Similar Datasets
Project description:In this work we analyzed PFS sequence recognized by SuCas12a2 Type V CRISPR-Cas nuclease
2023-01-11 | GSE178530 | GEO
Project description:In this work we analyzed crRNA processing by SuCasΩ Type V CRISPR-Cas nuclease in E. coli
2023-01-11 | GSE178531 | GEO
Project description:In this work we analyzed possible protospacer flanking sequences recognized by GeCas12a2 Type V CRISPR-Cas nuclease.
2026-03-18 | GSE325193 | GEO
Project description:We generated a collection of 13 plasmids, with each plasmid containing a variant of a CRISPR protospacer targeted by spacer 8 of the E. coli CRISPR-I array. We transformed the plasmids as a pool into delta cas3 E. coli cells expressing all other cas genes constitutively. We then transformed these cells with either an empty vector or a plasmid expressing the Cas3 nuclease. DNA surrounding the protospacers was PCR-amplified and sequenced.
2017-08-24 | E-MTAB-5969 | biostudies-arrayexpress
Project description:Our results show that compared to wild type, a deletion mutant of the cas3 gene, an essential nuclease part of the class 1 type I CRISPR-Cas system, increases the virulence of P gingivalis.
2020-09-16 | GSE154569 | GEO
Project description:The type V-I CRISPR-Cas system is becoming increasingly attractive for its potential utility in gene editing. However, natural nucleases often exhibit low efficiency, limiting their application. Here, we utilized structure-guided rational design and combinatorial protein engineering to optimize an uncharacterized Cas12i nuclease, Cas12i3. Accordingly, we developed Cas-SF01, a Cas12i3 variant that exhibits significantly improved gene-editing activity in mammalian cells and plants. Cas-SF01 displays comparable or superior editing performance compared to SpCas9 or recently engineered Cas12 nucleases. Further analysis of PAM recognition showed that Cas-SF01 has an expanded PAM range and effectively recognizes NTTN and noncanonical NATN and TTVN PAMs. Additionally, we identified an amino acid substitution, D876R, that markedly reduced the off-target effect while maintaining high on-target activity, leading to the development of Cas-SF01HiFi (high-fidelity Cas-SF01). Finally, we demonstrated that Cas-SF01 has robust gene-editing activity in both the monocot plant rice and dicot plant pepper. Our results suggest that Cas-SF01 can serve as a robust gene-editing platform with high efficiency and specificity for future genome editing applications across different organisms.
2023-11-05 | GSE236755 | GEO
Project description:CRISPR–Cas systems provide adaptive immunity in prokaryotes by targeting and degrading invasive genetic elements. Among them, the type I-F2 system represents the most compact type I CRISPR–Cas system, distinguished by the absence of small and large subunits. These subunits are otherwise essential for target recognition and nuclease recruitment. To elucidate how this minimal system mediates DNA interference, we determined the cryo-electron microscopy (cryo-EM) structure of the I-F2 Cascade complex bound to its target DNA and the effector helicase-nuclease Cas3. Our structure reveals a unique mechanism of Cas3 recruitment, predominantly mediated by the Cas7 subunit. We show how the helicase and C-terminal domains of Cas3 engage the single-stranded DNA to initiate directional DNA-unwinding and degradation. These findings uncover key mechanistic adaptations that enable efficient interference in the absence of canonical subunits and emphasize the mechanistic diversity among closely related type I systems, including I-E, I-F1, and I-F2. These insights provide a structural basis for engineering the minimal I-F2 system for genome editing and biotechnological applications.
2026-03-09 | PXD068253 | Pride
Project description:Base editors are RNA-guided deaminases that enable site-specific nucleotide transitions. The targeting scope of these Cas-deaminase fusion proteins critically depends on the availability of a protospacer adjacent motif (PAM) at the selected genomic locus, and is limited to a window within the CRISPR-Cas R-loop where single stranded (ss)DNA is accessible to the deaminase. Here, we reason that the Cas9-HNH nuclease domain sterically constrains ssDNA accessibility, and demonstrate that omission of this domain expands the editing window. By exchanging the HNH nuclease domain with an adenosine deaminase, we furthermore engineer adenine base editor variants (HNHx-ABE) with PAM-proximally shifted editing windows. HNHx-ABEs are substantially reduced in size, and expand the targeting scope of base editors. Our finding that the HNH domain is replaceable could moreover benefit future protein engineering efforts, where Cas9 operates together with other enzyme domains.
2020-11-12 | GSE161380 | GEO
Project description:Base editors are RNA-guided deaminases that enable site-specific nucleotide transitions. The targeting scope of these Cas-deaminase fusion proteins critically depends on the availability of a protospacer adjacent motif (PAM) at the selected genomic locus, and is limited to a window within the CRISPR-Cas R-loop where single stranded (ss)DNA is accessible to the deaminase. Here, we reason that the Cas9-HNH nuclease domain sterically constrains ssDNA accessibility, and demonstrate that omission of this domain expands the editing window. By exchanging the HNH nuclease domain with an adenosine deaminase, we furthermore engineer adenine base editor variants (HNHx-ABE) with PAM-proximally shifted editing windows. HNHx-ABEs are substantially reduced in size, and expand the targeting scope of base editors. Our finding that the HNH domain is replaceable could moreover benefit future protein engineering efforts, where Cas9 operates together with other enzyme domains.
2020-11-12 | GSE161379 | GEO