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HSF1-related Assays (Heat shock)

The LightSwitch™ Validated Heat Shock (HSF) Pathway Collection is a set of sequence-verified, transfection-ready, promoter reporter constructs that can be used to accurately quantify transcriptional activation in response to induction of the heat shock pathway. These constructs are a subset of the 18,000-member LightSwitch Promoter Reporter GoClone™ Collection that were selected based on sequence motif analysis, published information and in-house analysis of each promoter's activity* using our LightSwitch Luciferase Assay. The Heat Shock Biomarker Set contains 8 of the highest responding promoters under our chosen test conditions, as well as the promoters of 2 housekeeping genes (Figure 1). Active Motif also offers a HSF Pathway Reporter Stable Cell Line to facilitate high-throughput screening of the heat shock pathway.

Graph showing the induction ratios of the promoter reporter constructs of the Heat Shock Biomarker Set after transfection into HT1080 human fibrosarcoma cells and induction by growing the cells at 43 degrees.
Figure 1: Induction ratios for each member of the Heat Shock Biomarker Set.

HT1080 human fibrosarcoma cells were transfected with the promoter reporter constructs shown. After 12 hours, three replicates of each transfected construct were moved to 43°C while three replicates were grown as normal (kept at 37°C). After 8 hours at their respective temperatures, each treatment for each reporter construct was assayed in triplicate using the LightSwitch Luciferase Assay Kit. The graph shows the ratio of the average values for each treated/untreated signal on a log scale; error bars represent one standard deviation.

*Experimental results and the absolute values of luciferase units may vary depending on cell line, treatment and protocols. All data shown is solely for use as a guide for selecting candidate promoters and panels for your particular experimental design.

 
Name Format Cat No. Price  
LightSwitch™ Heat Shock (HSF1) Biomarker Set 1 set 32086 $3,500 Buy Now

Heat Shock (HSF) Pathway

The regulation of heat-shock genes is critical in cell stress response and the maintenance of cellular homeostasis. Heat-shock gene transcription is regulated by HSF (heat shock factor) transcription factors in response to various types of environmental stress such as heat, starvation, hypoxia, infection and toxin exposure. Heat shock proteins (HSPs) are also well-characterized for their roles as chaperones for protein synthesis, assembly, secretion and degradation. As specific HSPs have been implicated as central players in apoptosis, tumor survival, and immune response, understanding the regulation of the heat shock response is critical.

Applications

The collection of validated LightSwitch heat shock-related promoter reporters enables you to:

  • Understand the mechanisms by which heat shock regulated genes are induced or repressed
  • Quantify the functional consequences of transcription factor binding – while many other technologies give a qualitative view of transcription factor level, our promoter reporter assays quantify the effects of transcription factor binding
  • Confirm data from ChIP, ChIP-chip or ChIP-Seq experiments
  • Measure the effect of sequence variants and mutagenesis on promoter function
  • Screen for promoter activation or develop activity profiles across the heat shock response pathway for many compounds or conditions in parallel

LightSwitch Luciferase Assay System

The LightSwitch Luciferase Assay System is a complete solution for studying transcriptional & post-transcriptional gene regulation in living mammalian cells. It enables you to directly measure the functional activity of promoters and 3´UTRs through use of an engineered luciferase gene (RenSP) and optimized assay & transfection reagents that ensure superior, reproducible results. In addition to Validated Pathway Collections, the LightSwitch System includes collections of over 30,000 cloned human promoter reporter constructs and human 3´UTR reporter constructs, miRNA mimics and inhibitors, stable cell lines, as well as collections of synthetic long-range response element constructs and synthetic 3´UTRs target validation constructs that can provide more sensitivity than endogenous sequences. Comprehensive Custom Services are also available

Diagram of the LightSwitch Luciferase Promoter Reporter Assay.
LightSwitch Promoter Reporter Assays.

To study a promoter's regulation, it is cloned into a LightSwitch Reporter vector upstream of the RenSP luciferase gene and then transfected into mammalian cells. The cells are stimulated, if required, to induce transcription of RenSP luciferase from the cloned promoter. Promoter activity is then quantified using a LightSwitch Luciferase Assay Kit and a luminometer to measure the amount of luminescence produced.

Advantages of the LightSwitch Luciferase Assay System

  • Quantitative – Novel RenSP luciferase technology allows you to measure promoter activity with industry-leading sensitivity and dynamic range.
  • Simple, fast, complete solution – With pre-cloned LightSwitch Reporter vectors and optimized transfection & assay reagents, you can study regulation of your gene today. No cloning, DNA preparation or optimization is needed, and most studies do not require any internal transfection controls.
  • Comprehensive and verified – The genome-wide LightSwitch Reporter Collections are sequence-verified, transfection-ready promoter and 3´UTR reporter vectors.
  • Functionally insightful – Learn about the actual effects of transcription factor binding to verify computational predictions and supplement microarray or sequencing data.
  • Cost-effective – Efficiently screen for activation and/or repression using a multitude of conditions.

The Heat Shock Biomarker Set contains 5 µg aliquots each of 8 promoter reporter constructs and 2 housekeeping promoter constructs (ACTB & RPL10) for use as controls. Click on the links below for clone information.