Abstract: Disclosed are vectors, kits and methods useful in the construction of recombinant cells and DNAs via enhanced efficiency homologous recombination. The vectors are targeting vectors that contain a gene-of-interest spliced between two ends that are homologous to a genome target site. The ends of the vector may be protected from exonuclease attack by deploying a cap, such as a hair pin structure. The vector is linked to a nuclear localization signal sequence, and preferably, a bait peptide that binds to RAD51, to facilitate homologous recombination. The vector may be deployed in myriad genetic transformation applications, such as site-directed mutagenesis, gene therapy, and the like.
Abstract: Disclosed are vectors, kits and methods useful in the construction of recombinant cells and DNAs via enhanced efficiency homologous recombination. The vectors are targeting vectors that contain a gene-of-interest spliced between two ends that are homologous to a genome target site. The ends of the vector may be protected from exonuclease attack by deploying a cap, such as a hair pin structure. The vector is linked to a nuclear localization signal sequence, and preferably, a bait peptide that binds to RAD51, to facilitate homologous recombination. The vector may be deployed in myriad genetic transformation applications, such as site-directed mutagenesis, gene therapy, and the like.
Abstract: Disclosed are embryoid bodies having a uniform size of approximately 415 nm and comprising genetically modified embryonic stem cells, and methods of making same. The genetically uniform embryoid bodies can be multiplexed as one embryoid body per well in a multiwell format, and used as a high to medium throughput screen for test agents that affect the development and homeostasis of animals, including humans. The genetic modification of the embryonic stem cells is a promoter-report-selection construct that enables the selection and detection of cells of a particular lineage in the EB, to determine the effects of a test agent.