GENETIC ENGINEERING Crispr-cas9
• Genome engineering is just about its dawn of golden age. New and exciting tools are arising and piling up to its arsenal.
• .After TALENs, Zing-finger a new tool is coming to add up, the newest addition is the CRISPR system, a system that firstly discovered at the Streptococcus thermophilus and its natural role is the sequence-specific adaptive immunity against foreign DNA in bacteria.
• In the present report, CRISPR Cas9, type II system, used in an attempt to knock out the HPRT1 gene expressed in lymphocytes as a proof of concept. HAT medium in combination with Thioguanine used as a selection medium enabling a quick and simple read-out screening method, to verify that a successful editing event took place in the mammalian cells, a detection DNA cleavage kit used in combination with an enzymatic assay.