS9 system with high efficiency for diploid S.cerevisiae CWY132 strain for industrial production of 2-PE was constructed. When the length of the homology arm of donor DNA is increased from 60bp to 500bp, the efficiency of gene editing increased from 0–100%. Using CRISPR/CAS9 technology, the branched acetaldehyde dehydrogenase genes ALD2 and ALD3 and the terminal acetyltransferase gene ATF1 in the Ehrlich pathway of S.cerevisiae strains with different 2-PE yields were knocked out. The results showed that in the high-yielding CWY-132 strain, the 2-PE yield decreased from 3.50 g/L to 1.65 g/L when double ALD2 and ALD3 were knocked out, a decrease of 52.8%. When ATF1 was knocked out, the yield of 2-PE decreased to 0.83 g/L, a decrease of 76.2%; In the low-yielding strain PK-2C, the yield of 2-PE increased from 0.21 g/L to 1.20 g/L when ALD2 was knocked out, an increase of 471%. When ATF1 was knocked out, the yield of 2-PE increased to 0.45g/L, an increase of 114%. The results show that the same genetic manipulation strategy for strains with different 2-PE yeilds backgrounds produces significantly different or even opposite effects. In addition, we found that the insufficient supply of NADH in cells can significantly affect the production of 2-PE, and the tolerance of cells to 2-PE is also a key factor that limits the further increase of 2-PE production in high-yielding strain. Conclusions This study shows that the length of the Donor DNA homology arm is a key factor affecting the efficiency of CRISPR/CAS9 gene editing in industrial diploid S. cerevisiae strains. Our result also shows that it is not feasible to increase the 2-PE production in high-yielding strains by blocking the branch pathway in the Ehrlich pathway. Breakthrough in the 2-PE yield of the high-yielding strains requires improved strains’ tolerance to 2-PE and increase the cellular NADH level.

scholarly journals Applications of CRISPR/Cas Technology to Research the Synthetic Genomics of Yeast

2021 ◽  
Author(s):  
Huafeng Lin ◽  
Haizhen Wang ◽  
Aimin Deng ◽  
Minjing Rong ◽  
Lei Ye ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Synthetic Biology ◽  
Gene Editing ◽  
Functional Genes ◽  
Whole Genome ◽  
Yeast Saccharomyces Cerevisiae ◽  
Synthetic Genome ◽  
The World ◽  
Synthetic Genomics ◽  
Genome Projects

The whole genome projects open the prelude to the diversity and complexity of biological genome by generating immense data. For the sake of exploring the riddle of the genome, scientists around the world have dedicated themselves in annotating for these massive data. However, searching for the exact and valuable information is like looking for a needle in a haystack. Advances in gene editing technology have allowed researchers to precisely manipulate the targeted functional genes in the genome by the state-of-the-art gene-editing tools, so as to facilitate the studies involving the fields of biology, agriculture, food industry, medicine, environment and healthcare in a more convenient way. As a sort of pioneer editing devices, the CRISPR/Cas systems having various versatile homologs and variants, now are rapidly giving impetus to the development of synthetic genomics and synthetic biology. Firstly, in the chapter, we will present the classification, structural and functional diversity of CRISPR/Cas systems. Then we will emphasize the applications in synthetic genome of yeast (Saccharomyces cerevisiae) using CRISPR/Cas technology based on year order. Finally, the summary and prospection of synthetic genomics as well as synthetic biotechnology based on CRISPR/Cas systems and their further utilizations in yeast are narrated.

scholarly journals Enhanced scale and scope of genome engineering and regulation using CRISPR/Cas in Saccharomyces cerevisiae

2019 ◽  
Vol 19 (7) ◽  
Author(s):  
Matthew Deaner ◽  
Hal S Alper
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Regulation ◽  
Transcriptional Regulation ◽  
Gene Editing ◽  
Genome Engineering ◽  
Guide Rna ◽  
On Demand ◽  
Large Numbers ◽  
Genetic Modifications ◽  
Genome Scale

ABSTRACT Although only 6 years old, the CRISPR system has blossomed into a tool for rapid, on-demand genome engineering and gene regulation in Saccharomyces cerevisiae. In this minireview, we discuss fundamental CRISPR technologies, tools to improve the efficiency and capabilities of gene targeting, and cutting-edge techniques to explore gene editing and transcriptional regulation at genome scale using pooled approaches. The focus is on applications to metabolic engineering with topics including development of techniques to edit the genome in multiplex, tools to enable large numbers of genetic modifications using pooled single-guide RNA libraries and efforts to enable programmable transcriptional regulation using endonuclease-null Cas enzymes.

scholarly journals Anaerobic growth of Saccharomyces cerevisiae alleviates the lethal effect of phosphotyrosyl phosphatase activators depletion.

2001 ◽  
Vol 48 (4) ◽  
pp. 1043-1049
Author(s):  
B Rempola ◽  
A Kaniak ◽  
J P Di Rago ◽  
J Rytka
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitochondrial Genome ◽  
Lethal Effect ◽  
Heme Biosynthesis ◽  
Anaerobic Growth ◽  
Double Mutation ◽  
Synthetic Lethal ◽  
Per Se

Saccharomyces cerevisiae homologues of phosphotyrosyl phosphatase activator (PTPA) are encoded byRRD1 and RRD2, genes whose combined deletion is synthetic lethal. Previously we have shown that the lethality of rrd1,2delta can be suppressed by increasing the osmolarity of the medium. Here we show that the lethality of rrd1,2delta is also suppressed under oxygen-limited conditions. The absence of respiration per se is not responsible for the suppression since elimination of the mitochondrial genome or a block in heme biosynthesis fail to rescue the rrd1,2delta double mutation.

scholarly journals Cas9-mediated gene editing in Saccharomyces cerevisiae

2017 ◽  
Author(s):  
Ranjith Anand ◽  
Gonen Memisoglu ◽  
James Haber
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Editing
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