VEGAS: A Platform for Continuous Directed Evolution in Mammalian Cells (RNAseq)
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA507988 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR10126416_1.fastq.gz | Fastqsanger.gz | |||
| SRR10126416_2.fastq.gz | Fastqsanger.gz | |||
| SRR10126417_1.fastq.gz | Fastqsanger.gz | |||
| SRR10126417_2.fastq.gz | Fastqsanger.gz | |||
| SRR10126418_1.fastq.gz | Fastqsanger.gz |
Items per page: 5 1 - 5 of 43 |
Similar Datasets
Project description:Sindbis virus is a mutagenic ssRNA arbor virus. The structural genome component of this virus can be exchanged for any transgene of < 6kb in size. Here we sequence transgenic elements inserted to the virus. Our EGFP transgene was analyzed as a function of time during viral replication to quantify the mutagenic rate of the virus. Our nanobody transgene was analyzed to assess cDNA library complexity and identity.
2018-12-04 | GSE123269 | GEO
Project description:The VEGAS platform is not suitable for mammalian directed evolution
| PRJEB47639 | ENA
Project description:Replicative RNA enables directed evolution and Darwinian adaptation in mammalian cells
| PRJNA985713 | ENA
Project description:Directed evolution, artificial selection toward designed objectives, is routinely used to develop new molecular tools and therapeutics. Successful directed molecular evolution campaigns repeatedly test diverse sequences with a designed selective pressure. Unicellular organisms and their viral pathogens are exceptional for this purpose and have been used for decades. However, many desirable targets of directed evolution perform poorly or unnaturally in unicellular backgrounds. Here, we present a system for facile directed evolution in mammalian cells. Using the RNA alphavirus Sindbis as a vector for heredity and diversity, we achieved 24-h selection cycles surpassing 10-3 mutations per base. Selection is achieved through genetically actuated sequences internal to the host cell, thus the system's name: viral evolution of genetically actuating sequences, or "VEGAS." Using VEGAS, we evolve transcription factors, GPCRs, and allosteric nanobodies toward functional signaling endpoints each in less than 1 weeks' time.
| S-EPMC6660416 | biostudies-literature
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: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. Using REPLACE, we attempted continuous intracellular evolution of the negative dominant mutant KRAS (S17N). To analyze the process of mutation accumulation, we performed amplicon sequencing on experimental materials at different stages and under different treatment conditions. The results indicated that the mutations generated by this system were primarily induced by Monanunavir, and the addition of Monanunavir significantly accelerated the rate of evolution.
2024-07-07 | GSE235327 | GEO
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:Directed evolution of orthogonal aminoacyl-tRNA synthetases (AARSs) enables site-specific installation of noncanonical amino acids (ncAAs) into proteins. Traditional evolution techniques typically produce AARSs with greatly reduced activity and selectivity compared to their wild-type counterparts. We designed phage-assisted continuous evolution (PACE) selections to rapidly produce highly active and selective orthogonal AARSs through hundreds of generations of evolution. PACE of a chimeric Methanosarcina spp. pyrrolysyl-tRNA synthetase (PylRS) improved its enzymatic efficiency (kcat/KMtRNA) 45-fold compared to the parent enzyme. Transplantation of the evolved mutations into other PylRS-derived synthetases improved yields of proteins containing noncanonical residues up to 9.7-fold. Simultaneous positive and negative selection PACE over 48 h greatly improved the selectivity of a promiscuous Methanocaldococcus jannaschii tyrosyl-tRNA synthetase variant for site-specific incorporation of p-iodo-L-phenylalanine. These findings offer new AARSs that increase the utility of orthogonal translation systems and establish the capability of PACE to efficiently evolve orthogonal AARSs with high activity and amino acid specificity.
| S-EPMC5724969 | biostudies-literature
Project description:Evolution of mammalian miRNA genes
| PRJNA174234 | ENA
Project description:MicroRNAs (miRNAs) are major post-transcriptional regulators of gene expression, yet their origins and functional evolution in mammals remain little understood due to the lack of appropriate comparative data. Using RNA sequencing, we have generated extensive and comparable miRNA data for five organs in six species that represent all main mammalian lineages and birds (the evolutionary outgroup), with the aim to unravel the evolution of mammalian miRNAs. Our analyses reveal an overall expansion of miRNA repertoires in mammals, with three-fold accelerated birth rates of miRNA families in placentals and marsupials, facilitated by the de novo emergence of miRNAs in host gene introns. Generally, our analyses suggest a high rate of miRNA family turnover in mammals, with many newly emerged miRNA families being lost soon after their formation. Selectively preserved mammalian miRNA families gradually evolved higher expression levels as well as altered mature sequences and target gene repertoires, and were apparently mainly recruited to exert regulatory functions in nervous tissues. However, miRNAs that originated on the X chromosome evolved high expression levels and potentially diverse functions during spermatogenesis, including meiosis, through selectively driven duplication-divergence processes. Overall, our study thus provides detailed insights into the birth and evolution of mammalian miRNA genes and the associated selective forces. 30 main samples from five adult tissues (brain, cerebellum, heart, kidney and testis) collected in six species (human, macaque, mouse, opossum, platypus and chicken) + 5 biological replicates + 3 samples from spermatogenic cells in adult mouse testis (Sertoli cells, spermatocytes and spermatids)
2012-10-01 | E-GEOD-40499 | biostudies-arrayexpress