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Förstärkning, Nästa generations sekvensering, och Genomic DNA Kartläggning av retroviral integration webbplatser
JoVE Journal
Biologia
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JoVE Journal Biologia
Amplification, Next-generation Sequencing, and Genomic DNA Mapping of Retroviral Integration Sites
DOI:

09:31 min

March 22, 2016

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Capitoli

  • 00:05Titolo
  • 00:39Generation of Retroviral Stock
  • 02:20Viral Infection of HEK293T Cells
  • 04:30Genomic DNA Fragmentation
  • 05:33Ligation of Linker Oligonucleotide to Fragmented Genomic DNA
  • 06:58Amplification of Viral LTR-host Genomic DNA Junctions by Semi-nested PCR
  • 08:02Results: Sequence Logos Depicting HIV-1 Base Preferences from Representative Experiment Libraries
  • 08:52Conclusion

Summary

Traduzione automatica

We describe a protocol for amplifying retroviral integration sites from the genomic DNA of infected cells, sequencing the amplified virus-host junctions, and then mapping these sequences to a reference genome. We also describe techniques to quantify the distribution of integration sites relative to various genomic annotations using BEDTools.

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