Ontology highlight
ABSTRACT:
PROVIDER: PRJEB13825 | ENA |
REPOSITORIES: ENA
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Project description:Directed evolution in mammalian cells can facilitate the engineering of mammalian-compatible biomolecules and can enable synthetic evolvability for mammalian cells. We engineered an orthogonal alphaviral RNA replication system to evolve synthetic RNA-based devices, enabling RNA replicase-assisted continuous evolution (REPLACE) in live mammalian cells. we employed REPLACE to drive the continuous intracellular evolution of the cancer-related protein MEK1 with the aim of conferring resistance to Cobimetinib. To investigate the accumulation of mutations during this evolutionary process, we conducted amplicon sequencing on experimental materials collected at different stages. The results revealed intricate relationships among different mutations, highlighting the complex nature of the evolutionary landscape.
2024-07-07 | GSE235328 | GEO
Project description:Studies of the RNA polymerase-binding molecule ppGpp in bacteria and plants have shown that changes to the kinetics of the RNA polymerase can have dramatic biological effects in the short-term as a stress response. Here we describe the reprogramming of the kinetic parameters of the RNAP through mutations arising during laboratory adaptive evolution of Escherichia coli in minimal media. The mutations cause a 10- to 30-fold decrease in open complex stability at a ribosomal promoter and approximately a 10-fold decrease in transcriptional pausing in the his operon. The kinetic changes coincide with large scale transcriptional changes, including strong downregulation of motility, acid-resistance, fimbria, and curlin genes which are observed in site-directed mutants containing the RNA polymerase mutations as well as the evolved strains harboring the mutations. Site-directed mutants also grow 60% faster than the parent strain and convert the carbon-source 15% to 35% more efficiently to biomass. The results show that long-term adjustment of the kinetic parameters of RNA polymerase through mutation can be important for adaptation to a condition. Mutations in the RNA polymerase beta prime subunit (rpoC) were discovered in E. coli following adaptation to continual logarithmic growth (OD <= 0.3) in glycerol M9 minimal media at 30 C. We used site-directed mutagenesis to make strains of E. coli isogenic to wild-type except for single adaptive rpoC mutations. We found they increase growth rate in this condition by 60%. In order to understand how the mutations affect gene expression in this condition, we extracted total mRNA from the strains, which had been growing in the adaptive evolution condition, at OD=0.3. There were three RNAP mutants and the wild-type (E. coli K-12 MG1655). Each strain had three flasks from which RNA was extracted (three biological replicates). There were no technical replicates. The mRNA was synthesized into cDNA, labeled, and hybridized to an Affymetrix E. coli 2.0 GeneChip.
2010-04-12 | E-GEOD-15500 | biostudies-arrayexpress
Project description:Directed biomolecular evolution is widely used to tailor and enhance proteins but has hitherto not been applied in the reprogramming of mammalian cells. Here we describe a novel method to identify artificially enhanced and evolved reprogramming factors by pooled screens with randomised protein libraries, cell selection based on phenotypic readouts and genotyping by amplicon sequencing. We benchmark this approach by identifying artificially enhanced Sox2 and Sox17 factors in pluripotency reprogramming.
2018-07-12 | GSE107987 | GEO
Project description:Studies of the RNA polymerase-binding molecule ppGpp in bacteria and plants have shown that changes to the kinetics of the RNA polymerase can have dramatic biological effects in the short-term as a stress response. Here we describe the reprogramming of the kinetic parameters of the RNAP through mutations arising during laboratory adaptive evolution of Escherichia coli in minimal media. The mutations cause a 10- to 30-fold decrease in open complex stability at a ribosomal promoter and approximately a 10-fold decrease in transcriptional pausing in the his operon. The kinetic changes coincide with large scale transcriptional changes, including strong downregulation of motility, acid-resistance, fimbria, and curlin genes which are observed in site-directed mutants containing the RNA polymerase mutations as well as the evolved strains harboring the mutations. Site-directed mutants also grow 60% faster than the parent strain and convert the carbon-source 15% to 35% more efficiently to biomass. The results show that long-term adjustment of the kinetic parameters of RNA polymerase through mutation can be important for adaptation to a condition.
2010-04-01 | GSE15500 | GEO
Project description:This SuperSeries is composed of the following subset Series: GSE36129: An IDN2-containing complex involved in RNA-directed DNA methylation in Arabidopsis [leaves RNA-seq] GSE36143: An IDN2-containing complex involved in RNA-directed DNA methylation in Arabidopsis [BS-seq] GSE37206: An IDN2-containing complex involved in RNA-directed DNA methylation in Arabidopsis [flowers RNA-seq] Refer to individual Series
2012-05-25 | E-GEOD-36148 | biostudies-arrayexpress
Project description:DNA methylation is a conserved epigenetic mark in plants and many animals. How parental alleles interact in progeny to influence the epigenome is poorly understood. We analyzed the DNA methylomes of Arabidopsis Col and C24 ecotypes, and their hybrid progeny. Hy- brids displayed nonadditive DNA methylation levels, termed meth- ylation interactions, throughout the genome. Approximately 2,500 methylation interactions occurred at regions where parental DNA methylation levels are similar, whereas almost 1,000 were at differ- entially methylated regions in parents. Methylation interactions were characterized by an abundance of 24-nt small interfering RNAs. Furthermore, dysfunction of the RNA-directed DNA methylation pathway abolished methylation interactions but did not affect the increased biomass observed in hybrid progeny. Methylation interac- tions correlated with altered genetic variation within the genome, suggesting that they may play a role in genome evolution. Whole genome bisulfite sequencing and small RNA sequencing of the wild type and nrpd1nrpe1 double mutant background of parent Col ,C24, the hybrid ColXC24 and C24XCol to explore the role of the RdDM pathway in DNA methylation interactions.
2016-06-15 | E-GEOD-72993 | biostudies-arrayexpress