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.
Project description:It is known that current the-art-of-the-state TadA8 and TadA8e which evolved from E. coli TadA. They inherited the 'YA' context from tRNA deaminase. We started with wildtype E. coli TadA and designed an evolution campaign to force TadA variants to deaminate “GA” with fast kinetics. Three rounds of de novo directed evolution followed by DNA shuffling led to TadA8r, a TadA variant of superior “RA” deamination activity. TadA8r acts on a broadened editing window when fused to Streptococcus pyogenes Cas9 (SpCas9) and delivers robust editing at PAM distal positions. While highly active at on-target sites, ABE8r shows off-target DNA and RNA editing much lower than ABE8e. The off-target effects of ABE8r can be further mitigated by introducing a V106W substitution23, a R153 deletion22, or by mRNA delivery. Lastly, we demonstrate ABE8r-mediated correction of G1961E in ABCA4, the most prevalent mutation driving Stargardt disease (STGD1), in a “GA” context. ABE8r, with its superior activity and broadened context compatibility, complements and expands the current ABE family.
Project description:Primary human macrophages in in vitro conditions have been exposed to hypergravity and microgravity during the 28th DLR parabolic flight campaign
Project description:This data set contains 1376 mass spectrometry reads from root, rhizosphere and leaf sample of Populus Trichocarpa, as well as associated controls. This metabolomics data set was collected as part of a larger campaign which complements the metabolomics data with metagenome sequencing, transcriptomics, and soil measurement data.
Project description:Illumina-based whole genome bisulfite sequencing libraries for 4 RNA-directed DNA methylation mutants Examining DNA methylation levels in leaf tissue of RNA-directed DNA methylation mutants
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.
Project description:Using in vitro directed evolution, we show that mismatch repair-deficient Pseudomonas aeruginosa can engage novel resistance mechanisms to ceftazidime-avibactam.
