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Lecture 26 Transgenesis with high capacity vectors Transgenesis with High-Capacity Vectors • transgenes are complementary DNAs (cDNAs), small genes (<20 kb) or parts of genes • cDNAs are poorly expressed in mammalian cells • In transgenesis, important gene- specific regulatory sequences retained as part of the insert • • complete genes and multigene complexes (<100 kb) large for conventional vectors • need for high-capacity vectors • eg. bacterial, P1 bacteriophage- derived, mammalian, and yeast artificial chromosomes [ BACs, PACs,MACs and YACs, respectively]) • carry genomic DNA ranging in size from 100 to >1,000kb • used for transgenesis YAC transgenesis • transgenic mice produced by microinjection of the pronucleus of the fertilized egg or transfection of ES cells with YACs • carry either an array of related genes or a single large gene • used to study developmental processes • as models for human disorders • and for the production of human therapeutic agents. YAC transgenesis • For example, transgenic mice with the human globin gene cluster, • covers about 250kb and contains five different functional globin genes, • express genes in a tissue-specific and timedependent manner that corresponds to the site specificity and temporal pattern in humans. YAC transgenesis • production of mice that synthesize only human antibodies • Human monoclonal antibodies • Xenomouse Schematic representation of the κ and H human immunoglobulin genes.
