Modified Oligonucleotides for Guiding RNA Cleavage Using Bacterial RNase P

Новопашина, Д. С.; Nazarov, Anton; Vorobjeva, M. A.; Kuprushkin, Maxim; Davydova, Anna; Lomzov, Alexander A.; Pyshnyi, D. V.; Altman, Sidney; Venyaminova, Alya G.

doi:10.1134/s0026893318060134

Cited by 5 publications

(2 citation statements)

References 22 publications

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“…Nuclease resistance is one of the essential characteristics determining the possibility of in vitro and in vivo applications for therapeutic nucleic acids. Model serum-containing solutions are widely used for fast and convenient testing of stability of modified oligonucleotides in biological media [ 22 ].…”

Section: Resultsmentioning

confidence: 99%

Improving Stability and Specificity of CRISPR/Cas9 System by Selective Modification of Guide RNAs with 2′-fluoro and Locked Nucleic Acid Nucleotides

Sakovina

Vokhtantsev

Vorobyeva

et al. 2022

IJMS

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The genome editing approach using the components of the CRISPR/Cas system has found wide application in molecular biology, fundamental medicine and genetic engineering. A promising method is to increase the efficacy and specificity of CRISPR/Cas-based genome editing systems by modifying their components. Here, we designed and chemically synthesized guide RNAs (crRNA, tracrRNA and sgRNA) containing modified nucleotides (2’-O-methyl, 2’-fluoro, LNA—locked nucleic acid) or deoxyribonucleotides in certain positions. We compared their resistance to nuclease digestion and examined the DNA cleavage efficacy of the CRISPR/Cas9 system guided by these modified guide RNAs. The replacement of ribonucleotides with 2’-fluoro modified or LNA nucleotides increased the lifetime of the crRNAs, while other types of modification did not change their nuclease resistance. Modification of crRNA or tracrRNA preserved the efficacy of the CRISPR/Cas9 system. Otherwise, the CRISPR/Cas9 systems with modified sgRNA showed a remarkable loss of DNA cleavage efficacy. The kinetic constant of DNA cleavage was higher for the system with 2’-fluoro modified crRNA. The 2’-modification of crRNA also decreased the off-target effect upon in vitro dsDNA cleavage.

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Section: Resultsmentioning

confidence: 99%

Improving Stability and Specificity of CRISPR/Cas9 System by Selective Modification of Guide RNAs with 2′-fluoro and Locked Nucleic Acid Nucleotides

Sakovina

Vokhtantsev

Vorobyeva

et al. 2022

IJMS

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“…We chose two most effective modified oligonucleotide inhibitors m-inh (IC 50 = 90 nM) and pgd-inh3 (IC 50 = 100 nM) for further studies. These oligomers combine RNase P inhibiting activity with good stability in biological media (Novopashina et al, 2018) (see Figure S2 ), so we consider them as a prospective basis for the development of antibacterial agents. Of note, the high level of structural and functional conservation of bacterial RNase P (Altman, 2011) permits to extrapolate the regularities obtained for E.coli enzyme to the other bacterial species, particularly A. baumannii (Davies-Sala et al, 2018).…”

Section: Resultsmentioning

confidence: 99%

Novel Peptide Conjugates of Modified Oligonucleotides for Inhibition of Bacterial RNase P

et al. 2019

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Novel alternatives to traditional antibiotics are now of great demand for the successful treatment of microbial infections. Here, we present the engineering and properties of new oligonucleotide inhibitors of RNase P, an essential bacterial enzyme. The series of 2’- O -methyl RNA (2’-OMe-RNA) and phosphoryl guanidine oligonucleotides were targeted to the substrate-binding region of M1 RNA subunit of the RNase P. Uniformly modified 2’-OMe RNA and selectively modified phosphoryl guanidine oligonucleotides possessed good stability in biological media and effectively inhibited RNase P. Their conjugates with transporting peptides were shown to penetrate bacterial cells ( Escherichia coli and Acinetobacter baumannii ) and inhibit bacterial growth.

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Structure and hybridization properties of phosphoryl guanidine oligonucleotides under crowding conditions

Kanarskaya

Golyshev

Pyshnyi

et al. 2021

Biochemical and Biophysical Research Communications

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Modified Oligonucleotides for Guiding RNA Cleavage Using Bacterial RNase P

Cited by 5 publications

References 22 publications

Improving Stability and Specificity of CRISPR/Cas9 System by Selective Modification of Guide RNAs with 2′-fluoro and Locked Nucleic Acid Nucleotides

Improving Stability and Specificity of CRISPR/Cas9 System by Selective Modification of Guide RNAs with 2′-fluoro and Locked Nucleic Acid Nucleotides

Novel Peptide Conjugates of Modified Oligonucleotides for Inhibition of Bacterial RNase P

Structure and hybridization properties of phosphoryl guanidine oligonucleotides under crowding conditions

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