pCrU6.3-SpCas9 cloning (pPH159)
From Simon Kelterborn, Peter Hegemann lab, Humboldt University-Berlin October 2017
Cloning vector for Streptococcus pyogenes Cas9 (SpCas9) single guide RNA transcription, controlled by the U6 snRNA promoter #3 (3’ of Cre08.g371751). The 20 bp target site can be inserted in a cut-ligation reaction as annealed oligos into an Esp3I cloning site following the protocol of Ann Ran (Ran et al. 2013). The immediate 4-bp sequence upstream of the transcriptional start site was changed to ACTT in all U6 constructs to simplify the cloning procedures.