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Reporter Genes

Reporter Genes

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.

Cell Specific Gene Expression

Cell Specific Gene Expression

Multicellular organisms contain a variety of structurally and functionally distinct cell types, but the DNA in all the cells originated from the same parent cells. The differences in the cells can be attributed to the differential gene expression. Liver cells, whose functions include detoxification of blood, production of bile to metabolize fats, and synthesis of proteins essential for metabolism, must express a specific set of genes to perform their functions. Gene expression also varies with...

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Transient Gene Expression in Tobacco using Gibson Assembly and the Gene Gun

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Transient gene expression in electroporated Picea glauca protoplasts.

F Bekkaoui¹, M Pilon, E Laine

¹Plant Biotechnology Institute, 110 Gymnasium Road, Saskatoon, S7N OW9, Saskatoon, Canada.

Plant Cell Reports

|November 19, 2013

Summary

White spruce protoplasts were successfully transformed using electroporation with a chloramphenicol acetyltransferase (CAT) reporter gene. Linearized DNA and specific voltage pulses optimized CAT gene expression for plant transformation studies.

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In vivo Electroporation of Developing Mouse Retina

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Published on: June 24, 2011

Area of Science:

Plant Biotechnology
Molecular Biology
Genetics

Background:

Efficient gene delivery into plant cells is crucial for genetic modification.
Electroporation is a common physical method for introducing DNA into plant protoplasts.
Reporter genes are essential tools for assessing transformation efficiency.

Purpose of the Study:

To optimize electroporation parameters for transient gene expression in white spruce protoplasts.
To evaluate the effectiveness of different plasmid constructs (linear vs. circular) for gene delivery.
To assess the suitability of reporter genes, specifically chloramphenicol acetyltransferase (CAT) and β-glucuronidase (GUS), for transformation studies in white spruce.

Main Methods:

Introduction of the chloramphenicol acetyltransferase (CAT) reporter gene into white spruce (Picea glauca) protoplasts via electroporation.
Optimization of plasmid concentration (pCaMVCN) and applied voltage (350V.cm(-1) with a 105ms decay constant).
Comparison of transient gene expression levels between linearized and circular plasmid constructs and assessment of β-glucuronidase (GUS) activity.

Main Results:

Transient CAT gene expression in white spruce protoplasts was significantly influenced by plasmid concentration and voltage.
Optimal CAT activity was achieved with a specific voltage pulse (350V.cm(-1), 105ms decay constant).
Linearized plasmid DNA resulted in substantially higher CAT activity compared to circular DNA, indicating improved transformation efficiency.
High background β-glucuronidase-like activity in protoplasts, potentially masking true transformation events, was observed.

Conclusions:

Electroporation is a viable method for introducing reporter genes into white spruce protoplasts.
Optimized voltage and linearized DNA constructs enhance transient gene expression efficiency.
The native β-glucuronidase-like activity in white spruce protoplasts presents a challenge for using GUS as a reliable reporter gene in this species.