Excellent, Fast Assay for Measuring Promoter Activity

September 19, 2017

University of Kansas Medical Center
Pathology
Postdoctoral Fellow

Overall

Quality of Results

Ease-of-Optimization

What do these ratings mean?
Write a Review

Company:

SwitchGear Genomics

Product Name:

LightSwitch™ Transfection Optimization Kit

Catalog Number:

TFXOPT

This research is focused on identifying novel transcriptional targets of an oncoprotein, with a goal of developing novel therapeutics to treat cancer. In these assays, we determined whether knockdown of the oncoprotein affected promoter activity of our genes of interest in osteosarcoma cell lines. We chose this product because of the quick availability and because it does not require transfection of dual reporters, which was a concern as our cells are difficult to transfect.

Experimental Design and Results Summary

Application

Luciferase-based reporter assay

Starting Material

Human tumor cells

Protocol Overview

A tumor cell line was transfected with either the positive or negative promoter vectors to optimize transfection at the following condition (5,000 cells/well 3:1 vector:transfection reagent; 15,000 cells/well 3:1 vector:transfection reagent; 5,000 cells/well 6:1 vector:transfection reagent; 15,000 cells/well 6:1 vector:transfection reagent). Cells were incubated for 24 hours, the plates were evaluated per manufacturer's protocol.

Tips

It is helpful to measure cell viability alongside this assay to ensure there is no cell death.

Results Summary

We were able to optimize the assay with a very large signal to noise window.

DOI or PMID #

N/A

Additional Notes

N/A

Related Categories

Image Gallery

Summary

The Good

Assay is very easy to perform, data is easy to interpret. This is not a dual assay, so there is no built-in transfection control.

The Bad

The assay can be costly (reporter constructs are expensive), particularly if you're evaluating multiple cell lines.

The Bottom Line

The assay is great, and very quick. Easy to use for any scientific background or level of experience. Cost may be prohibitive.

Join the discussion