Homo sapiens
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1307969 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:Bridge recombinases from the IS110 family of transposons, such as IS621, associate with a bridge RNA (bRNA) to mediate programmable recombination between donor and target DNAs. While insertion is mediated by the recombinase–bRNA complex, it remains unknown how these elements are excised from genomes to form circular intermediates. Here, we show that bRNA is weakly expressed from IS621 loci in the Escherichia coli genome and that the IS621 recombinase–bRNA complex mediates excision less efficiently than insertion. Furthermore, we determine the cryo-electron microscopy structures of the IS621 recombinase–bRNA complex bound to excision DNA substrates, providing mechanistic insights into the excision reaction. Similar to the previously reported donor- and target-bound insertion complex, the excision complex comprises two recombinase dimers, each accommodating target- and donor-binding loops of the bRNA. However, DNA recognition differs notably between the two complexes. While the donor and target DNA form a bent U-shape during insertion, the excision substrates adopt linear conformations and bind across both bRNA loops, forming an X-shaped structure, which reduces the efficiency of top-strand exchange and contributes to the naturally observed bias in reaction direction against excision. Despite such differences, the efficiencies of both reactions are similarly modulated by base pairing between the bRNA handshake guides and the top strands of DNA. Overall, this study provides mechanistic insights into the complete IS110 transposition cycle, and facilitates the optimal design of programmable bridge editing applications.
2026-08-12 | GSE328474 | GEO
Project description:Chromosomal rearrangements including large DNA-fragment inversions, deletions, and duplications by Cas9 with paired sgRNAs are important to investigate structural genome variations and developmental gene regulation, but little is known about the underlying mechanism. Here we report that disrupting CtIP or FANCD2, which is thought to function in NHEJ, enhances precise DNA-fragment deletion. In addition, by analyzing the inserted nucleotides at the junctions of DNA-fragment deletions, inversions, duplications, and characterizing the cleaved products, we find that Cas9 endonucleolytically cleaves the noncomplementary strand with a flexible scissile profile upstream of -3 position of the PAM site in vivo and in vitro, generating overhanged DSB ends. Moreover, we find that engineered Cas9 nucleases have distinct cleavage profiles. Finally, Cas9-mediated nucleotide insertions are nonrandom and are equal to the combined sequences upstream of both PAM sites with predicted frequencies. Thus, precise and predictable DNA-fragment editing could be achieved by perturbing DNA repair genes and using appropriate PAM configurations. These findings have important implications regarding 3D chromatin folding and enhancer insulation during gene regulation.
2018-07-26 | GSE113698 | GEO
Project description:Improvement of precise large-fragment deletion with a versatile nuclease prime editor
| PRJNA1184455 | ENA
Project description:Improvement of precise large-fragment deletion with a versatile nuclease prime editor
| PRJNA1483649 | ENA