title: Transfer of an expression YAC into goat fetal fibroblasts by cell fusion for mammary gland bioreactor.
abstract: Yeast artificial chromosomes (YACs) as transgenes in transgenic animals are likely to ensure optimal expression levels. Microinjection of YACs is the exclusive technique used to produce YACs transgenic livestock so far. However, low efficiency and high cost are its critical restrictive factors. In this study, we presented a novel procedure to produce YACs transgenic livestock as mammary gland bioreactor. A targeting vector, containing the gene of interest-a human serum albumin minigene (intron 1, 2), yeast selectable marker (G418R), and mammalian cell resistance marker (neo(r)), replaced the alpha-lactalbumin gene in a 210kb human alpha-lactalbumin YAC by homogeneous recombination in yeasts. The chimeric YAC was introduced into goat fetal fibroblasts using polyethylene glycol-mediated spheroplast fusion. PCR and Southern analysis showed that intact YAC was integrated in the genome of resistant cells. Perhaps, it may offer a cell-based route by nuclear transfer to produce YACs transgenic livestock.
authors.affiliation: Shanghai Transgenic Research Center, China.
authors.forename: Xu-Feng
authors.initials: XF
authors.lastname: Zhang
authors.forename: Guo-Xiang
authors.initials: GX
authors.lastname: Wu
authors.forename: Jian-Quan
authors.initials: JQ
authors.lastname: Chen
authors.forename: Ai-Min
authors.initials: AM
authors.lastname: Zhang
authors.forename: Si-Guo
authors.initials: SG
authors.lastname: Liu
authors.forename: Bing-Hua
authors.initials: BH
authors.lastname: Jiao
authors.forename: Guo-Xiang
authors.initials: GX
authors.lastname: Cheng
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified / Q000378:metabolism*; D019149:Bioreactors; D002459:Cell Fusion / Q000379:methods; D002478:Cells, Cultured; D018244:Chromosomes, Artificial, Yeast / Q000235:genetics*; D003001:Cloning, Molecular / Q000379:methods; D005347:Fibroblasts / Q000378:metabolism*; D018390:Gene Targeting / Q000379:methods; D018014:Gene Transfer Techniques; D006041:Goats / Q000196:embryology / Q000235:genetics* / Q000378:metabolism*; D007768:Lactalbumin / Q000096:biosynthesis* / Q000235:genetics; D008321:Mammary Glands, Animal / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis
chemical_list: D011994:Recombinant Proteins; D007768:Lactalbumin