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Culture of Bovine Fetal Skin Fibroblasts in vitro and Ipr1 Gene Transfection

畜牧兽医学报 2011年第S1期 7-12页

作者： SONG Yong-li 1,2,HE Xiao-ning 2,HUA Song 2,LAN Jie 2,ZHENG Yue-mao 2,LI Ji-xia 2,HE Xiao-ying 2,QUAN Fu-sheng 2,ZHANG Yong 2(1. College of Veterinary Medicine,Jilin Agricultural Science and Technology College,Jilin132101,China 2. Key Laboratory of Animal Reproductive Physiology & Embryo Technology,Ministry of Agriculture,College of Veterinary Medicine,Northwest Agricuture and Forestry University,Yangling 712100,China)

In order to generate transgenic donor cells for nuclear transfer, bovine fetal fibroblasts were isolated in vitro and transfected with the eukaryotic expression vector pSRA-EGFP-Ipr1. The mouse Ipr1 gene and human SR-A promoter were successfully cloned and then used to construct this macrophage-specific eukaryotic expression vector. Bovine fetal fibroblasts in stable primary culture (4th passage) were transfected with pSRA-EGFP-Ipr1 by electroporation. Fluorescence from GFP was observed after 24h. Transgenic cells were selected using G418 and the resultant monoclones were picked and expanded. The transgenic cells, at the 9 th passage, were evaluated by PCR and flow cytometry. The inserted Ipr1 was confirmed by PCR, indicating stable integration of the transgene into the genome and cells had normal karyotypes and very good appearance, which indicate no deleterious result of the transgenesis. In conclusion, the cells obtained could be used as donor cells for nuclear transfer for further research of transgenic cattle.

关键词： donor cells eunuclear expression vector Ipr1 gene bovine fetal fibroblasts cells electroporation EGFP

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X-ray irradiation selectively kills thymocytes of different stages and impairs the maturation of donor-derived CD4^+CD8^+ thymocytes in recipient thymus

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The Journal of Biomedical Research 2012年第5期26卷 355-364页

作者： Jinbo Li Hongquan Cai Jianliang Jin Qian Wang Dengshun Miao The Research Center for Bone and Stem cells Department of Human Anatomy Nanjing Medical University Nanjing Jiangsu 210029 China Department of Preventive Medieine School of Public Health Nanjing Medical University Nanjing Jiangsu 210029 China

The aim of the present study was to determine whether the sensitivity of thymocytes to X-ray radiation depends on their proliferative states and whether radiation impairs the maturation of donor-derived thymocytes in recipient *** assigned 8-week-old C57BL/6J mice into three treatment groups：1） untreated;2） X-ray radiation;3） X-ray radiation plus bone marrow transplantation with donor bone marrow cells from transgenic mice express-ing enhanced green fluorescent protein（GFP） on a universal *** 4 weeks,the size of the thymus,the number and proliferation of thymocytes and ratios of different stage thymocytes were analyzed by immunohisto-chemistry and flow *** results showed that：1） CD4＋CD8＋ thymocytes were more sensitive to X-ray radiation-induced cell death than other thymocytes;2） the proliferative capacity of CD4＋CD8＋ thymocytes was higher than that of other thymocytes;3） the size of the thymus,the number of thymocytes and ratios of thymo-cytes of different stages in irradiated mice recovered to the normal level of untreated mice by bone marrow trans-plantation;4） the ratio of GFP-positive CD4＋CD8＋ thymocytes increased significantly,whereas the ratio of GFP-positive CD4＋ or CD8＋ thymocytes decreased *** results indicate that the degree of sensitivity of thymocytes to X-ray radiation depends on their proliferative states and radiation impairs the maturation of donor-derived CD4＋CD8＋ thymocytes in recipient thymus.

关键词： thymus radiation CD4^+CD8^+ thymocytes sensitivity donor cells

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