US 12,378,612 B2
Oligonucleotide-based probes and methods for detection of microbes
James O. McNamara, Iowa City, IA (US); Katie R. Flenker, Iowa City, IA (US); Hyeon Kim, Iowa City, IA (US); Alexander R. Horswill, Iowa City, IA (US); Frank J. Hernandez, Iowa City, IA (US); Mark Behlke, Coralville, IA (US); Lingyan Huang, Coralville, IA (US); Richard Owczarzy, Coralville, IA (US); Elliot Burghardt, Iowa City, IA (US); and Karen Clark, Iowa City, IA (US)
Assigned to UNIVERSITY OF IOWA RESEARCH FOUNDATION, Iowa City, IA (US)
Filed by UNIVERSITY OF IOWA RESEARCH FOUNDATION, Iowa City, IA (US)
Filed on Oct. 5, 2021, as Appl. No. 17/494,626.
Application 16/794,677 is a division of application No. 15/117,414, granted, now 10,619,219, issued on Apr. 14, 2020, previously published as PCT/US2015/015062, filed on Feb. 9, 2015.
Application 17/494,626 is a continuation of application No. 16/794,677, filed on Feb. 19, 2020, granted, now 11,155,882.
Claims priority of provisional application 61/937,359, filed on Feb. 7, 2014.
Claims priority of provisional application 61/980,498, filed on Apr. 16, 2014.
Claims priority of provisional application 61/992,034, filed on May 12, 2014.
Prior Publication US 2022/0098648 A1, Mar. 31, 2022
This patent is subject to a terminal disclaimer.
Int. Cl. C12Q 1/689 (2018.01); A61K 49/00 (2006.01); C07H 21/02 (2006.01); C07H 21/04 (2006.01); C12Q 1/70 (2006.01); G01N 21/64 (2006.01); G01N 33/58 (2006.01)
CPC C12Q 1/689 (2013.01) [A61K 49/0054 (2013.01); C07H 21/02 (2013.01); C07H 21/04 (2013.01); C12Q 1/701 (2013.01); G01N 21/6428 (2013.01); G01N 33/581 (2013.01); G01N 2021/6432 (2013.01)] 29 Claims
 
1. A method of detecting endonuclease activity in a test sample, comprising:
(a) contacting the test sample with a probe for detecting a microbial endonuclease comprising an oligonucleotide, a fluorophore operably linked to the oligonucleotide, and a quencher operably linked to the oligonucleotide, wherein the oligonucleotide comprises one or more modified nucleotides, is capable of being cleaved by a microbial nuclease, and has a DNA TT di-nucleotide, DNA AT di-nucleotide, DNA AA di-nucleotide or DNA TA di-nucleotide to form a digested probe, and
(b) measuring the fluorescence emitted by the digested probe.