The UCSF ES Cell Targeting Core provides high-quality ES cell services with a high probability of germline transmission. The core offers ES cell targeting, genomic DNA extraction from 96-well plates, expansion of targeted ES cells, chromosome counts, and preparation of ES cells for microinjection, to all UCSF investigators as well as external investigators.
Prior to initiation of a project, consultation is available from the director on the entire procedures of generating knockout mice. This meeting will outline specifics of the project including knockout strategy, KO construct design, screening assays, and other issues relevant to microinjection and chimera breeding.
We work with the Gladstone Transgenic Gene Targeting Core for your microinjections to deliver a full-range gene targeting service. On the day of injection, we prepare cells and deliver them to Gladstone.
Recent publications by Core Facility users:
We can extract genomic DNA from ES cells on 96-well plate with minimal cost for your own Southern or PCR genotyping.
Service available only to UCSF researchers. Tell us where you would like to make double-strand breaks in the genome. We design and make TALEN expression plasmids that you can use for either “knockout” (by non-homologous end joining mechanism) or “knock-in” (by homologous recombination mechanism with a donor vector/oligos) approaches to edit the genome. TALENs can be applied to cell lines and embryos of fly, worm, fish, and mammals.
Investigators provide one vial of frozen ES cells with information about culture condition from original resource. We will revive, nurture, and refreeze ES cells (5 vials) when they are ready. In addition, we will give you 1~2 million cells for your genotyping verification.
Investigator’s targeting construct will be electroporated by core personnel. We have two feeder-independent ES cell lines, E14 (129-derived) and JM8A3.N1 (C57BL/6-derived) you can choose from. After drug selection for about one week, up to 300 colonies will be picked. When they are about to be confluent, we will split them as duplicate, one master plate to freeze for future expansion of positive clones and one plate for genotyping to identify targeted ES cell clones. Your plates for genotyping will be ready for pick-up 2-3 weeks after the electroporation date.
Tell us where you would like to make a double-strand cut for knockout, knockin, or a larger genomic deletion. We will design and subclone sgRNAs to modified PX330/PX335 and sequence-verify them. We make 3~4 gRNAs per target site.
ES Cell Targeting Core has not received any reviews.
ES Cell Targeting Core has not received any endorsements.