To examine whether or not the regulatory sequence of chicken ovalbumin gene can drive transgene expression specifically in hen oviduct, the authors constructed an oviduct-specific expression vector (pOV), containing 3...To examine whether or not the regulatory sequence of chicken ovalbumin gene can drive transgene expression specifically in hen oviduct, the authors constructed an oviduct-specific expression vector (pOV), containing 3.0 kilobases (kb) of the 5'-flanking sequence and 3.0 kb of the 3'-flanking sequence of the chicken ovalbumin gene. Jellyfish green fluorescence protein (EGFP) reporter gene and bacterial LacZ reporter gene were respectively inserted into the downstream of the 5'-regulatory region. The recombinants were named as pOVEGFP and pOVLacZ. Two transfer systems, in vitro and in vivo, were used to verify the function of the vector. In vitro, the plasmid DNA pOVEGFP and pEGFP-N1 were transfected respectively by the polyethyle-neimine procedure into the primary chicken oviduct epithelium (PCOE) and fibroblasts cells isolated from laying hens. In vivo, the recombinant vector pOVLacZ was injected into egg-laying hens via wing vein and the tissues were collected for RT-PCR analysis. The results showed that expression of pEGFP-Nl was achieved at low level in oviduct epithelial cells and at high level in fibroblasts, but that the recombinant vector was not expressed in both cells. RT-PCR analysis showed that the LacZ gene was transcribed in the oviduct, but not in the heart, liver, kidney and spleen of the injected hens. Accordingly, the β-galactosidase activity was only detected in the oviduct magnum (116.7 mU/ml) and eggs (16.47 mU/ml). These results indicated that the cloned regulation regions of chicken ovalbumin gene could drive exogenous gene expression specifically in the oviducts of hens. In vivo gene injection via wing vein may serve as a rapid production system of recombinant proteins in chicken eggs. In addition, the cultured primary oviduct cells from laying hens were not efficient temporary expression systems for analyzing the function of regulating elements of ovalbumin gene.展开更多
Experiments were conducted to study the effects of dietary taurine and housing density on oviduct function in laying hens. Green-shell laying hens were randomly assigned to a free range group and two caged groups, one...Experiments were conducted to study the effects of dietary taurine and housing density on oviduct function in laying hens. Green-shell laying hens were randomly assigned to a free range group and two caged groups, one with low-density and the other with high-density housing. Each group was further divided into control（C） and taurine treatment（T） groups. All hens were fed the same basic diet except that the T groups＇ diet was supplemented with 0.1% taurine. The experiment lasted 15 d. Survival rates, laying rates, daily feed consumption, and daily weight gain were recorded. Histological changes, inflammatory mediator levels, and oxidation and anti-oxidation levels were determined. The results show that dietary taurine supplementation and reduced housing density significantly attenuated pathophysiological changes in the oviduct. Nuclear factor-κB（NF-κB） DNA binding activity increased significantly in the high-density housing group compared with the two other housing groups and was reduced by taurine supplementation. Tumor necrosis factor-α（TNF-α） m RNA expression in the high-density and low-density C and T groups increased significantly. In the free range and low-density groups, dietary taurine significantly reduced the expression of TNF-α m RNA. Supplementation with taurine decreased interferon-γ（IFN-γ） m RNA expression significantly in the low-density groups. Interleukin 4（IL-4） m RNA expression was significantly higher in caged hens. IL-10 m RNA expression was higher in the high-density C group than in the free range and low-density C groups. Supplementation with taurine decreased IL-10 m RNA expression significantly in the high-density group and increased superoxide dismutase（SOD） activity in the free range hens. We conclude that taurine has important protective effects against oviduct damage. Reducing housing density also results in less oxidative stress, less inflammatory cell infiltration, and lower levels of inflammatory mediators in the oviduct. Therefore, both dietary taurine and reduced housing density can ameliorate oviduct injury, enhance oviduct health, and promote egg production in laying hens.展开更多
文摘To examine whether or not the regulatory sequence of chicken ovalbumin gene can drive transgene expression specifically in hen oviduct, the authors constructed an oviduct-specific expression vector (pOV), containing 3.0 kilobases (kb) of the 5'-flanking sequence and 3.0 kb of the 3'-flanking sequence of the chicken ovalbumin gene. Jellyfish green fluorescence protein (EGFP) reporter gene and bacterial LacZ reporter gene were respectively inserted into the downstream of the 5'-regulatory region. The recombinants were named as pOVEGFP and pOVLacZ. Two transfer systems, in vitro and in vivo, were used to verify the function of the vector. In vitro, the plasmid DNA pOVEGFP and pEGFP-N1 were transfected respectively by the polyethyle-neimine procedure into the primary chicken oviduct epithelium (PCOE) and fibroblasts cells isolated from laying hens. In vivo, the recombinant vector pOVLacZ was injected into egg-laying hens via wing vein and the tissues were collected for RT-PCR analysis. The results showed that expression of pEGFP-Nl was achieved at low level in oviduct epithelial cells and at high level in fibroblasts, but that the recombinant vector was not expressed in both cells. RT-PCR analysis showed that the LacZ gene was transcribed in the oviduct, but not in the heart, liver, kidney and spleen of the injected hens. Accordingly, the β-galactosidase activity was only detected in the oviduct magnum (116.7 mU/ml) and eggs (16.47 mU/ml). These results indicated that the cloned regulation regions of chicken ovalbumin gene could drive exogenous gene expression specifically in the oviducts of hens. In vivo gene injection via wing vein may serve as a rapid production system of recombinant proteins in chicken eggs. In addition, the cultured primary oviduct cells from laying hens were not efficient temporary expression systems for analyzing the function of regulating elements of ovalbumin gene.
基金Project supported by the National Natural Science Foundation of China(Nos.31101783 and 31472164)the Special Fund for Independent Innovation of Agricultural Science and Technology in Jiangsu Province of China(No.CX(13)3070)
文摘Experiments were conducted to study the effects of dietary taurine and housing density on oviduct function in laying hens. Green-shell laying hens were randomly assigned to a free range group and two caged groups, one with low-density and the other with high-density housing. Each group was further divided into control（C） and taurine treatment（T） groups. All hens were fed the same basic diet except that the T groups＇ diet was supplemented with 0.1% taurine. The experiment lasted 15 d. Survival rates, laying rates, daily feed consumption, and daily weight gain were recorded. Histological changes, inflammatory mediator levels, and oxidation and anti-oxidation levels were determined. The results show that dietary taurine supplementation and reduced housing density significantly attenuated pathophysiological changes in the oviduct. Nuclear factor-κB（NF-κB） DNA binding activity increased significantly in the high-density housing group compared with the two other housing groups and was reduced by taurine supplementation. Tumor necrosis factor-α（TNF-α） m RNA expression in the high-density and low-density C and T groups increased significantly. In the free range and low-density groups, dietary taurine significantly reduced the expression of TNF-α m RNA. Supplementation with taurine decreased interferon-γ（IFN-γ） m RNA expression significantly in the low-density groups. Interleukin 4（IL-4） m RNA expression was significantly higher in caged hens. IL-10 m RNA expression was higher in the high-density C group than in the free range and low-density C groups. Supplementation with taurine decreased IL-10 m RNA expression significantly in the high-density group and increased superoxide dismutase（SOD） activity in the free range hens. We conclude that taurine has important protective effects against oviduct damage. Reducing housing density also results in less oxidative stress, less inflammatory cell infiltration, and lower levels of inflammatory mediators in the oviduct. Therefore, both dietary taurine and reduced housing density can ameliorate oviduct injury, enhance oviduct health, and promote egg production in laying hens.
