“Parallel” and “Antiparallel Tail-Clamps” Increase the Efficiency of Triplex Formation with Structured DNA and RNA Targets

ChemBioChem ◽  
2005 ◽  
Vol 6 (6) ◽  
pp. 1034-1042 ◽  
Author(s):  
Anna Nadal ◽  
Ramon Eritja ◽  
Teresa Esteve ◽  
Maria Pla
Keyword(s):  
Triplex Formation ◽  
Dna And Rna ◽  
Rna Targets

2′-N-(Pyren-1-yl)acetyl-2′-Amino-α-L-LNA: Synthesis and Detection of Single Nucleotide Mismatches in DNA and RNA Targets

ChemBioChem ◽  
2007 ◽  
Vol 8 (10) ◽  
pp. 1122-1125 ◽  
Author(s):  
T. Santhosh Kumar ◽  
Jesper Wengel ◽  
Patrick J. Hrdlicka
Keyword(s):  
Single Nucleotide ◽  
Dna And Rna ◽  
Rna Targets

High Affinity Of Backbone-Modified α-Anomeric Oligodeoxynucleottoes For Dna And Rna Targets

1998 ◽  
Vol 17 (9-11) ◽  
pp. 1645-1649 ◽  
Author(s):  
A. Laurent ◽  
F. Debart ◽  
J.-C. Bologna ◽  
J.-J. Vasseur ◽  
B. Rayner
Keyword(s):  
High Affinity ◽  
Dna And Rna ◽  
Rna Targets

scholarly journals A programmable pAgo nuclease with universal guide and target specificity from the mesophilic bacterium Kurthia massiliensis

2021 ◽  
Author(s):  
Ekaterina Kropocheva ◽  
Anton Kuzmenko ◽  
Alexei A. Aravin ◽  
Daria Esyunina ◽  
Andrey Kulbachinskiy
Keyword(s):  
Nucleic Acid ◽  
Nucleic Acid Detection ◽  
Bacterial Cells ◽  
Dna And Rna ◽  
Rna Targets ◽  
Wide Range ◽  
Mesophilic Bacterium ◽  
Strict Specificity ◽  
Programmable Nucleases

ABSTRACTArgonaute proteins are programmable nucleases that are found in both eukaryotes and prokaryotes and provide defense against invading genetic elements. Although some prokaryotic Argonautes (pAgos) were shown to recognize RNA targets in vitro, the majority of studied pAgos have strict specificity toward DNA, which limits their practical use in RNA-centric applications. Here, we describe a unique KmAgo nuclease from the mesophilic bacterium Kurthia massiliensis that can be programmed with either DNA or RNA guides and can precisely cleave both DNA and RNA targets. KmAgo preferentially binds 16-20 nt long 5′-phosphorylated guide molecules with no strict specificity for their sequence and is active in a wide range of temperatures. In bacterial cells, KmAgo is loaded with small DNAs with no obvious sequence preferences suggesting that it can uniformly target genomic sequences. Target cleavage by KmAgo depends on the formation of secondary structure indicating that KmAgo can be used for structural probing of RNA targets. Mismatches between the guide and target sequences greatly affect the efficiency and precision of target cleavage, depending on the mismatch position and the nature of the reacting nucleic acid. These properties of KmAgo open the way for its use for highly specific nucleic acid detection and cleavage.

scholarly journals DNA and RNA editing without sequence limitation using the flap endonuclease 1 guided by hairpin DNA probes

2020 ◽  
Vol 48 (20) ◽  
pp. e117-e117
Author(s):  
Kun Tian ◽  
Yongjian Guo ◽  
Bingjie Zou ◽  
Liang Wang ◽  
Yun Zhang ◽  
...  
Keyword(s):  
Rna Editing ◽  
Human Cells ◽  
Hairpin Dna ◽  
Flap Endonuclease 1 ◽  
Dna And Rna ◽  
Rna Targets ◽  
Single Base Mismatch ◽  
Compact Size

Abstract Here, we characterized a flap endonuclease 1 (FEN1) plus hairpin DNA probe (hpDNA) system, designated the HpSGN system, for both DNA and RNA editing without sequence limitation. The compact size of the HpSGN system make it an ideal candidate for in vivo delivery applications. In vitro biochemical studies showed that the HpSGN system required less nuclease to cleave ssDNA substrates than the SGN system we reported previously by a factor of ∼40. Also, we proved that the HpSGN system can efficiently cleave different RNA targets in vitro. The HpSGN system cleaved genomic DNA at an efficiency of ∼40% and ∼20% in bacterial and human cells, respectively, and knocked down specific mRNAs in human cells at a level of ∼25%. Furthermore, the HpSGN system was sensitive to the single base mismatch at the position next to the hairpin both in vitro and in vivo. Collectively, this study demonstrated the potential of developing the HpSGN system as a small, effective, and specific editing tool for manipulating both DNA and RNA without sequence limitation.

Highly sensitive detection of rare mutant alleles by combining argonaute-based enrichment and XNA-PCR.

2019 ◽  
Vol 37 (15_suppl) ◽  
pp. e14605-e14605
Author(s):  
Jinzhao Song ◽  
Michael Joseph Powell ◽  
Wei Liu ◽  
Junman Chen ◽  
Haim Bau
Keyword(s):  
Nucleic Acids ◽  
High Efficiency ◽  
Elevated Temperatures ◽  
Thermus Thermophilus ◽  
Thermophilic Bacterium ◽  
Detection Methods ◽  
Single Nucleotide ◽  
Complementary Dna ◽  
Dna And Rna ◽  
Rna Targets

e14605 Background: Characterization of disease-associated, cell-free nucleic acids (liquid biopsy) provides a powerful, minimally-invasive means for early disease detection, genotyping, and personalized therapy. Detection of alleles of clinical interest is often challenged by their low concentration and sequence homology with the much more abundant wildtype nucleic acids. Methods: Argonuate (Ago) from the thermophilic bacterium Thermus thermophilus ( TtAgo) utilizes short DNA guides to specifically cleave complementary DNA and RNA targets. We found that under optimized conditions, TtAgo cleaves DNA and RNA complementary to the guide DNA with high efficiency, but spares nucleic acids with a single nucleotide mismatch at and around its catalytic site with high sensitivity. Based on these findings, we designed a new multiplexed enrichment assay, dubbed NAVIGATER (Nucleic Acid enrichment Via DNA Guided Argonaute from Thermus thermophilus), that utilizes TtAgo, to specifically cleave perfectly complementary DNA and RNA while sparing alleles of interest. Results: NAVIGATER greatly increases the fractions of rare mutant alleles with single nucleotide precision enhancing the sensitivity of downstream detection methods such as XNA-PCR. We demonstrate 60-fold enrichment of KRAS G12D in blood samples from pancreatic cancer patients and over ten-fold improved sensitivity of XNA-PCR, enabling multiplex detection of KRAS and EGFR mutants at 0.01% fractions. Conclusions: NAVIGATER has important advantages over other mutant allele enrichment assays such as the ones based on CRISPR-Cas. It does not require the target to contain a protospacer-adjacent motif; is a true (turnover) catalyst; can cleave both DNA and associated exosomal RNA targets, improving sensitivity; and can operate at elevated temperatures for higher selectivity and compatibility with detection schemes.

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