Roger Craig

Tagged Mutagenesis by Efficient Minos-Based Germ Line Transposition

ABSTRACT Germ line gene transposition technology has been used to generate “libraries” of flies and worms carrying genomewide mutations. Phenotypic screening and DNA sequencing of such libraries provide functional information resulting from insertional events in target genes. There is also a great need to have a fast and efficient way to generate mouse mutants in vivo to model developmental defects and human diseases. Here we describe an optimized mammalian germ line transposition system active during early mouse spermatogenesis using the Minos transposon. Transposon-positive progeny carry on average more than 2 new transpositions, and 45 to 100% of the progeny carry an insertion in a gene. The optimized Minos-based system was tested in a small rapid dominant functional screen to identify mutated genes likely to cause measurable cardiovascular “disease” phenotypes in progeny/embryos. Importantly this system allows rapid screening for modifier genes.

Stable expression of human growth hormone over 50 generations in transgenic insect larvae

Developments in insect transgenesis using transposons combined with available mass rearing technology for insects such as the Medfly, Ceratitis capitata, provide opportunity for the production of protein for industrial, agricultural and healthcare purposes on a very large scale. In this study, we report the germ-line transformation and expression of a cDNA encoding human growth hormone (hGH) in transgenic Drosophila using the Minos transposon. Production and secretion of a bioactive hGH into the haemolymph of transgenic larvae was demonstrated by immunoblot analysis, ELISA and a proliferation bioassay. Stable expression of hGH was observed over 50 generations. The results indicate that mass reared transgenic diptera with a rapid period of larval growth could provide cost effective production systems for the manufacture of therapeutic and other high value proteins.