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Hypoxia-Induced Reporter Genes with Different Half-Lives

  • Balaji KrishnamacharyEmail author
  • Pierre Danhier
  • Samata Kakkad
  • Santosh K. Bharti
  • Zaver M. Bhujwalla
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1790)

Abstract

The utility of reporter genes has gained significant momentum over the last three decades. Reporter genes are used to understand the transcriptional activity of a gene both in vitro and in vivo, and in pathway analysis and drug screening for diseases involving protozoan parasites, and in anti-cancer drug developments. Here, using a human prostate cancer xenograft model (PC3), we describe a method to construct and validate hypoxia reporter genes with different half-lives. Using molecular biology and optical imaging techniques, we have validated the expression of long half-life enhanced green fluorescence protein (EGFP) expression and short half-life luciferase gene expression to report on the spatial and temporal evolution of hypoxia in vivo.

Key words

Bioluminescence Lentivirus Luciferase assay Hypoxia Hypoxia response elements (HRE) Reporter gene 

Notes

Acknowledgments

This work was supported by National Institutes of Health R01 CA73850, R01 CA82337, and P50 CA103175.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Balaji Krishnamachary
    • 1
    Email author
  • Pierre Danhier
    • 1
  • Samata Kakkad
    • 1
  • Santosh K. Bharti
    • 1
  • Zaver M. Bhujwalla
    • 1
    • 2
  1. 1.JHU ICMIC Program, Division of Cancer Imaging Research, The Russell H. Morgan Department of Radiology and Radiological ScienceThe Johns Hopkins University School of MedicineBaltimoreUSA
  2. 2.Sidney Kimmel Comprehensive Cancer CenterThe Johns Hopkins University School of MedicineBaltimoreUSA

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