牛疱疹病毒Ⅰ型Bartha Nu/67株gD基因的克隆及在大肠杆菌中的表达

用PCR方法从感染牛疱疹病毒Ⅰ型Bartha Nu／67株的MDBK细胞中扩增gD基因，将之克隆到pGEM-T Easy载体。根据对gD基因的结构分析，设计合成3对引物，以获得的克隆质粒为模板，PCR扩增gD胞外域基因，将之分别亚克隆到pGEX-4T-2和pET-32a表达载体中构建成表达质粒pGEX-4T—gDQ、pGEX-4T—gDQB、pGEX-4T—gDHB和pET—gDQ、pET—gDQB、pET—gDHB，利用上述质粒分别转化大肠杆菌BL21和BL21（DE3），IPTG诱导后SDS—PAGE检测，结果pGEX-4T—gDQ、pGEX-4T—gDQB、pGEX-4T—gDHB和pET—gDQ四个表达质粒出现表达且符合预期大小，Western blot表明表达产物均有抗原性。     

微孢子虫糖蛋白的研究进展

随着蛋白质组学和糖组学研究的迅速发展,糖蛋白已成为众多研究的中心.糖蛋白的糖链结构直接影响其功能,因此糖蛋白中糖链的结构研究成为糖生物学研究的重点之一.就糖蛋白研究的基本策略方法作了简要介绍,并且简要概括了近年来微孢子虫糖蛋白的研究进展.     

Organic C and N stabilization in a forest soil: Evidence from sequential density fractionation

In mineral soil, organic matter (OM) accumulates mainly on and around surfaces of silt- and clay-size particles. When fractionated according to particle density, C and N concentration (per g fraction) and C/N of these soil organo-mineral particles decrease with increasing particle density across soils of widely divergent texture, mineralogy, location, and management. The variation in particle density is explained potentially by two factors: (1) a decrease in the mass ratio of organic to mineral phase of these particles, and (2) variations in density of the mineral phase. The first explanation implies that the thickness of the organic accumulations decreases with increasing particle density. The decrease in C/N can be explained at least partially by especially stable sorption of nitrogenous N-containing compounds (amine, amide, and pyrrole) directly to mineral surfaces, a phenomenon well documented both empirically and theoretically. These peptidic compounds, along with ligand-exchanged carboxylic compounds, could then form a stable inner organic layer onto which other organics could sorb more readily than onto the unconditioned mineral surfaces (“onion” layering model).To explore mechanisms underlying this trend in C concentration and C/N with particle density, we sequentially density fractionated an Oregon andic soil at 1.65, 1.85, 2.00, 2.28, and 2.55 g cm⁻³ and analyzed the six fractions for measures of organic matter and mineral phase properties.All measures of OM composition showed either: (1) a monotonic change with density, or (2) a monotonic change across the lightest fractions, then little change over the heaviest fractions. Total C, N, and lignin phenol concentration all decreased monotonically with increasing density, and ¹⁴C mean residence time (MRT) increased with particle density from ca. 150 years to >980 years in the four organo-mineral fractions. In contrast, C/N, ¹³C and ¹⁵N concentration all showed the second pattern. All these data are consistent with a general pattern of an increase in extent of microbial processing with increasing organo-mineral particle density, and also with an “onion” layering model.X-ray diffraction before and after separation of magnetic materials showed that the sequential density fractionation (SDF) isolated pools of differing mineralogy, with layer-silicate clays dominating in two of the intermediate fractions and primary minerals in the heaviest two fractions. There was no indication that these differences in mineralogy controlled the differences in density of the organo-mineral particles in this soil. Thus, our data are consistent with the hypothesis that variation in particle density reflects variation in thickness of the organic accumulations and with an “onion” layering model for organic matter accumulation on mineral surfaces. However, the mineralogy differences among fractions made it difficult to test either the layer-thickness or “onion” layering models with this soil. Although SDF isolated pools of distinct mineralogy and organic-matter composition, more work will be needed to understand mechanisms relating the two factors.     

马立克氏病病毒(MDV)糖蛋白gB在重组鸡痘病毒中的表达

将马立克氏病病毒gB基因用EcoR I从质粒pUR-MDVEgB中切下。两端补平后，插入到质粒pEFgpt12s的Pst I酶切位点。构建了含有完整的MDVgB基因的转移载体pEFgpt12s-gB。这个载体的特点是，它含有两个启动子，在强的复合启动子Spromoter的下游是插入的gB基因，另一个反向启动子vvP7.5的下游 是标记基因Ecogpt，它的两端是FPV的非必需区。携带gB基因的质粒pEFgpt12s-gB通过磷酸钙的方法转染用282e4株FPV感染3-4h的鸡胚成纤维细胞。通过药物筛选 ，得到含有gB基因的重组病毒。通过PCR、免疫荧光检测，证实了重组病毒中含有gB基因，并且gB糖蛋白也得到了表达。     

海蜇头糖蛋白超声辅助提取工艺研究

为提高海蜇头糖蛋白提取效果,采用超声辅助提取工艺,在单因素试验的基础上,采用超声处理时间、超声功率和提取时间三因素三水平响应面分析试验以优化此工艺条件.结果表明:海蜇头目标糖蛋白超声辅助提取的最佳工艺条件为超声处理时间15 min,超声功率300 W,提取时间60 min;在此工艺条件下,糖蛋白实际得率为9.14%,与模型预测值之间具有较好的拟合性.在3个因素中,超声处理时间,超声功率对糖蛋白得率的影响极显著,提取时间影响不显著,且相互之间无交互效应.     

旋毛虫抗原性质的分析试验

本试验应用聚丙安垂直板电泳分离,分析了旋毛虫抗原的组分。电泳后经考马斯亮蓝染色的旋毛虫幼虫抗原呈现２０条蛋白质色带,经Ｓｃｈｉｆｆ试剂染色呈现６条红色的糖蛋白色带。将电泳后的凝胶均匀分割Ａｇ１,Ａｇ２,Ａｇ３,和Ａｇ４４个组分。分别测定其蛋白质浓度为１２．３７μｇ,２２．５４μｇ,２６．３６μｇ,２６．０４μｇ。     

林蛙输卵管糖蛋白的纯化方法研究

林蛙输卵管糖蛋白是由输卵管上皮细胞分泌的一类高分子量的特异性蛋白质,对配子的成熟和运输、受精以及早期胚胎发育具有重要影响。长白山林蛙输卵管糖蛋白具有特殊药用价值和广泛的应用前景,目前对其提取和纯化工艺的研究也比较深入。简述输卵管糖蛋白的性质及生物学功能,着重论述分离纯化方法。     

牛传染性鼻气管炎病毒gD基因表达质粒免疫小鼠的试验研究

基因疫苗是近几年来发展起来的一种新型疫苗，具有使用安全，生产、运输、贮存方便，免疫效果稳定持久的特点。实验采用PCR方法将从洛阳种牛场西门塔尔牛精液中分离的牛传染性鼻气管炎(Infectious bovine rhino—traeheitis virus，IBRV)洛精株gD基因进行了扩增，经序列测定确证后构建了gD基因的真核表达质粒。将其单独或连同脂质体肌肉注射于BALB／c小鼠，应用ELISA定期检测血清抗体水平。结果表明，含IBRV gD基因的表达质粒能够引起小鼠的特异性血清抗体反应。     

猪瘟病毒cF114株E2基因在家蚕杆状病毒表达系统中的融合表达

为进一步利用家蚕杆状病毒表达系统研制猪瘟病毒(CSFV)新型的亚单位疫苗,将猪瘟病毒cF114株囊膜糖蛋白E2基因克隆至带有GST标签的分泌表达杆状病毒转移载体pAcSecG2T的EcoRⅠ和BamHⅡ位点之间,获得了带有杆状病毒囊膜蛋白gp67信号肽-GST-E2元件的重组杆状病毒转移载体pAcSecG2T-E2,与线性化家蚕杆状病毒Bm-BacPAK6 DNA共转染家蚕BmN细胞,筛选重组病毒(Bm-BacPAK6-E2),PCR证实所分离的重组病毒含有目的片段E2基因。SDS-PAGE图谱显示,感染重组病毒后,在细胞培养液上清和家蚕幼虫、蛹的血淋巴中均可检测到一条分子量为63 kD左右的特异性条带;感染Bm-BacPAK6-E2家蚕蛹的血淋巴可检测到GST活性,接种病毒148 h后重组蛋白的表达水平达到17.11μg/mL。     

玉竹糖蛋白的微波提取及抗氧化作用研究

[目的]微波辅助提取玉竹糖蛋白并对其抗氧化能力进行研究。[方法]以玉竹为原料,采用微波辅助提取糖蛋白,通过单因素和正交试验得到最佳提取工艺,并考察玉竹糖蛋白的抗氧化性。[结果]微波辅助提取玉竹糖蛋白的最佳工艺为料液比1∶30 g/m L、微波功率420 W、提取时间4 min、提取次数3次,在此条件下,验证玉竹糖蛋白得率为8.85%。玉竹糖蛋白具有显著的还原力和清除DPPH自由基的能力,但弱于VC。[结论]微波辅助提取玉竹糖蛋白是一种经济有效的办法,可为进一步开发玉竹产品提供依据。