Lux expression in cells and methods of use

A kit for producing a stem cell having an autonomous luminescent phenotype, comprising:

• at least one vector comprising at least one of a luxA nucleic acid, a luxB nucleic acid, a luxC nucleic acid, a luxD nucleic acid, a luxE nucleic acid, nucleic acid, and a flavin-reductase nucleic acid.

A method for producing a stem cell having autonomous and constitutive luminescence, comprising:

• providing a stem cell;
• and
• transfecting the stem cell with at least one vector comprising at least one of a luxA nucleic acid, a luxB nucleic acid, a luxC nucleic acid, a luxD nucleic acid, a luxE nucleic acid, nucleic acid, and a flavin-reductase nucleic acid.

A method for producing a stem cell comprising an autonomous luminescent phenotype comprising a tissue-specific signal, comprising:

• providing a stem cell;
• and
• transfecting the stem cell with at least one vector comprising at least one of a luxA nucleic acid, a luxB nucleic acid, a luxC nucleic acid, a luxD nucleic acid, a luxE nucleic acid, nucleic acid, and a flavin-reductase nucleic acid, wherein at least one of the nucleic acids is operatively linked to a tissue-specific promoter.

A kit for producing a stem cell comprising an autonomous luminescent phenotype comprising a tissue-specific signal, comprising:

• at least one vector comprising at least one of a luxA nucleic acid, a luxB nucleic acid, a luxC nucleic acid, a luxD nucleic acid, a luxE nucleic acid, nucleic acid, and a flavin-reductase nucleic acid, wherein at least one of the nucleic acids is operatively linked to a tissue-specific promoter.

A kit for producing a stem cell emitting an autonomous luminescent signal inducible by an analyte, comprising:

• at least one vector comprising a luxA nucleic acid, a luxB nucleic acid, a luxC nucleic acid, a luxD nucleic acid, a luxE nucleic acid, and a flavin reductase nucleic acid, and wherein at least one of the luxA nucleic acid, the luxB nucleic acid, the luxC nucleic acid, the luxD nucleic acid, the luxE nucleic acid, and the flavin reductase nucleic acid are operatively linked to at least one analyte-responsive response element;
• and
• a second vector comprising at least one analyte-responsive reverse transactivator that, when exposed to the analyte, activates the at least one analyte-responsive response element.

A kit for producing a stem cell emitting an autonomous luminescent signal repressible by an analyte, comprising:

• at least one vector comprising a luxA nucleic acid, a luxB nucleic acid, a luxC nucleic acid, a luxD nucleic acid, a luxE nucleic acid, and a flavin reductase nucleic acid, and wherein at least one of the luxA nucleic acid, the luxB nucleic acid, the luxC nucleic acid, the luxD nucleic acid, the luxE nucleic acid, and the flavin reductase nucleic acid are operatively linked to at least one analyte-responsive response element;
• and
• a second vector comprising at least one analyte-responsive transactivator that, when exposed to the analyte, does not activate the at least one analyte-responsive response element.

A stem cell-derived autonomously luminescent cell, comprising:

• an autonomously luminescent eukaryotic cell differentiated from an autonomously luminescent stem cell, wherein the differentiated autonomously luminescent eukaryotic cell and the autonomously luminescent stem cell both express a constitutive luminescent signal in the absence of an exogenous luminescent stimulator.

A method for producing a stem cell-derived autonomously luminescent cell from an autonomously luminescent stem cell, comprising:

• constructing an autonomously luminescent stem cell, and
• differentiating the autonomously luminescent stem cell into the stem cell-derived autonomously luminescent cell, wherein the stem cell-derived autonomously luminescent cell emits a luminescent signal in the absence of an exogenous luminescent stimulator.

Any one of the preceding claims, wherein the stem cell-derived autonomously luminescent cell expresses luxA, luxB, luxC, luxD, luxE, and flavin reductase, such that the stem cell-derived autonomously luminescent cell luminesces in the absence of an exogenous luminescent stimulator.

Any one of the preceding claims, wherein a combined production level of luxC, luxD, luxE, and flavin reductase ranges from ten to forty times greater than a combined production level of luxA and luxB.

Any one of the preceding claims, wherein a combined production level of luxC, luxD, luxE, and flavin reductase ranges from twenty to thirty times greater than a combined production level of luxA and luxB.

Any one of the preceding claims, wherein one or more of the nucleic acids encoding each of luxA, luxB, luxC, luxD, luxE has a greater than 80% sequence identity with the corresponding nucleic acid in Photorhabdus luminescens.

Any one of the preceding claims, wherein one or more of the nucleic acids encoding each of luxA, luxB, luxC, luxD, luxE has a 100% sequence identity with the corresponding nucleic acid in Photorhabdus luminescens.

Any one of the preceding claims, wherein the stem cell emits the luminescent signal through transcription of the luxA nucleic acid, the luxB nucleic acid, the luxC nucleic acid, the luxD nucleic acid, the luxE nucleic acid, and the flavin reductase nucleic acid.

Any one of the preceding claims, wherein transcription levels of the luxC nucleic acid, the luxD nucleic acid, the luxE nucleic acid, and the flavin reductase nucleic acid range from ten to forty times greater than transcription levels of the luxA nucleic acid and the luxB nucleic acid.

Any one of the preceding claims, wherein transcription levels of the luxC nucleic acid, the luxD nucleic acid, the luxE nucleic acid, and the flavin reductase nucleic acid range from twenty to thirty times greater than transcription levels of the luxA nucleic acid and the luxB nucleic acid.

Any one of the preceding claims, wherein the nucleic acids encoding each of luxC, luxD, luxE, and flavin reductase are present in a combined level of from ten times to forty times a combined level of nucleic acids encoding luxA and luxB.

Any one of the preceding claims, wherein the nucleic acids encoding each of luxC, luxD, luxE, and flavin reductase are present in a combined level of from twenty times to thirty times a combined level of nucleic acids encoding luxA and luxB.

Any one of the preceding claims, wherein the stem cell or the at least one stem cell further comprises luxA, luxB, luxC, luxD, luxE, and flavin reductase.

Any one of the preceding claims, wherein the stem cell or the at least one stem cell expresses luxA, luxB, luxC, luxD, luxE, and flavin reductase.

Any one of the preceding claims, wherein a combined production level of luxC, luxD, luxE, and flavin reductase ranges from ten to forty times greater than a combined production level of luxA and luxB.

Any one of the preceding claims, wherein a combined production level of luxC, luxD, luxE, and flavin reductase ranges from twenty to thirty times greater than a combined production level of luxA and luxB.

Any one of the preceding claims, wherein at least one of luxC, luxD, luxE, and flavin reductase is present at a level greater than a level of at least one of luxA and luxB.

Any one of the preceding claims, wherein luxC, luxD, luxE, and flavin reductase are present at a combined level of from ten times to forty times greater than a combined level of luxA and luxB.

Any one of the preceding claims, wherein luxC, luxD, luxE, and flavin reductase are present at a combined level of from twenty times to thirty times greater than a combined level of luxA and luxB.

Any one of the preceding claims, wherein the stem cell or the at least one stem cell comprising luxA, luxB, luxC, luxD, luxE, and flavin reductase autonomously luminesces in the absence of an exogenous luminescent stimulator.

Any one of the preceding claims, wherein the constitutive promoter is a chicken beta-actin promoter.

Any one of the preceding claims, wherein the first constitutive promoter and the second constitutive promoter is a chicken beta-actin promoter.

Any one of the preceding claims, wherein at least one of the luxA nucleic acid, the luxB nucleic acid, the luxC nucleic acid, the luxD nucleic acid, the luxE nucleic acid, and the flavin reductase nucleic acid is operatively linked to at least one linker region.

Any one of the preceding claims, wherein the at least one linker region comprises a viral 2A peptide.

Any one of the preceding claims, wherein the agent comprises a chemotherapeutic agent, an antibiotic, an insecticide, a pesticide, an herbicide, or a fertilizer.

Any one of the preceding claims, wherein the stem cell or the at least one stem cell is an induced pluripotent stem cell, a mesenchymal stem cell, or a non-embryonic stem cell.

Any one of the preceding claims, wherein the exogenous luminescent stimulator is a fluorescent stimulation signal.

Any one of the preceding claims, wherein the exogenous luminescent stimulator is a chemical luminescent activator, and preferably wherein the chemical luminescent activator comprises an aldehyde functional group.