Search results for "Guide RNA"
showing 4 items of 4 documents
Engineering CRISPR guide RNA riboswitches for in vivo applications
2019
CRISPR-based genome editing provides a simple and scalable toolbox for a variety of therapeutic and biotechnology applications. Whilst the fundamental properties of CRISPR proved easily transferable from the native prokaryotic hosts to eukaryotic and multicellular organisms, the tight control of the CRISPR-editing activity remains a major challenge. Here we summarise recent developments of CRISPR and riboswitch technologies and recommend novel functionalised synthetic-gRNA (sgRNA) designs to achieve inducible and spatiotemporal regulation of CRISPR-based genetic editors in response to cellular or extracellular stimuli. We believe that future advances of these tools will have major implicati…
The RNA methyltransferase Dnmt2 methylates DNA in the structural context of a tRNA
2016
The amino acid sequence of Dnmt2 is very similar to the catalytic domains of bacterial and eukaryotic DNA-(cytosine 5)-methyltransferases, but it efficiently catalyzes tRNA methylation, while its DNA methyltransferase activity is the subject of controversial reports with rates varying between zero and very weak. By using composite nucleic acid molecules as substrates, we surprisingly found that DNA fragments, when presented as covalent DNA-RNA hybrids in the structural context of a tRNA, can be more efficiently methylated than the corresponding natural tRNA substrate. Furthermore, by stepwise development of tRNAAsp, we showed that this natural Dnmt2 substrate could be engineered to employ R…
Improvement of baculovirus as protein expression vector and as biopesticide by CRISPR/Cas9 editing
2019
The clustered regularly interspaced short palindromic repeats (CRISPR) system?associated Cas9 endonuclease is a molecular tool that enables specific sequence editing with high efficiency. In this study, we have explored the use of CRISPR/Cas9 system for the engineering of baculovirus. We have shown that the delivering of Cas9-single guide RNA ribonucleoprotein (RNP) complex with or without DNA repair template into Sf21 insect cells through lipofection might be efficient to produce knockouts as well as knock-ins into the baculovirus. To evaluate potential application of our CRISPR/Cas9 method to improve baculovirus as protein expression vector and as biopesticide, we attempted to knockout se…
Investigating REPAIRv2 as a Tool to Edit CFTR mRNA with Premature Stop Codons
2020
Cystic fibrosis (CF) is caused by mutations in the gene encoding the transmembrane conductance regulator (CFTR) protein. Some CF patients are compound heterozygous or homozygous for nonsense mutations in the CFTR gene. This implies the presence in the transcript of premature termination codons (PTCs) responsible for a truncated CFTR protein and a more severe form of the disease. Aminoglycoside and PTC124 derivatives have been used for the read-through of PTCs to restore the full-length CFTR protein. However, in a precision medicine framework, the CRISPR/dCas13b-based molecular tool &ldquo