title: Targeting the exogenous htPAm gene on goat somatic cell beta-casein locus for transgenic goat production.
abstract: Combining gene targeting of animal somatic cells with nuclear transfer technique has provided a powerful method to produce transgenic animal mammary gland bioreactor. The objective of this study is to make an efficient and reproducible gene targeting in goat fetal fibroblasts by inserting the exogenous htPAm cDNA into the beta-casein locus with liposomes or electroporation so that htPAm protein might be produced in gene-targeted goat mammary gland. By gene-targeting technique, the exogenous htPAm gene was inserted to milk goat beta-casein gene sequences. Fetal fibroblasts were isolated from Day 35 fetuses of Guanzhong milk goats, and transfected with linear gene-targeting vector pGBC4htPAm using Lipefectamin-2000 and electoporation, respectively. Forty-eight gene-targeted cell colonies with homologous recombination were obtained, and three cell colonies were verified by DNA sequence analysis within the homologous recombination region. Using gene-targeted cell lines as donor cells for nuclear transfer, a total of 600 reconstructed embryos had been obtained, and 146 developed cloned embryos were transferred to 16 recipient goats, and finally three goats showed pregnancy at Day 90.
authors.affiliation: Department of Animal Sciences, Institute of Animal Reproduction, Development and Genetic Engineering, Laiyang Agricultural University, Qingdao, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Shen
authors.forename: Guocheng
authors.initials: G
authors.lastname: Lan
authors.forename: Xueyi
authors.initials: X
authors.lastname: Yang
authors.forename: Lan
authors.initials: L
authors.lastname: Li
authors.forename: Lingjiang
authors.initials: L
authors.lastname: Min
authors.forename: Zhengtian
authors.initials: Z
authors.lastname: Yang
authors.forename: Liyuan
authors.initials: L
authors.lastname: Tian
authors.forename: Xiaojie
authors.initials: X
authors.lastname: Wu
authors.forename: Yujiang
authors.initials: Y
authors.lastname: Sun
authors.forename: Hong
authors.initials: H
authors.lastname: Chen
authors.forename: Jinghe
authors.initials: J
authors.lastname: Tan
authors.forename: Jixian
authors.initials: J
authors.lastname: Deng
authors.forename: Qingjie
authors.initials: Q
authors.lastname: Pan
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics; D002364:Caseins / Q000235:genetics*; D019976:Cloning, Organism; D005260:Female; D018390:Gene Targeting; D005818:Genetic Engineering / Q000379:methods; D005822:Genetic Vectors / Q000138:chemical synthesis; D006041:Goats / Q000235:genetics*; D006801:Humans; D050505:Mutant Proteins / Q000235:genetics; D053652:Nuclear Transfer Techniques; D010957:Plasmids / Q000138:chemical synthesis; D011247:Pregnancy; D011270:Pregnancy, Animal; D010959:Tissue Plasminogen Activator / Q000235:genetics*
chemical_list: D002364:Caseins; D050505:Mutant Proteins; D010959:Tissue Plasminogen Activator