Devices and Methods for Detecting Microorganisms Using Recombinant Reproduction-Deficient Indicator Bacteriophage
OwnerStephen Erickson, Jose S. Gil +2inventors
InventorsStephen Erickson, Jose S. Gil, Minh Mindy Bao Nguyen, Wendy S. Hahn
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Abstract
Disclosed herein are compositions, methods, kits and systems for rapid detection of microorganisms using a reproduction-deficient indicator bacteriophage. The specificity of such reproduction-deficient indicator bacteriophage for binding and infecting particular microorganisms of interest allows targeted and sensitive detection of a microorganism of interest.
Claims20
A recombinant phage comprising an indicator gene in a late gene region of genome of the phage, wherein the recombinant phage is reproduction-deficient, and wherein the recombinant phage is capable of specifically infecting a microorganism of interest.
A composition comprising at least two recombinant phages, each comprising an indicator gene in a late gene region of genome of the phage, wherein the recombinant phages are reproduction-deficient, and wherein the recombinant phages are capable of specifically infecting one or more microorganism of interest.
A method of preparing a recombinant phage, comprising:
- selecting a parent phage that specifically infects a target microorganism;
- altering a gene of the parent page to generate a recombinant reproduction-deficient phage;
- transforming an engineered strain of the target microorganism capable of expressing a product of the gene mutated in the reproduction-deficient phage with a homologous recombination (HR) plasmid comprising an indicator gene and HR sequences flanking the indicator gene and homologous to a desired sequence in the parent phage;
- infecting the transformed target microorganism with the parent phage or the reproduction-deficient parent phage, allowing HR to occur between the HR plasmid and the genome or the parent phage or the recombinant reproduction-deficient phage;
- and
- isolating a particular clone of recombinant phage that is both reproduction-deficient and is capable of expressing a product of the indicator gene.
Dependencies are inferred from the claim text.