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利用含有白喉毒素N端序列的基因作上游引物、含有αMSH全序列作下游引物,以pET28a/DAB389-EGF为模板,PCR扩增DAB389-αMSH基因片段,用限制性内切酶EcoR I和NcoI酶切,并插入原核表达载体pET28a的相应位点,构建了重组表达载体pET28a/DAB389-αMSH,在大肠杆菌中表达重组融合蛋白DAB389-αMSH,转化菌经1 mmol/L IPTG、30℃诱导5 h,用SDS-PAGE和Western blot鉴定表达的重组毒素。结果表明扩增的片段与理论值一致,重组质粒的DNA序列分析正确;SDS-PAGE表明重组毒素相对分子量为43.76 KD,且表达量达菌体总蛋白量的31.6%。Western blot分析显示,重组毒素能特异性地与抗白喉抗体结合,表明已成功构建表达了重组DAB389-αMSH的工程菌株,并获得表达蛋白。  相似文献   
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The main experiment assessed whether the inhibitory effects of the dopamine agonist, cabergoline, on prolactin and α-melanocyte stimulating hormone (MSH) concentrations would persist throughout a longer-term administration (65 days). The possible effect of cabergoline on insulin sensitivity was also studied. Ten mares known to be insulin insensitive were allotted to two groups (treated vs. control). An insulin challenge, a glucose tolerance test, and a sulpiride challenge were administered before treatment. On day 0, treated mares (n = 5) received an injection of 5 mg cabergoline in slow-release vehicle; control mares (n = 5) received an equivalent vehicle injection. Injections were repeated every 10 days for a total of seven injections. Sulpiride challenges were done 1 day before each cabergoline treatment to assess possible refractoriness to the treatment. Behavior and hair coat density were also monitored. Plasma prolactin was suppressed (P < .01) to undetectable levels in mares receiving cabergoline; control mares had robust prolactin responses to each sulpiride injection. There was no indication of refractoriness to cabergoline over time. Plasma MSH concentrations after sulpiride were also suppressed (P < .05) by cabergoline. After treatment, neither the glucose response to insulin nor the insulin response to glucose differed (P > .1) between groups. No behavioral changes were noted because of treatment. Weight of hair samples indicated that cabergoline perturbed (P < .05) winter coat growth. It is concluded that 5 mg of cabergoline in slow-release vehicle administered every 10 days is an effective way of delivering dopaminergic activity to mares that results in no noticeable detrimental effects and no refractoriness to the drug.  相似文献   
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针对池塘养殖牙鲆(Paralichthys olivaceus)无眼侧体色黑化消褪的现象,本研究从形态、血清和mRNA水平上对其生理学机制进行了研究.光镜观察显示,无眼侧消褪区域的黑色素细胞数量显著少于有眼侧和无眼侧黑化区域(P<0.05),电镜观察发现,无眼侧消褪区域皮肤中黑色素颗粒模糊,黑色素细胞中色素体存在凋亡现象.无眼侧黑化区域消褪过程中,鳞片类型经历了由栉鳞—弱栉鳞—圆鳞转变的过程,栉鳞上硬棘数量也随之减少.无眼侧黑化消褪的牙鲆血清中的黑色素聚集激素(MCH)肽含量显著高于无眼侧正常和无眼侧黑化的牙鲆(P<0.05),但无眼侧体色黑化的牙鲆血清中的黑色素细胞刺激激素(MSH)肽含量显著高于其他2种类型的牙鲆(P<0.05).基因表达分析显示,无眼侧黑化消褪鱼垂体MCH mRNA表达水平显著高于无眼侧黑化鱼(P<0.05),而垂体POMC1 mRNA表达水平显著低于无眼侧黑化鱼(P<0.05),但都与无眼侧正常鱼无显著差异.研究结果可为阐释池塘养殖牙鲆无眼侧体色黑化调控机制提供理论支持.  相似文献   
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使用DNASTAR软件,利用计算机模拟比较了带有亲和标记和连接肽的DAB389(Gly4Ser)2-αMSH与DAB389-αMSH两种融合蛋白的抗原性、亲水性、等电点及表位等分子生物学特性.结果显示,DAB389(Gly4Ser)2-αMSH的抗原性和表位明显低于DAB389-αMSH,而DAB389(Gly4Ser)2-αMSH的亲水性高于DAB389-αMSH,两者的等电点均为5.9.  相似文献   
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AIM: The present study was undertaken to explore the effects of α-MSH on partial biological activities of LPS. METHODS:Colorimetric method was used for the measurement of hydrogen peroxide(H2O2) production in mouse peritoneal macrophages, the apoptosis of polymorphonuclear leukocytes(PMNs) and the binding of LPS to monocytes were studied with flow cytometry. RESULTS: It was found that LPS strongly stimulated macrophages to release H2O2. When macrophages were cultured with α-MSH in the presence of LPS, the H2O2 release was markedly suppressed (P<0.01). Neither LPS nor α-MSH alone was capable of affecting the apoptosis of PMNs (P>0.05). In the presence of LPS, however, α-MSH significantly promoted the apoptosis of PMNs (P<0.01). α-MSH significantly inhibited the binding of LPS to monocytes as the binding rate of FITC-LPS and the mean surface fluorescence intensity of monocytes ( P<0.05 and P<0.01, respectively). CONCLUSION: In the presence of LPS, α-MSH not only effectively suppressed the release of H2O2 from macrophages , promoted the apoptosis of PMNs, but also interfered with the binding of LPS to monocytes. α-MSH may play an important role in the immunomodulation of the body .  相似文献   
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