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Cloning CRISPR-Cas Plasmids

114 bytes added, 19:41, 19 October 2017
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===Verification===
* Miniprep the transformed clones and digest 1 ug of purified plasmid for 1h with AgeI and BbsI in NEB Buffer 1.1 and then run on an agarose gel. Empty vectors will yield a ~1kb fragment, wheras vectors with insert will only be cut once. This is because the cloning removes the BbsI site.
* Send clones with insert for sequencing with the hU6 sequencing primer. Go to http://bridgeslab.sph.umich.edu/protocols/index.php/Submitting_Plasmids_for_Sequencing for further details. Align those clones with the empty vector full sequence or the hU6 sequencing of the empty vector, available at http://www.addgene.org/42335/sequences/. Doublecheck that your insert is correct, and is oriented in the correct manner.
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