Advanced searches left 3/3

Cas9 - Europe PMC

Summarized by Plex Scholar
Last Updated: 10 September 2022

* If you want to update the article please login/register

DUSP2 deletion with CRISPR/Cas9 promotes Mauthner cell axonal regeneration at the early stage of zebrafish.

Dusp2 was discovered as upregulated gene in zebrafish with spinal cord injury, according to a previous research. We found that dual specificity phosphatase 2 is a negative regulator of axon regeneration of the Mauthner cell in this region. Dusp2 axons of dusp2 -/- zebrafish had a faster recovery at the early stage after birth, while those of dusp2 +/- zebrafish did not. Overexpression of DUSP2 in Tg zebrafish by single-cell electroporation has slowed the regeneration of M-cell axons.

Source link: https://europepmc.org/article/MED/36018180


CRISPR/Cas9 - mediated knock-in method can improve the expression and effect of transgene in P1 generation of channel catfish (Ictalurus punctatus)

Transgenesis has a variety of uses in fish raising and the design of fish models. Channel catfish is an economically important farmed fish in the United States and is a highly popular farmed fish. Genomic real-time PCR found that the CRISPR/Cas9 transgenic fish had significantly higher average transgene copy numbers than randomly embedded transgenic fish. In addition, reverse transcription PCR and fatty acids analysis showed that CRISPR/Cas9 P1 fish had high OmElovl2 transgene expression in most tissues and 20% higher DHA relative to their controls, with 20 percent higher DHA among those controls, while randomly integrated P1 fish did not have detectable OmElovl2 expression in any of five tissues examined. There were no significant differences between transgenic fish produced by random integration and their non-transgenic controls.

Source link: https://europepmc.org/article/MED/IND607803567


High-throughput continuous evolution of compact Cas9 variants targeting single-nucleotide-pyrimidine PAMs.

Despite the availability of Cas9 variants with various protospacer-adjacent motif compatibilities, some genomic loci, particularly those with pyrimidine-rich PAM sequences, are unobtainable by high-activity Cas9 proteins. We created four Cas9 variants that enable targeting of the majority pyrimidine-rich PAM sequences in the human genome, with directed evolution. Variants eNme2-T. 1 and eNme2-T. 2 include N4TN PAM sequences with similar editing results as current variants, while eNme2-C and eNme2-C. NR's eNme2-T. 1 and eNme2-T. 1 include less stringent PAM specifications, comparable or higher up-target operation at N4CN PAM sequences, as well as lower off-target operation at N4CN PAM sequences, eNme2-T.

Source link: https://europepmc.org/article/MED/36076084


Multi-pathway DNA-repair reporters reveal competition between end-joining, single-strand annealing and homologous recombination at Cas9-induced DNA double-strand breaks.

Multiple pathways are used to repair error-free repair to mutagenesis and genomic loss, with results ranging from error-free repair to genetic modification, and genomic loss. DSB-repair by error-free canonical non-homologous end-joining versus homologous recombination, mutagenic repair versus HR, and mutagenic end-joining versus single strand annealing versus HR based on statistical comparison versus HR - a difference between DSB-repair by error-free canonical non-homologous end-joining versus HR versus DSB-he versus homologous end-logistic end-joining vs versus homologous end-joination versus recombination versus HR versus homologistic end-joining versus annealing versus single versus HR versus HR versus HR versus HR versus HR versus HR versus HR versus HR versus HR According to our results, SSA-mediated DSB-repair also occurs at endogenous genomic loci, owing to Alu elements or homologous gene regions. These new Cas9-based DSB-Spectrum reporters support the thorough analysis of repair pathway crosstalk and DSB-repair results.

Source link: https://europepmc.org/article/MED/36075911


Targeted dual base editing with Campylobacter jejuni Cas9 by a single AAV-mediated delivery

We created cjCas9-based base editors, as well as a cytosine base editor and an adenine base editor that can reliably induce endogenous base substitutions of up to 91. 2% in HEK293T cells. The active windows of cjCBEmax and cjABE8e were larger than those of spCas9-based base editors, according to a study of the base editing effectiveness of 13 endogenous target sites, and their specificities were marginally smaller than that of cjCas9. Engineered cjCas9 and gRNA scaffold can raise the base editing yield of cjABE8e by up to 6. 4 percent in HEK293T cells.

Source link: https://europepmc.org/article/PPR/PPR541555


A CRISPR/Cas9-based enhancement of high-throughput single-cell transcriptomics

Single-cell transcriptomics has suffered as a result of lapses in coverage of the full transcriptome, revealing an incomplete gene expression profile of the cell. We describe a validation of scCLEAN showing a 2. 1-fold increase in library complexity with negligible off-target effects, using immune cells. Unique isoform detection has increased by 4. 6 percent when applying scCLEAN to single-cell isoseq samples, resulting in a 4. 6-fold increase in unique isoform detection. We then illustrate the ability of scCLEAN to elucidate biological data by testing two participant cohorts of cardiovascular samples, exposing new molecular features, including inflammatory signatures.

Source link: https://europepmc.org/article/PPR/PPR541365


Fusing an exonuclease with Cas9 enhances homologous recombination in Pichia pastoris.

Conclusions The endogenous exonuclease Mre11 and Exo1 had the highest positive rates in seamless FAA1 removal, and bonding the MRE11 to the C-terminal of CAS9 had the highest positive rate and a small number of clones. When overexpressing RAD52, we discovered that the expression of CAS9-MRE11 remarkably raised success rates when simultaneously simulating double genes and three genes. Conclusions The fusion of the endogenous exonuclease Mre11 with Cas9 improves homologous recombination efficiency in P. pastoris. The objective set forth here would enable P. pastoris' metabolic engineering to lead to the production of value-added compounds at high levels.

Source link: https://europepmc.org/article/MED/36071435


Drosophila CRISPR/Cas9 mutants as tools to analyse cardiac filamin function and pathogenicity of human FLNC variants.

Filamins are large proteins with actin binding capabilities. We wanted to investigate these diseases in vivo here, using Drosophila melanogaster as a new in vivo model. We demonstrate that adult-specific cardiac RNAi-induced dilution, impaired systolic function, and sarcomeric changes were all present in the adult stage, raising the need for cardiac function and maintainer of sarcomere stability. Finally, we conclude that deletions of the C-term portion of the dFil's last four Ig-like domains are dispensable for cardiac function.

Source link: https://europepmc.org/article/MED/36066120


Efficient repair of human homozygous genetic mutation by CRISPR/Cas9 mediated interlocus gene conversion

The bulk of DNA double-strand breaks caused by gene editing software are mainly resolved by non-homologous end joining or homology-directed reconstruction with exogenous synthetic DNA templates. However, the therapeutic use of HDR could be particularly difficult given the requirement for the codelivery of exogenous DNA templates with toxicity into cells, as well as the low rate of HDR could also limit its clinical use. Here, we used hematopoietic stem cells with genetic mutations to produce u03b2-thalassemia in HBB coding regions, discovering that several cells are repaired by CRISPR/Cas9-mediated gene transfer independent of exogenous synthetic DNA templates. Cas9's conversion of mutations from ribonucleoprotein with sgRNA to haematopoietic stem and progenitor cells without exogenous DNA template has resulted in efficient conversion of mutations to normal wild-type sequences lacking exogenous DNA template.

Source link: https://europepmc.org/article/PPR/PPR540288

* Please keep in mind that all text is summarized by machine, we do not bear any responsibility, and you should always check original source before taking any actions

* Please keep in mind that all text is summarized by machine, we do not bear any responsibility, and you should always check original source before taking any actions