Fig. 1
From: A CRISPR interference strategy for gene expression silencing in multiple myeloma cell lines
Schematic protocol of sgRNA cloning into lentiviral plasmid. First, the annealing of the primers was required to generate the sgRNAs (A). Then, the lentiviral plasmid was digested with Esp3I enzyme to linearize, purify and quantify (B). The ligation of the sgRNA into the linearized lentiviral plasmid was made at 16ºC overnight in presence of T4 ligase enzyme (C). Finally, the ligated plasmids were transformed in DH5α competent E. coli (D) and colonies were checked by PCR and sequencing, before DNA amplification.