CRISPR Human - Deletion HPRT1_A

The RNA-guided CRISPR-Cas9 nuclease system has revolutionized the genome editing practices. For the most part, the Cas9-mediated genome editing is performed either via nonhomologous end joining (NHEJ) or homology-directed repair (HDR) in mammalian cells, However, designing of specific sgRNAs and minimizing off-target cleavage mediated mutagenesis are the major challenges in CRISPR-Cas based genome editing. To circumvent these issues, we can take advantages of many available tools and approaches for sgRNA construction and delivery.

Start discussion

Found 1 discussion for this experiment

Discussion

1 year ago

1 year ago by 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?

Share your thoughts or question with experts in your field by adding a discussion!

Found 1 matching solution for this experiment

pX330-U6-Chimeric_BB-CBh-hSpCas9 + Lipofectamine® LTX Reagent

Protocol tips
For goldengate reaction, there is a low and a high concentration mixture available for T4 ligase. For single inserts, the low concentration is just fine, but the follow-up article on golden gate cloning (PMID: 19436741) found that the high-concentration T4 was better for multiple insert cloning.
Can't find the product you've used to perform this experiment? It would be great if you can help us by Adding a product!
Become shareholder Discussions About us Contact Privacy Terms