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Visualization of Endoplasmic Reticulum Localized mRNAs in Mammalian Cells
JoVE Journal
Biology
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JoVE Journal Biology
Visualization of Endoplasmic Reticulum Localized mRNAs in Mammalian Cells
DOI:

10:24 min

December 17, 2012

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Chapters

  • 00:05Title
  • 01:18Preparation of Cells and Treatment with Translation Inhibitors
  • 02:29Digitonin Extraction and Cell Fixation
  • 04:47Fluorescent In situ Hybridization
  • 06:21Washing and Mounting the Stained Samples
  • 08:08Placental Alkaline Phosphatase and Cytochrome P450-8B1 mRNA ER-association
  • 09:50Conclusion

Summary

Automatic Translation

Here we describe a method to visualize endoplasmic reticulum-associated mRNAs in mammalian tissue culture cells. This technique involves the selective permeabilization of the plasma membrane with digitonin to remove cytoplasmic contents followed by fluorescent in situ hybridization to detect either bulk poly(A) mRNA or specific transcripts.

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