Project description:Whole-genome resequencing of eight transcription factor mutants and one wild-type, in order to verify the T-DNA insertion site and its uniqueness.
Project description:Copy Number Variations (CNVs) were identified performing Comparative Genomic Hybridization (CGH) on 225 patients after whole-genome amplification, using Agilent SurePrint G3 4x180K microarrays. CNVs were further integrated with gene expression (Affymetrix U133+2 arrays) and mutations (targeted DNA resequencing). Complete description of the methods, array quality checks and called segments are available as supplemental material in the corresponding publication.
Project description:Background: Evolutionary engineering is a powerful approach to isolate suppressor mutants and industrially relevant genotypes. Until recently, DNA microarray analysis was the only affordable genome-wide approach to identify the responsible mutations. This situation has changed due to the rapidly decreasing costs of whole genome (re)sequencing. DNA microarray-based mRNA expression analysis and whole genome resequencing were combined in a study on lactate transport in Saccharomyces cerevisiae. Jen1p is the only S. cerevisiae lactate transporter reported in literature. To identify alternative lactate transporters, a jen1Δ strain was evolved for growth on lactate. Results: Two independent evolution experiments yielded Jen1p-independent growth on lactate (μmax 0.14 and 0.18 h-1 for single-cell lines IMW004 and IMW005, respectively). Whereas mRNA expression analysis did not provide leads, whole-genome resequencing showed different single nucleotide changes (C755G/Leu219Val and C655G/Ala252Gly) in the acetate transporter gene ADY2. Analysis of mRNA levels and depth of coverage of DNA sequencing combined with karyotyping, gene deletions and diagnostic PCR showed that in IMW004 an isochromosome III (~475 kb), which contains two additional copies of ADY2C755G, was formed via crossover between YCLWΔ15 and YCRCΔ6. Introduction of the ADY2 alleles in a jen1 ady2 strain resulted in growth on lactate (μmax 0.14 h-1 for Ady2pLeu219Val and 0.12 h-1 for Ady2pAla252Gly). Conclusions: Whole-genome resequencing of yeast strains obtained from independent evolution experiments enabled rapid identification of a key gene that was not identified by mRNA expression analysis of the same strains. Reverse metabolic engineering showed that mutated alleles of ADY2 (C655G and C755G) encode efficient lactate transporters.
Project description:Illumina human Omni5Exome arrays were used to investigate CNVs in Sѐzary syndrome tumours as part of a larger study involving whole exome sequencing of the same samples and targeted resequencing of a further cohort.
Project description:Four groups of C57 mice were gavaged with CMC, ethanol, DEHP and ethanol+DEHP for 28 continuous days. At the end of the experiment, livers were harvested to carry out Whole genome resequencing, we found that metabolism related genes were significantly changed in the DEHP and ethanol+DEHP groups compared with control groups, but there was no significant change between them.
Project description:Current human pluripotent stem cells lack the transcription factor circuitry that governs the ground state of mouse embryonic stem cells (ESC). Here we report that short-term expression of two components, NANOG and KLF2, is sufficient to ignite other elements of the network and reset the human pluripotent state. Inhibition of ERK and protein kinase C signalling sustains a transgene-independent rewired state. Reset cells self-renew continuously without ERK signalling, are phenotypically stable and karyotypically intact. They differentiate in vitro and form teratomas in vivo. Metabolism is reprogrammed in reset cells with activation of mitochondrial respiration as in ESC. DNA methylation is dramatically reduced and transcriptome state is globally realigned across multiple cell lines. Depletion of ground state transcription factors, TFCP2L1 or KLF4 has marginal impact on conventional human pluripotent stem cells, but collapses the reset state. These findings demonstrate feasibility of installing and propagating functional control circuitry for ground state pluripotency in human cells. DNA methylation analysis in Conventional and Reset human embryonic stem cells by whole genome bisulfite sequencing, in triplicate, using the Illumina platform
Project description:Primary objectives: The primary objective is to investigate circulating tumor DNA (ctDNA) via deep sequencing for mutation detection and by whole genome sequencing for copy number analyses before start (baseline) with regorafenib and at defined time points during administration of regorafenib for treatment efficacy in colorectal cancer patients in terms of overall survival (OS).
Primary endpoints: circulating tumor DNA (ctDNA) via deep sequencing for mutation detection and by whole genome sequencing for copy number analyses before start (baseline) with regorafenib and at defined time points during administration of regorafenib for treatment efficacy in colorectal cancer patients in terms of overall survival (OS).