title: Tandem Oligonucleotide Probe Annealing and Elongation To Discriminate Viral Sequence.
abstract: New approaches for genomic DNA/RNA detection are in high demand in order to provide controls for existing enzymatic technologies and to create alternatives for emerging applications. In particular, there is an unmet need in rapid, reliable detection of short RNA regions which could open up new opportunities in transcriptome analysis, virology, and other fields. Herein, we report for the first time a "click" chemistry approach to oligonucleotide probe elongation as a novel approach to specifically detect a viral sequence. We hybridized a library of short, terminally labeled probes to Ebola virus RNA followed by click assembly and analysis of the read sequence by various techniques. As we demonstrate in this paper, using our new approach, a viral RNA sequence can be detected in less than 2 h without the need for cDNA synthesis or any other enzymatic reactions and with a sensitivity of <10 pM target RNA.
authors.affiliation: Department of Physics, Chemistry and Pharmacy, University of Southern Denmark , Campusvej 55, 5230 Odense M, Denmark.
authors.forename: Maria
authors.identifier: 0000-0002-9727-2496
authors.initials: M
authors.lastname: Taskova
authors.affiliation: Department of Physics, Chemistry and Pharmacy, University of Southern Denmark , Campusvej 55, 5230 Odense M, Denmark.
authors.forename: Jesper
authors.initials: J
authors.lastname: Uhd
authors.affiliation: Division of Oncology, Department of Medicine, Stanford University , 269 Campus Drive, Stanford, California 94305, United States.
authors.forename: Laura
authors.initials: L
authors.lastname: Miotke
authors.affiliation: Division of Oncology, Department of Medicine, Stanford University , 269 Campus Drive, Stanford, California 94305, United States.
authors.forename: Matthew
authors.initials: M
authors.lastname: Kubit
authors.affiliation: Stanford Genome Technology Center, Stanford University , 3165 Porter Drive, Palo Alto, California 94304, United States.
authors.forename: John
authors.initials: J
authors.lastname: Bell
authors.affiliation: Division of Oncology, Department of Medicine, Stanford University , 269 Campus Drive, Stanford, California 94305, United States.|Stanford Genome Technology Center, Stanford University , 3165 Porter Drive, Palo Alto, California 94304, United States.
authors.forename: Hanlee P
authors.initials: HP
authors.lastname: Ji
authors.affiliation: Department of Physics, Chemistry and Pharmacy, University of Southern Denmark , Campusvej 55, 5230 Odense M, Denmark.
authors.forename: Kira
authors.identifier: 0000-0003-4878-0301
authors.initials: K
authors.lastname: Astakhova
mesh_terms: D002232:Carbocyanines / Q000737:chemistry; D057930:Click Chemistry; D016002:Discriminant Analysis; D029043:Ebolavirus / Q000235:genetics*; D009693:Nucleic Acid Hybridization; D015345:Oligonucleotide Probes / Q000235:genetics / Q000378:metabolism*; D009841:Oligonucleotides / Q000737:chemistry; D020641:Polymorphism, Single Nucleotide; D012367:RNA, Viral / Q000032:analysis / Q000378:metabolism*
chemical_list: D002232:Carbocyanines; D015345:Oligonucleotide Probes; D009841:Oligonucleotides; D012367:RNA, Viral; C085321:cyanine dye 5; C477371:locked nucleic acid