Project description:We performed a large-scale genome-wide characterisation of indels generated following editing with CRISPR/Cas9. We used pools of sgRNAs and performed targeted capture and sequencing of the edited regions in HepG2 cells.
Project description:Genome editing was conducted on a t(3;8) K562 model to investigate the effects of deleting different modules or CTCF binding sites within the MYC super-enhancer. To check mutations after targeting with CRISPR-Cas9 we performed amplicon sequencing using the Illumina PCR-based custom amplicon sequencing method using the TruSeq Custom Amplicon index kit (Illumina). The first PCR was performed using Q5 polymerase (NEB), the second nested PCR with KAPA HiFi HotStart Ready mix (Roche). Samples were sequenced paired-end (2x 250bp) on a MiSeq (Illumina).
Project description:To compare the impact of CRISPR-egineered R175 TP53 mutant variants in HCT116 and H460 cells, mutations at the amino acid position 175 were generated systematically by CRISP/Cas9 editing. Here, genomic amplicon regions covering the TP53 Exons 5 were sequenced via targeted sequencing.
Project description:We recreated the t(7;12) translocation in K562 cells by CRISPR/Cas9 to understand its effects on haematopoietic cells, which is of relevance to understand how this cytogenetic abnormalities causes and promotes acute leukaemia in infants. Wild-type K562 were edited by electroporation of ribonucleoprotein complexes consisting of Cas9 enzyme and two guide RNAs targeting patient-specific breakpoint loci. K562 electroporated with Cas9 enzyme only were used as control. Edited K562 harbouring the t(7;12) were single-cell cloned to obtain homogeneous populations (hereby referred to as K562-t(7;12)). We performed RNA sequencing analysis of K562-t(7;12) compared to K562 control to uncover transcriptional changes associated with the translocation.