MLM3636

CRISPR Mouse - Deletion Neuro 2a Prnp

Experiment
CRISPR Mouse - Deletion Neuro 2a Prnp
Product
MLM3636 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 gRNA Expression Vectors
The SpCas9 plasmid JDS246 (Plasmid 43861) and the gRNA expression plasmid MLM3636 (Plasmid 43860) were obtained from a non-profit plasmid share repository (Addgene, Cambridge, MA, USA). Suitable CRISPR target sites within Prnp Exon 3 positive and negative strands were identified using the ‘CRISPR Design Tool’ (http://crispr.mit.edu/) described in the Results section. The respective oligonucleotide pairs were obtained from Life Technologies (Burlington, ON, Canada) and were customized to include overhangs compatible for ligation into MLM3636 linearized by digestion with BsmB1 (cat. no. R0580S; New England BioLabs, Ipswich, MA, USA), a cloning site located in this vector on the 3′ side of a U6 promoter element. Oligonucleotides were phosphorylated with polynucleotide kinase (cat. no. EK0031; Fermentas, Ottawa, ON, Canada), annealed and inserted into the gRNA plasmid using T4 DNA ligase (cat no. M0202S; New England Biolabs) and transformed into Turbo competent E. coli (cat no. C2984H; New England Biolabs).

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Reviews

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Discussion

Discussion

4 years ago

Author: Mario Udinese Italy

Floxing mice with CRISPR

Hi everyone! I am planning on floxing mice with CRISPR but I am having trouble deciding which region to target. Do you have any tips on choosing?

Discussion

5 years ago

Author: Ben Saar Israel

How to choose a region to target for CRISPR

Hi everyone! I am planning on floxing mice with CRISPR but I am having trouble deciding which region to target. Do you have any tips on choosing?

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

Papers

Check out relevant papers found by Labettor's AI that are relevant for performing CRISPR Mouse - Deletion Neuro 2a Prnp using MLM3636 from Addgene.

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Manufacturer protocol

Download the product protocol from Addgene for MLM3636 below.

Download PDF Download manufacturer protocol

Videos

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