A method for high efficiency YAC lipofection into murine embryonic stem cells

We describe a modified protocol for introducing yeast artificial chromosomes (YACs) into murine embryonic stem (ES) cells by lipofection. With a decreased DNA:cell ratio, increased concentration of condensing agents and altered culture conditions, this protocol reduces the requirement for YAC DNA to a few micrograms, improves the recovery of neomycin-resistant ES colonies and increases the yield of clones containing both flanking vector markers and insert. These modifications enable generation of sufficient ‘intact ’ transgenic clones for biological analysis with a single experiment. Large DNA fragments carried on yeast artificial chromosomes (YACs) have been introduced into the mammalian genome (1,2)