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Reporter genes are a type of protein-coding gene that are often tagged to a gene of interest. Once inside a target cell, reporter genes usually produce visually identifiable characteristics like fluorescence and luminescence when expressed along with the gene of interest. Thus, reporter genes “report” the presence or absence of genes of interest in an organism, determine the gene expression pattern, or track the physical location of a DNA segment or protein in the cell.
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In vivo Application of the REMOTE-control System for the Manipulation of Endogenous Gene Expression
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Customizable gene sensing and response without altering endogenous coding sequences.

Fabio Caliendo1,2, Elvira Vitu1,2, Junmin Wang3

  • 1Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.

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Researchers developed a novel method to integrate genetic biosensors into cells without altering gene coding sequences. This synthetic biology approach enhances cellular control for applications in biomanufacturing and therapeutics.

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Area of Science:

  • Synthetic biology
  • Genetic engineering
  • Cellular engineering

Background:

  • Synthetic biology utilizes genetic circuits to control cellular functions.
  • Integrating biosensors into endogenous genes via CRISPR-Cas9 allows gene expression monitoring but can disrupt native regulation.
  • A method is needed to embed biosensors without impacting gene coding sequences.

Purpose of the Study:

  • To develop a novel method for integrating genetic biosensors into endogenous genes without altering their coding sequences.
  • To enable precise control and prediction of biosensor responses using mathematical modeling.
  • To engineer a functional cell stress sensor and actuator system in CHO-K1 cells.

Main Methods:

  • Engineered a system to insert single-guide RNAs into the terminator regions of endogenous genes, activating downstream circuits.
  • Developed a mathematical model to predict and fine-tune sensor dosage responses.
  • Constructed a cell stress sensor and actuator in CHO-K1 cells to conditionally activate the antiapoptotic protein BCL-2.

Main Results:

  • Successfully integrated genetic biosensors into endogenous genes without modifying coding sequences.
  • Demonstrated fine-tuning and prediction of sensor dosage responses using a mathematical model.
  • Engineered a functional cell stress sensor and actuator that enhances cell survival under stress by activating BCL-2.

Conclusions:

  • The developed platform allows for the integration of genetic biosensors into endogenous genes by targeting terminator regions.
  • This approach avoids disruption of native gene coding sequences and allows for predictable biosensor responses.
  • The engineered cell stress sensor and actuator system shows potential for biomanufacturing, cell fate control, and cell-based therapeutics.