Project description:We analyzed the transcriptome of dormant and after-ripened imbibed seeds of the Arabidopsis accession Cape verde Islands. We analyzed the transcriptional changes in temporal and spatial detail by sampling at four imbibition time-points (3, 7, 12 and 24 hours after sowing) and two seed compartments (RAD and MCE).
Project description:Many Arabidopsis thaliana accession show sensitvity to the air pollutant ozone, including the accession Cvi-0 from the Cape Verde Islands. To understand and assist in genetic mapping of loci causing the ozone sensitvity of Cvi-0, transcript profiling was performed in Cvi-0, the tolerant Col-0, and a near isogenic line (Col-S) where ozone sensitivity was introgressesed from Cvi-0 to Col-0 through eight rounds of backcrossing.
Project description:We analyzed the transcriptome of dormant and after-ripened imbibed seeds of the Arabidopsis accession Cape verde Islands. We analyzed the transcriptional changes in temporal and spatial detail by sampling at four imbibition time-points (3, 7, 12 and 24 hours after sowing) and two seed compartments (RAD and MCE). Gene expression in imbibed dormant and after-ripened seeds was compared. For each sample three replicates were used.
Project description:Generation of transgenic cell lines is limited by inefficient gene editing requiring genotypic screening of hundreds to thousands of colonies to isolate correctly gene-edited cells. Here, we describe a novel method called CRISPRa On-Target Editing Retrieval (CRaTER) that enriches for cells with on-target knock-in of a promoterless cDNA-fluorescent reporter transgene by transiently overexpressing the targeted endogenous genetic locus and sorting for fluorescent cells. We use CRaTER to enrich for rare cells with heterozygous, biallelic-editing of the endogenous, transcriptionally-inactive MYH7 locus in human induced pluripotent stem cells (hiPSC), resulting in a 9-fold enrichment compared to antibiotics selection alone. We leveraged CRaTER to enrich for heterozygous knock-in of a library of single nucleotide variants (SNV) in MYH7, a gene encoding for sarcomeric MHC-β wherein autosomal dominant missense mutations cause cardiac and skeletal myopathies. CRaTER enabled 90% enrichment of heterozygous, biallelically-edited hiPSCs – a 38.6-fold enrichment compared to antibiotics selection alone – to generate 113 SNVs comprising 78 missense variants in MYH7. hiPSCs that have undergone CRaTER enrichment can differentiate to cardiomyocytes and exhibit expected localization and expression of MHC-β fusion proteins. Together, CRaTER substantially reduces the screening required for isolation of gene-edited cells, enabling the generation of transgenic cell lines at unprecedented scale.