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Simultaneous in vivo enzyme and host engineering through laboratory evolution

ABSTRACT: Simultaneous in vivo enzyme and host engineering through laboratory evolution

PROVIDER: PRJEB26704 | ENA |

REPOSITORIES: ENA

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Simultaneous in vivo enzyme and host engineering through laboratory evolution

Project description:Two genetic selection systems that couple metabolite hydroxylation or methylation of small molecules to growth of Escherichia coli are presented in this study. One system targets pterin-dependent hydroxylation (tBPt) while another focuses on S-adenosylmethionine-dependent methylation (SAM). Using adaptive laboratory evolution with growth selection, these two systems are demonstrated to not only achieve in vivo directed evolution of enzymes involved in human hormone biosynthesis but also reveal non-intuitive host factors that elude existing synthetic biology approaches. Raw sequencing data for the relevant strains generated in this study are presented here.

2019-01-30 | E-MTAB-6779 | biostudies-arrayexpress

Simultaneous multifunctional transcriptome engineering by CRISPR RNA scaffold

Project description:RNA processing and metabolism are subjected to precise regulation in the cell to ensure integrity and functions of RNA. Though targeted RNA engineering has become feasible with the discovery and engineering of the CRISPR-Cas13 system, simultaneous modulation of different RNA processing steps remains unavailable. In addition, the off-target events resulting from the effectors fused with dCas13 limit its application. Here we developed a novel platform, Combinatorial RNA Engineering via Scaffold Tagged gRNA (CREST), which can simultaneously execute multiple RNA modulation functions on different RNA targets. In CREST, RNA scaffolds are appended to the 3’ end of Cas13 gRNA and their cognate RNA binding proteins are fused with enzymatic domains for manipulation. We show that CREST is capable of simultaneously manipulating RNA alternative splicing and A-to-G or C-to-U base editing. Furthermore, by fusing two split fragments of the deaminase domain of ADAR2 to dCas13 and PUFc respectively, we reconstituted its enzyme activity at target sites. This split design can reduce more than 90% of off-target events otherwise induced by a full-length effector. The flexibility of the CREST framework will enrich the transcriptome engineering toolbox for the study of RNA biology and the development of RNA-focused therapeutics

2022-06-26 | GSE206745 | GEO

Simultaneous multifunctional transcriptome engineering by CRISPR RNA scaffold

Project description:Simultaneous multifunctional transcriptome engineering by CRISPR RNA scaffold

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Identification of a novel metabolic engineering target for carotenoid production in Saccharomyces cerevisiae via ethanol-induced adaptive laboratory evolution

Project description:Carotenoids are a large family of health-beneficial compounds that have been widely used in the food and nutraceutical industries. There have been extensive studies to engineer Saccharomyces cerevisiae for the production of carotenoids, which already gained high level. However, it was difficult to discover new targets that were relevant to the accumulation of carotenoids. Herein, a new, ethanol-induced adaptive laboratory evolution was applied to boost carotenoid accumulation in a carotenoid producer BL03-D-4, subsequently, an evolved strain M3 was obtained with a 5.1-fold increase in carotenoid yield. Through whole-genome resequencing and reverse engineering, loss-of-function mutation of phosphofructokinase 1 (PFK1) was revealed as the major cause of increased carotenoid yield. Transcriptome analysis was conducted to reveal the potential mechanisms for improved yield, and strengthening of gluconeogenesis and downregulation of cell wall-related genes were observed in M3. This study provided a classic case where the appropriate selective pressure could be employed to improve carotenoid yield using adaptive evolution and elucidated the causal mutation of evolved strain.

2021-01-09 | GSE164470 | GEO

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Project description:We used RNA-seq to profile E. coli K-12 MG1655 strains subjected to adaptive laboratory evolution after knockout of endogenous glucose-6-phosphate isomerase (pgi) and subsequent expression of heterologous version of the pgi gene from Pseudomonas aeruginosa and Bacillus megaterium.

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Adaptive laboratory evolution of chorismate synthase knockouts

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2023-02-08 | GSE224889 | GEO

The Ministry of Education key laboratory of dairy biotechnology and engineering

Project description:The Ministry of Education key laboratory of dairy biotechnology and engineering Raw sequence reads

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Project description:We have performed adaptive laboratory evolution of E. coli pdhR gene deletion strain to examine the adaptive strategies of E. coli.

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