lentiCRISPR v2

CRISPR Human - Deletion STING exon 5

Experiment
CRISPR Human - Deletion STING exon 5
Product
lentiCRISPR v2 from Addgene
Manufacturer
Addgene

Protocol tips

Protocol tips
There is no need to perform a negative control golden-gate reaction (without
insert) as it will always contain colonies so not a good indicator of cloning success.

Publication protocol

Generation of STING-deficient cells with CRISPRs.
A single guide RNA (sgRNA) targeting STING exon 5 (sgRNASTING) (5′-AATATGACCATGCCAGCCCA-3′) was cloned into the LentiCRISPR vector (catalog number 49535; Addgene) (26) to generate pLVCR-sgSTING. Human telomerase reverse transcriptase (hTERT)-immortalized HUVECs (tHUVECs) were transduced with either the empty LentiCRISPR vector (control) or pLVCR-sgSTING (STING knockout [STINGKO]) and selected with puromycin (1 μg/ml) for 7 days. To obtain single-cell clones, cells were plated at 0.6 cells per well into 96-well plates, and all STINGKO clones used in this study (clones 9 and 11) were verified by Sanger sequencing for deletion and by Western blotting for STING expression.

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Reviews

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Discussion

Discussion

4 years ago

Author: Milena Alexeyeva Russian Federation

DNA insert using CRISPR

I would like to excise a large strand of DNA and insert a new one using CRISPR. My problem is that my strand will be a little over 1kb and I am not sure if this is going to be a limiting factor. Also, how long should the homology arms be for a region of this size?

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Papers

Check out relevant papers found by Labettor's AI that are relevant for performing CRISPR Human - Deletion STING exon 5 using lentiCRISPR v2 from Addgene.

Paper title
cGAS-STING Signaling Regulates Initial Innate Control of Cytomegalovirus Infection
Full paper
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Manufacturer protocol

Download the product protocol from Addgene for lentiCRISPR v2 below.

Download PDF Download manufacturer protocol

Videos

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