pX330-U6-Chimeric_BB-CBh-hSpCas9

Protocol tips

Upstream tips	Protocol tips	Downstream 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.

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 gRNAs expressing vector
Three gRNAs were designed using http://crispr.mit.edu. The sequences of the gRNAs are as follow:

gRNA1: catacaattatgtttccccgTGG;

gRNA2: gggtcccttgtggtgctttcAGG;

gRNA3: ttcactcctctgcaaaacccTGG.

The Cas9 and gRNA expression vector pX330-U6-Chimeric_BB-CBh-hSpCas9 was a gift from Feng Zhang [27] (Addgene plasmid # 42230). The linearized vector pX330 was ligated with these gRNAs following Feng Zhang’s protocol.

The gRNA expression vector pDR274, a gift from Keith Joung [33] (Addgene plasmid # 42250), was linearized and ligated with the gRNAs as described above.

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Reviews

Discussion

Papers

Check out relevant papers found by Labettor's AI that are relevant for performing CRISPR Mouse - Deletion ES (embryonic stem) cells Slx2 using pX330-U6-Chimeric_BB-CBh-hSpCas9 from Addgene.

Manufacturer protocol

Download the product protocol from Addgene for pX330-U6-Chimeric_BB-CBh-hSpCas9 below.

Download manufacturer protocol

Videos

Check out videos that might be relevant for performing CRISPR Mouse - Deletion ES (embryonic stem) cells Slx2 using pX330-U6-Chimeric_BB-CBh-hSpCas9 from Addgene. Please note that these videos are representative and steps or experiment specific processes must be kept in mind to expect desired results.