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Måling af BK-Polyomavirus ikke-kodende kontrol region drevet Transkriptional aktivitet via flowcytometri
JoVE Journal
Genetics
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JoVE Journal Genetics
Measurement of BK-polyomavirus Non-Coding Control Region Driven Transcriptional Activity Via Flow Cytometry
DOI:

11:54 min

July 13, 2019

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Chapters

  • 00:00Title
  • 01:15Collection of Blood and or Urine Samples and Isolation of BKPyV-DNA
  • 02:11Amplification and Sequencing of the Non-coding Control Region (NCCR)
  • 03:37Cloning of the Non-coding Control Region (NCCR) into the Dual Fluorescence Reporter
  • 05:26Transient Transfection of HEK293T Cells with the Dual Fluorescence Reporter Plasmid and Treatment with Potential Antiviral Agents
  • 07:23Fluorescence Microscopy and Flow Cytometry
  • 09:22Results and Data Interpretation
  • 11:26Conclusion

Summary

Automatic Translation

I dette manuskript er en protokol præsenteret for at udføre FACS-baserede måling af BK-Polyomavirus transkriptional aktivitet ved hjælp af HEK293T celler transficeret med en tovejs reporter plasmid udtrykker tdtomato og egfp. Denne metode giver yderligere mulighed for at kvantitativt bestemme indflydelsen af nye forbindelser på viral transkriptionen.

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