Project description:We performed ChIP-seq to measure H3K9me3 levels in wild-type HeLa cells and HeLa cells lacking TASOR, MPP8, periphilin and SETDB1 generated through CRISPR/Cas9-mediated gene disruption.
Project description:Ensuring genome safety during gene editing is crucial for clinical translation of the high-efficient CRISPR-Cas9 toolbox. Therefore, the undesired events including chromosomal translocations, vector integrations, and large deletions arising during therapeutic gene editing remain to be adequately addressed or tackled in vivo. Here, we apply CRISPR-Cas9TX in comparison to CRISPR-Cas9 to target Vegfa for the treatment of age-related macular degeneration (AMD) disease in a mouse model. AAV delivery of both CRISPR-Cas9 and CRISPR-Cas9TX can efficiently inhibit laser-induced neovascularization. Importantly, Cas9TX almost eliminates chromosomal translocations that occur at a frequency of approximately 1% in Cas9-edited mouse retinal cells. Strikingly, the widely observed AAV integration at the target Vegfa site is also greatly dropped from nearly 50% of edited events to the background level during Cas9TX editing. Our findings reveal that chromosomal structural variations routinely occur during in vivo genome editing and highlight Cas9TX as a superior form of Cas9 for in vivo gene disruption.
Project description:Mitochondrial supercomplexes are functionally important, but how supercomplexes being assembled is not known. Mitochondrial Complex I has an important role in the assembly of supercomplex. In this study, we generated a series of cell model with shRNA mediated gene silencing or CRISPR-Cas9 gene knockout technology. We performed a series of mitochondrial proteome analyses in different cell lines with disruption of gene that encodes subunit of Complex I. Also, we performed Co-IP of exogenous TIMMDC1, which was an assembly factor of Complex I.
Project description:By comparing HeLa cells lacking MORC2 or SETDB1 generated through CRISPR/Cas9-mediated gene disruption to wild-type HeLa cells, the goal of the experiment was to determine the effect of loss of MORC2 on the distribution of the repressive H3K9me3 histone modification.
Project description:By comparing HeLa cells lacking ATF7IP or SETDB1 generated through CRISPR/Cas9-mediated gene disruption to wild-type HeLa cells, the goal of the experiment was to determine the effect of loss of the SETDB1•ATF7IP complex on the distribution of the repress
Project description:HeLa cells lacking MORC2 generated through CRISPR/Cas9-mediated gene disruption were reconstituted with either wild-type or R252W mutant MORC2, and re-repression of HUSH target genes assessed by RNA-seq
Project description:By comparing HeLa cells lacking MORC2 generated through CRISPR/Cas9-mediated gene disruption to wild-type HeLa cells, the goal of the experiment was to determine the effect of loss of MORC2 on the transcriptome.
Project description:Human acute myeloid leukemia cell lines OCI-AML2 and OCI-AML3 were used in a CRISPR/Cas9-mediated approach to specifically target DDX3X’s gene sequences encoding the RNA binding domain of the helicase. DDX3X RNA binding domain is bipartite in the two halves of the helicase core. sgRNAs were designed to target both halves of the domain (named RNA binding domain A and B – RBDA and RBDB). We performed RNA-seq to observe the gene expression changes in both OCI-AML2 and OCI-AML3 cell lines following the not-combined CRISPR/Cas9 –mediated targeting of both regions of the DDX3X RNA binding domain. Control CRISPR/Cas9 performed with no sgRNA expressing vector (named “empty vector”) was performed in both cell lines. The latter condition was used as a control for gene expression changes analysis, for each cell line.
