US 11,898,211 B2
Biosensors for detecting and/or neutralizing bioavailable uranium and related U-sensitive genetic molecular components, gene cassettes, vectors, genetic circuits, compositions, methods and systems
Dan Mcfarland Park, Dublin, CA (US)
Assigned to LAWRENCE LIVERMORE NATIONAL SECURITY, LLC, Livermore, CA (US)
Filed by LAWRENCE LIVERMORE NATIONAL SECURITY, LLC, Livermore, CA (US)
Filed on Feb. 4, 2020, as Appl. No. 16/781,950.
Claims priority of provisional application 62/801,077, filed on Feb. 4, 2019.
Prior Publication US 2020/0248277 A1, Aug. 6, 2020
This patent is subject to a terminal disclaimer.
Int. Cl. C12Q 1/6897 (2018.01); C12N 1/20 (2006.01); C12N 15/52 (2006.01); C12N 9/12 (2006.01); C12N 15/63 (2006.01); G21F 9/18 (2006.01); G21F 9/30 (2006.01); C12N 15/74 (2006.01)
CPC C12Q 1/6897 (2013.01) [C12N 1/20 (2013.01); C12N 9/12 (2013.01); C12N 15/52 (2013.01); C12N 15/635 (2013.01); C12N 15/74 (2013.01); C12Y 207/13003 (2013.01); G21F 9/18 (2013.01); G21F 9/30 (2013.01); C12N 2830/002 (2013.01); C12N 2830/55 (2013.01); C12N 2840/002 (2013.01); C12N 2840/105 (2013.01); G01N 2520/00 (2013.01)] 40 Claims
 
1. A UO2F2 biosensor comprising a genetically engineered bacterial cell capable of heterologously and/or natively expressing histidine kinase 1363, and U-sensitive transcriptional regulator 1362;
wherein the bacterial cell is an engineered bacterial cell including a 1362 U-sensing genetic reportable molecular component and/or a 1362 U-sensing/U-neutralizing genetic molecular component, each comprising a U-sensitive promoter in a configuration wherein the U-sensitive promoter directly initiates expression of the U-sensing reportable molecular component and/or of the U-sensing U-neutralizing molecular component in presence of bioavailable U;
wherein the U-sensitive promoter is
a 1362 U-sensitive prompter comprising a U-sensitive transcriptional 1362 binding site having a DNA sequence
(SEQ ID NO: 1)
N1N2N3N4N5N6N7N8N9N10N11N12N13N14N15N16N17N18,
in which
N1 is C or T;
N2 is G or A;
N3 is T or C;
N4 is C;
N5 is A or G;
N6 is G or C;
N7 C or G;
N8 is any nucleotide;
N9 is any nucleotide;
N10 is any nucleotide;
N11 is any nucleotide;
N12 is T or C;
N13 is G;
N14 is T or C;
N15 is C;
N16 is A or C;
N17 is G; and
N18 is C or G,
and in which N1 to N17 are selected independently
and wherein the genetically modified bacterial cell is an engineered bacterial cell further comprising an F-sensing riboswitch within at least one of
the U-sensing reportable genetic molecular component in a configuration wherein the U-sensing reportable genetic molecular component, is transcribed in presence of an effective amount of bioavailable fluoride,
an F-sensing reportable genetic molecular component in a configuration wherein the F-sensing reportable genetic molecular component is transcribed in presence of an effective amount of bioavailable fluoride, and
an F sensitive genetic circuit in which at least one molecular component is a reportable molecular component (and in particular one or more reportable genetic molecular component and/or one or more reportable cellular molecular component), the reportable molecular component expressed when the genetic circuit operates according to the circuit design in presence of bioavailable F.