The invention relates to compositions and methods for making and use of a real-time cellular sensor. Components of a multipart enzyme are sequestered in different cellular compartments and only come together after receptor activation; a pool of substrate is made available in the cell to ensure real-time enzymatic output.

Modified or mutant bacterial luciferases having improved activity, as compared to wild type or unmodified bacterial luciferases, are described. The modified or mutant bacterial luciferases display increased light production and/or slower signal decay. Employing these modified or mutant bacterial luciferases improve a luminescence reporter system assay by increasing the detection sensitivity, resulting in improved bioreporter/reporter assays. The invention further provides methods for using the modified or mutant bacterial luciferases, reporter assays using the modified or mutant bacterial luciferases, and kits and articles of manufacture.

Cells, including stem cells, comprising an autobioluminescent phenotype, wherein the cells emit a luminescent signal in the absence of an exogenous luminescent stimulator, are provided. The luminescent signal may be constitutive, inducible, repressible, or tissue-specific. The cells express a synthetically engineered bacterial luciferase (lux) cassette, i.e., the luxCDABEfrp gene cassette. The cells may comprise luxA, luxB, luxC, luxD, luxE, and flavin reductase. The cells may each express a combined expression level of luxC, luxD, luxE, and flavin reductase that is from ten to forty times greater than a combined expression level of luxA and luxB. Further, methods of making and using the cells comprising an autobioluminescent phenotype are disclosed herein.

The present disclosure relates to cells, including stem cells, comprising an autobioluminescent phenotype, wherein the cells emit a luminescent signal in the absence of an exogenous luminescent stimulator. The luminescent signal may be constitutive, inducible, repressible, or tissue-specific. The cells express a synthetically engineered bacterial luciferase (lux) cassette, i.e., the luxCDABEfrp gene cassette. The cells may comprise luxA, luxB, luxC, luxD, luxE, and flavin reductase. The cells may each express a combined expression level of luxC, luxD, luxE, and flavin reductase that is from ten to forty times greater than a combined expression level of luxA and luxB. Further, methods of making and using the cells comprising an autobioluminescent phenotype are disclosed herein.

Cells, including stem cells, comprising an autobioluminescent phenotype, wherein the cells emit a luminescent signal in the absence of an exogenous luminescent stimulator, are provided. The luminescent signal may be constitutive, inducible, repressible, or tissue-specific. The cells express a synthetically engineered bacterial luciferase (lux) cassette, i.e., the luxCDABEfrp gene cassette. The cells may comprise luxA, luxB, luxC, luxD, luxE, and flavin reductase. The cells may each express a combined expression level of luxC, luxD, luxE, and flavin reductase that is from ten to forty times greater than a combined expression level of luxA and luxB. Further, methods of making and using the cells comprising an autobioluminescent phenotype are disclosed herein.

Modified or mutant bacterial luciferases having improved activity, as compared to wild type or unmodified bacterial luciferases, are described. The modified or mutant bacterial luciferases display increased light production and/or slower signal decay. Employing these modified or mutant bacterial luciferases improve a luminescence reporter system assay by decreasing the detection sensitivity, resulting in improved bioreporter/reporter assays. The invention further provides methods for using the modified or mutant bacterial luciferases, reporter assays using the modified or mutant bacterial luciferases, and kits and articles of manufacture.

A system for stable expression of gene pathways in cell lines, methods of making cell lines with stable expression of gene pathways, and methods of using the same are disclosed herein. The system comprises a nucleic acid construct configured to encode at least two genes of a multigene pathway in a cell. The nucleic acid construct comprises a plurality of nucleic acid sequences, wherein the plurality of nucleic acid sequences comprises: a first nucleic acid sequence encoding at least one gene of the multigene pathway; a first protease recognition nucleic acid sequence encoding a protease recognition site; a first linker nucleic acid sequence encoding a linker region, wherein the linker region comprises a viral 2A peptide; and a second nucleic acid sequence encoding at least one gene of the multigene pathway.

Provided are methods for detecting and treating disease states, including cancer, diabetes, fibrosis, and autoimmune diseases, by detecting increased mechanical modulus, or stiffness, or targeting tissues having increased mechanical modulus, or stiffness. Practicing these methods provides specific and localized detection assays and therapies for these disease states. Provided are mechano-responsive cell systems that can selectively detect and treat cancer metastases by targeting the unique biophysical and mechanical properties in the tumor microenvironment. Provided are methods for making mechano-sensitive CAR T cells by using mechano-responsive promoter logic. Provided are blood tests using engineered stem cells that express reporters after the cells home to a specific niche and secrete the reporter into the blood, which can be then be detected with a blood test. In alternative embodiments, provided are ultrasensitive detection platforms, able to detect target molecules or cells in blood with single-molecule or single-cell sensitivity.

The present disclosure relates compositions and methods of engineering microbial strains to detect and quantify glycan molecules.