Lenti v2 insertion into the genome

Hello all - I can't find this answer elsewhere. Hoping someone here can help.

We're following the lenti v2 protocol, and are curious about how much of the v2 plasmid is inserted into the host genome. Is it just the cas9 and sgRNA or more? How do you determine where it was inserted? We want to make sure it's not disrupting any coding genes.


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Bronze Ghoulover 4 years ago

Only the region from the 5' to 3' LTR of the lentiviral construct will be integrated.

Determining integration site is not so straightforward without doing a variation of inverse PCR. You can buy a kit that does it: http://www.clontech.com/US/Products/Viral_Transduction/Lentiviral_Transduction_Tools/Lentivirus_Integration_Site?sitex=10020:22372:US

Note that each cell will have a different integration site so you would need to perform this on an isolated clone.

Yellow Wendigoover 4 years ago

Thanks so much - answered my question!

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