猪传染性胃肠炎病毒spike蛋白亲和肽的原核表达     

于天飞  董慧莹  谢鹏宇 《西北农林科技大学学报(自然科学版)》2020,48(1):49-56

【目的】原核表达猪传染性胃肠炎病毒(transmissible gastroenteritis virus of swine,TGEV)spike蛋白亲和肽(SQHT),检测其病毒亲和性,为建立TGEV诊断方法奠定理论和物质基础。【方法】人工合成TGEV spike蛋白亲和肽基因,亚克隆后将该基因分别插入至原核表达载体pET-32a和pGEX-6p-1中,构建重组质粒pET-32a-SQHT和pGEX-6p-SQHT,对重组质粒进行BamHⅠ单酶切和BamHⅠ/XhoⅠ双酶切及测序鉴定。将重组质粒分别转化至大肠杆菌Rosetta(DE3),用IPTG进行诱导表达,对其表达产物的生物学活性进行检测。【结果】亚克隆获得了150 bp的TGEV spike蛋白亲和肽基因。成功构建了重组质粒pET-32a-SQHT和pGEX-6p-SQHT,并诱导表达出重组蛋白TRX-SQHT和GST-SQHT,其分子质量分别为25和31 ku。Western blot分析表明,2种重组蛋白与TGEV病毒粒子具有良好的亲和性;Dot-ELISA分析表明,2种重组蛋白与TGEV病毒粒子的最低结合滴度TCID_(50)为5×10~2 mL~(-1)。特异性试验表明,重组蛋白TRX-SQHT和GST-SQHT均可与TGEV产生特异性结合,而不与猪流行性腹泻病毒(PEDV)和猪轮状病毒(PoRV)结合。【结论】使用原核细胞成功表达了TGEV spike蛋白亲和肽,表达的重组蛋白具有很好的TGEV特异亲和性。  相似文献   

Enzootic bovine leukosis application of E.L.I.S.A. technique for the detection of antibodies     

P Maris  P Nougayrede  G Perrin 《Comparative immunology, microbiology and infectious diseases》1983,6(1):45-50

A simple microplate method of enzyme-linked immunosorbent assay (E.L.I.S.A.) for detecting antibodies to bovine leukosis virus (B.L.V.) is described. The antigen consisted of a solution containing the two major antigens of the B.L.V. (gp 51 and p 24) obtained by a technique of purification using CN-Br activated Sepharose 4B. This E.L.I.S.A. was compared with the agar gel immunodiffusion test (A.G.I.D.T.) in a study of 545 bovine sera. The total discrepancy rate between the two tests was 11% with a better sensitivity for E.L.I.S.A.  相似文献   

IDENTIFICATION OF FOUR LARCH SPECIES SEEDS WITH PEROXIDASE ISOENZYMES     

Guo WeimingNortheast Forestry University  Dept of Basie Courses  Nanjing Agricultural University Nanjing  P.R.ChinaCheng FengqinForestry Designing Institute of Hcilongjiang Province. 《林业研究》1992,3(2):41-46

The POD isoymograms of shelled seeds and embryos could be used to distinguish seeds of four specices of Larix.The period of sampling isozymes was critical and the two periods B and C in radical lengths of germinating seeds were shown to be optimal for sampling.Based on the number of common zones,the degree of similarity of isozymograms and double axises system or-dinations,the relationship of four species of the genus was also discussed.  相似文献   

Application of molecular recognition theory in thyrotropin and its receptor system     

LU Feng-xian  DAI Cheng-kai  TANG Te  YE Jing  WANG Wei 《园艺学报》2004,20(9):1587-1592

AIM: The selective recognition of the sense peptides which are located in special regions of thyrotropin receptor (TSHR) by their corresponding antisense peptides has been investigated. Three pairs of sense and antisense peptides were named TR1 (aa37-45) and RT1 (aa45-37), TR2 (aa353-366) and RT2 (aa366-353), TR3 (aa648-655) and RT3 (aa655-648). METHODS: To prepare three affinity chromatography columns, antisense peptides were immobilized, called RT1-sepharose 4B, RT2-sepharose 4B and RT3-sepharose 4B, respectively and investigate the retardative behavior for each of native peptide TR1, TR2 or TR3 on above columns with stepwise elution. RESULTS: Each of the three immobilized antisense peptides recognized and retarded its corresponding sense peptide-TR1, TR2 or TR3 instead of those non-complementary peptides. Immobilized RT1 recognized free TSHR protein molecule as well. In additional, bovine thyrotropin was recognized by immobilized TR1. CONCLUSION: The results indicate that molecular recognition theory exsits in thyrotropin receptor system. It may be useful to isolate biological molecules and to locate epitopes of TSH on TSHR molecule. Otherwise, antisense peptide may be used for treatment of experimental autoimmunolized thyroid disease (AITD) in the rat.  相似文献   

拟南芥NF-YC9重组蛋白的制备     

张晓娅  张国芳  司琪  崔素娟  赵红桃 《河北农业大学学报》2016,(2):15-20

NF-YC9是模式植物拟南芥核因子Y家族成员之一,在开花调控等过程中发挥重要作用,但其蛋白的特性及其作用的分子机制尚待探究。本文首先扩增获得了NF-YC9基因的序列,采用重组技术构建了与麦芽糖结合蛋白MBP编码基因融合的重组表达载体,将其转化到大肠杆菌BL21中,进行了MBP-NF-YC9融合蛋白的诱导表达,并借助直链淀粉亲和层析介质纯化技术获得了电泳纯的MBP-NF-YC9重组蛋白,以备进一步对NF-YC9蛋白的生化特性进行全面检测。  相似文献   

蛋壳残留溶菌酶的磁性亲和分离   总被引：2，自引：0，他引：2  

王炜军方颖  黄国林 《华南农业大学学报》2005,26(4):62-66

采用包埋法制备了具有磁响应性能的几丁质凝胶亲和吸附介质,并对其颗粒表观结构、吸附特性等物化特性进行了分析．试验表明：介质中的Fe3O4颗粒对杂蛋白存在非特异性吸附,而含醋酸铵的磷酸盐缓冲液的充分洗涤可有效消除非特异性吸附．利用此介质所建立的方法对蛋壳清洗液中的溶菌酶进行了分离．结果表明：通过外加磁场可快速地实现亲和吸附介质与粘稠清洗液及其中蛋壳碎片和泥沙等固形物杂质的分离,大大简化了操作步骤,酶活力回收可达26％,纯化酶对蛋白的比活性为50894U／mg,经SDS-聚丙烯酰胺凝胶电泳显示单一条酶蛋白带．  相似文献   

葡萄柚不同砧木嫁接亲和性与保护酶活性相关性分析   总被引：1，自引：0，他引：1  

严毅  张南新  何承忠  李贤忠 《北方园艺》2012,(11):8-11

以"哈路比"为接穗,分别嫁接在"曼赛龙"柚、美国酸橙、当地酸柚、枳壳砧木上,对不同砧木与葡萄柚品种嫁接愈合过程亲和性与保护酶活性的相关性及嫁接愈合过程中保护酶活性变化规律与嫁接成活的关系进行分析。结果表明:"哈路比"与4种砧木嫁接愈合过程中3种保护酶活性大小变化趋势为:愈合部>接穗>砧木;愈合部酶活较高的组合,其嫁接成活率也相应较高;相关性分析表明,愈合部保护酶活性大小与嫁接成活率一样均可作为嫁接亲和性的判定指标;综合多项指标发现,葡萄柚"哈路比"与"曼赛龙"柚嫁接亲和性最高,与美国酸橙和枳壳亲和性一般,与当地酸柚亲和性最差。  相似文献   

构建和谐教育方式，增强社会主义意识形态的吸引力和凝聚力     

叶飞霞 《福建农业大学学报(哲学社会科学版)》2008,11(1):9-12

通过解析调控教育场中诸多教育要素及相互关系，提出了坚持适应社会与引导潮流的和谐统一、社会教育目标与个人成才目标的和谐统一、尊重价值取向多样化和价值导向一元化的和谐统一、教育继承与创新的和谐统一、教育者“言教”与“行教”的和谐统一等5个方面要求，探索符合教育发展规律的和谐教育方式，以达到增强社会主义意识形态的吸引力和凝聚力的目的。  相似文献   

Charge Distribution and the Interactive Effect of pH and Ionic Strength on Phosphate Adsorption Properties of Two Benchmark Soils from Botswana     

《Communications in Soil Science and Plant Analysis》2012,43(22):2821-2836

The constraint imposed by phosphate (PO₄) deficiency on soil fertility in Botswana necessitated the study of ionic strength (μ) and pH effects on electric charge distribution and PO₄ adsorption of two representative soils: Typic Pellustert and Typic Haplustalf. Electric charge increased with increasing soil pH, but the effect of µ on electric charge distribution was indistinct. The point-of-zero-salt-effect (PZSE) occurred at pH 4.8 in the Pellustert and at almost 6.0 in the Haplustalf. Ionic strength decreased PO₄ adsorption in the Pellustert, but increased it in the Haplustalf. The Scatchard and reciprocal linearization of the Langmuir model adequately described PO₄ adsorption of the soils under varying µ and pH × µ conditions. The PO₄ adsorption maxima (M) and affinity index (b) decreased with increasing µ in the Pellustert, but not in the Haplustalf. The decreasing M and b with increasing µ in the Pellustert was explained by electrostatic repulsion from the highly negative charge surface, whereas the increasing M and b with increasing µ in the Haplustalf was attributed to enhanced access to the surface sorption sites due to compression of diffuse double layer. Under varying pH × µ conditions, PO₄ sorption sites were resolved into low- and high-energy sites, depending on the concentration of added PO₄. Because b is related to the ease of PO₄ detachment from soil surfaces into solution for plant uptake or leaching, PO₄ added to this Pellustert at µ > 0.01 would likely leach freely, and hence µ monitoring is imperative for PO₄ fertilization strategy of this Pellustert.  相似文献   

美国不同轻质集料的磷吸附特性比较     

M. D. BAKER  S. SIMKINS  L. A. SPOKAS  P. L. M. VENEMAN  B. S. XING 《土壤圈》2014,24(6):808-816

In this study, phosphorus(P) sorption of thirteen light-weight aggregates(LWAs) from USA was compared during batch equilibrium experiments in order to identify those materials which had the highest P sorption capacity for further study. Seven different levels of sorption activity were observed, which were broadly grouped into three categories—high performing, middle performing, and low performing aggregates. Chemical analysis of Ca, Al, Fe, and Mg was used to describe the differences between LWAs. There was a significant correlation between cation(especially Al, Ca, Fe, and Mg) content and P sorbed. Langmuir isotherms were used to describe P sorption maximum and binding affinity for four of the top five performing LWAs, Universal, Stalite "D", Stalite "Mix", and TXI.The fifth aggregate, Lehigh, exhibited more complex sorption, and was better described by the Freundlich isotherm. Universal had a mean P sorption at the highest concentration of 824 mg kg-1, well above its calculated sorption maximum(702 mg kg-1), and also had the highest binding affinity(1.1 L mg-1). This experiment suggests that the top performing LWA(Universal) may perform poorly in column and field studies due to observed precipitates, which could degrade constructed wetland performance. Other LWAs may exhibit superior field performance due to their high calculated sorption maxima. In general, these results highlight the importance of batch experiments as a first step in identifying materials suitable for column and field experiments.  相似文献