Gene knock-out in hard-to-transfect cells
Gene knock-out, which is gene editing leading to loss of function, just requires delivering into cells the 2 CRISPR system actors: the CAS9 endonuclease and the specific guide RNA to target the gene of interest. Thus, simple transfection of CleanCap CAS9 mRNA and gRNA followed by cell isolation with the smart aliquotor is a convenient and robust method. Howevery, what can you do if the cells are hard-to-transfect?
CAS9 expressing lentiviral particules
With a very high titer >10^9TU/ml, Signagen laboratories provide CAS9 expressing lentiviral particles under CMV or EF1a promoter.
Through the transduction, lentiviral particles provide high level of delivery into most hard-to-transfect cells. To test this solution on a cell type, control lentiviral particles are available such as the LV-EF1a-GFP-Puro from Signagen laboratories. You can discover them right here and download the manual.
Alternatively, if it is preferable to express CAS9 and the eGFP reporter in the same lentiviral particles, Genecopoeia produces this.
Lentivirus for gRNA
Genecopoeia designs the sgRNA and produce the lentiviral particles from the plasmid pCRISPR-LvSG03 below. Contact me through the form below to get a specific quotation.
The gene editing can be performed with lentiviral particles. Some researchers even use it to develop CAS9 expressing cell lines. It is a very good solution for hard-to-transfect cells, however, due to the packaging size, it is only for knock out, not for knock-in.