副猪嗜血杆菌外膜蛋白P5对豚鼠树突状细胞成熟及功能影响

为研究副猪嗜血杆菌(Haemophilus parasuis,Hps)外膜蛋白P5对豚鼠骨髓来源的树突状细胞(Dendritic cell,DC)刺激淋巴细胞增殖功能及细胞因子分泌的影响,以GM-CSF、IL-4联合诱导骨髓细胞生成未成熟DC,加入不同剂量副猪嗜血杆菌外膜蛋白P5,观察DC形态,检测混合淋巴细胞反应,ELISA法测IL-6、IL-12p70、TNF-α分泌情况.结果显示:经副猪嗜血杆菌外膜蛋白P5刺激后,可获得具有典型树突状突起形态的DC;经5μg·mL-1P5刺激的DC较对照IL-6、TNF-α分泌量极显著增加(P＜0.01),IL-12的分泌量有所增加(P＞0.05);50 μg·mL-1P5刺激的DC较对照IL-6分泌量极显著增加(P＜0.01),IL-12p70和TNF-α分泌量均极显著下降(P＜0.01).诱导后的DC刺激同种异体T淋巴细胞增殖的能力极显著增强(P＜0.01).本研究证明P5能影响骨髓细胞来源的DC的分化、成熟及功能的发挥,且不同剂量的P5对DC的成熟程度影响有差异.     

鸭腔上囊胚胎及胚后发育期凋亡相关蛋白的免疫组化分析

研究天府肉鸭腔上囊胚胎及胚后发育期Bcl-2、Bax、Fas、FasL蛋白的表达。将20只天府肉鸭分为4组,即24天胚龄(E24),胚后3、8、29周龄(P3、P8、P29),采用免疫组化技术。结果显示,Bcl-2、Bax、Fas、FasL在各组滤泡淋巴细胞中均有表达,FasL还表达于滤泡间上皮。滤泡皮质和髓质淋巴细胞Bcl-2阳性率呈递减的变化趋势(髓质P3～8除外);皮质和髓质淋巴细胞Bax阳性率在E24～P8恒定,P29上升;皮质和髓质淋巴细胞Bcl-2/Bax值呈下降趋势,但皮质在P3～8恒定,髓质在P3～8上升;滤泡皮质和髓质淋巴细胞Fas阳性率在E24～P3上升,P3～8下降,P8～29上升;皮质和髓质淋巴细胞FasL阳性率变化规律与Fas相似,但皮质在P3～8恒定。滤泡皮质淋巴细胞Bcl-2阳性率及Bcl-2/Bax值在E24～P8明显高于髓质,在P29则显著低于髓质;皮质淋巴细胞Bax阳性率在E24～P8与髓质无显著差异,在P29则明显高于髓质;皮质淋巴细胞Fas和FasL阳性率在E24～P8明显低于髓质,在P29则显著高于髓质。结果提示,Bcl-2、Bax、Fas、FasL是鸭腔上囊胚胎及胚...     

牛GM-CSF与金黄色葡萄球菌黏附素FnBPB共表达质粒的构建与原核表达

采用PCR方法对牛粒细胞-巨噬细胞集落刺激因子(GM-CSF)和金黄色葡萄球菌FnBPB的D区进行特异性的扩增,并通过重叠延伸PCR扩增GM-CSF-FnBPB串联基因,构建了克隆质粒pMD19-GM-CSF-FnBPB。利用表达载体pET-32a(+)对该融合基因片段进行原核表达,SDS-PAGE分析表明,在1mmol/L IPTG诱导浓度下,在39ku处出现了与目的蛋白一致的外源蛋白带,Western blot分析表明,该蛋白具有反应原性,进而证明该融合基因成功在原核细胞中表达。     

高通量筛选差异表达基因技术及其在家畜中的应用

差异表达基因筛选技术是研究生物重要经济性状相关基因或抗病相关基因的有力工具。作者总结了近年来发展起来的几种高通量筛选差异表达基因的技术,并对其原理、流程及在家畜中的应用作了简要介绍。     

Effect of hot air,UV-C,white light and modified atmosphere treatments on expression of chlorophyll degrading genes in postharvest broccoli (Brassica oleracea L.) florets

Several treatments were applied in order to delay postharvest degreening in broccoli florets and investigate their effects on genes associated with chlorophyll catabolism. Degradation of chlorophylls is the most evident visual manifestation of broccoli postharvest deterioration, occurring rapidly due to the immature stage in which the material is harvested. Treatments such as storage in modified atmosphere, exposure to hot air, UV-C and white lamps were employed in the current work to induce a delay in degreening and chlorophyll degradation. Expression of genes possibly related to chlorophyll catabolism was analyzed in these samples and discussed. Chlorophyllases, the enzymes traditionally believed to remove the phytol side chain from chlorophyll appear to have a gene expression that was not regulated by postharvest treatments. Pheophytinase, a recently discovered new enzyme in this metabolic pathway, correlated chlorophyll loss accurately in heat, UV-C and white-light treatments, but not in modified atmospheres. Results presented in this work indicate that postharvest treatments that delay chlorophyll degradation have a higher effect on the expression of pheophytinase rather than on chlorophyllase genes.     

Analysis of differential protein expression during the early stage of in vitro tuberization in taro

Tuberization is a complex and multilevel developmental process. Many important metabolic changes in the early stage of tuberization are crucial to the tuber differentiation and development. In this study, we attempted to identify proteins differentially expressed in the early stage of in vitro tuberization in taro (Colocasia esculenta var. antiquorum) by two-dimensional polyacrylamide gel electrophoresis (2-DE) and mass spectrometry (MS). Protein samples from shoot tips cultured in 8% sucrose media at 0, 2, 4, 6 and 8 d were separated with 2-DE. A total of 13 differentially expressed proteins were analyzed with MS. Four proteins via, glyceraldehyde-3-phosphate dehydrogenase, alcohol dehydrogenase, chloroplast protein synthesis elongation factor (EF-Tu), and ankyrin repeat protein HBP1 were successfully identified during in vitro tuberization of taro. This implies that important metabolic changes, including sucrose metabolism, signal transduction and cell defense, occurred in the early stage of in vitro tuberization in taro.     

Protective role of heat-shock protein 70 in gastric mucosal lesion induced by endotoxin in cirrhotic rats with portal hypertensive gastropathy

AIM: To investigate the protective role of heat-shock protein 70 (HSP70) in the pathogenesis of gastric mucosal damage in cirrhotic rats with portal hypertensive gastropathy (PHG).METHODS: The rat model of liver cirrhosis with PHG was established by injection with tetrachloride.The animals were divided into normal control group, PHG group, PHG+heat treatment group, PHG+BPI21 group and PHG+endotoxin groups.The endotoxin used in the experiment was at the dose of 3 mg/kg and endotoxin antagonist BPI21 was at the dose of 2 mg/kg.HSP70 was induced by pre-treating the animals with mild whole-body heating.The levels of HSP70 and tumor necrosis factor alpha (TNF-α) in the gastric mucosa were measured by ELISA.Furthermore, the pathological changes of the gastric mucosa were observed under microscope with HE staining.RESULTS: Compared with the normal control rats, the rats in PHG group showed obvious gastric pathological lesion, decrease in HSP70 production and increase in TNF-α level in the gastric mucosa, and increased endotoxin concentration in the plasma.Compared with PHG+endotoxin group, the gastric mucosal lesion in PHG+BPI21 group was significantly attenuated, accompanied by the increase in HSP70 production and decrease in TNF-α level in the gastric mucosa.Heat treatment increased HSP70 production and decreased TNF-α concentration in the PHG rats, thus attenuating the gastric mucosal damage.CONCLUSION: HSP70 alleviates the gastric mucosal lesion induced by endotoxin in cirrhotic rats with PHG and decreases the concentration of TNF-α in gastric mucosa, indicating a protective role of HSP70 in the pathogenesis of gastric mucosal damage in PHG.     

喷雾干燥鸡血浆蛋白粉工艺优化

为了开发利用鸡血蛋白质资源,该文探讨了鸡血浆蛋白分离及喷雾干燥工艺。首先研究了不同的离心转速和离心时间对鸡血浆蛋白分离效果的影响;使用SD-BASIC喷雾干燥器(英国Labplant公司),研究了喷雾干燥过程热风温度、进料速率对鸡血浆蛋白粉提取率、含水率、粗蛋白质量分数、氮溶解指数、颜色各项指标的影响。结果表明:离心转速和离心时间对血浆蛋白分离效果有显著影响(p<0.05);最佳分离条件为:离心转速3000r/min,离心时间15min。喷雾干燥热风温度、进料速率及两者之间的交互作用对鸡血浆蛋白粉品质均有显著影响(p<0.05);喷雾压力0.3MPa,进料温度为25℃,热风温度150℃、进料速率0.167mL/s时效果最好,此时血浆蛋白粉提取率、含水率、粗蛋白质量分数、氮溶解指数分别为64.289%、4.514%、72.644%、97.295%。研究结果为鸡血浆蛋白粉的工业化生产提供了相关的基础性数据。     

预热处理对甘薯蛋白酶解特性的影响

以碱性蛋白酶（Alcalase）为水解酶,研究了不同预热处理温度和时间对甘薯蛋白酶解特性的影响。采用单因素试验及响应面分析法,对Alcalase水解预热处理条件进行了研究,建立了水解度（DH）及三氯乙酸氮溶解指数（TCA-NSI）与预热处理的回归模型,分析了模型的有效性及因子间的交互作用。结果表明,预热处理对Alcalase酶解甘薯蛋白的DH和TCA-NSI的影响显著（P＜0.05）;甘薯蛋白预热处理的最优工艺参数为：温度92℃,时间13?min。在此条件下,DH与TCA-NSI分别为26.32%和84.33%。与直接酶解天然甘薯蛋白相比,在预热处理的最佳工艺条件下所得到的DH和TCA-NSI分别提高了15.89%和53.37%。     

番茄果实成熟衰老相关蛋白酶鉴定

为进一步研究蛋白酶与成熟衰老的关系,试验以‘918’番茄(Lycopersicum esculentum Mill.cv.918)果实为材料,采用改进的含明胶的十二烷基硫酸钠聚丙烯酰胺凝胶电泳方法对番茄果实中的蛋白酶进行检测,结果发现番茄果实中有3个蛋白酶,命名为SP-1、SP-2和SP-3,其活性伴随着果实后熟衰老发生变化。乙烯处理可提高番茄果实蛋白酶SP-2的活性,使SP-1和SP-3活性被提前检出,而1-甲基环丙烯处理对蛋白酶起相反作用。番茄果实蛋白酶受果实后熟衰老和乙烯的双重调控,是衰老相关蛋白酶。该文为进一步研究蛋白酶与果蔬衰老的关系提供参考。