Characterization of bovine ileal epithelial cell line for lectin binding,susceptibility to enteric pathogens,and TLR mediated immune responses

In this study, primary and immortalized bovine intestinal epithelial cells (BIECs) were characterized for the expression of surface carbohydrate moieties. Primary BIEC-c4 cells showed staining greater than 90 % for 16 lectins but less than 50 % staining for four lectins. Immortalized BIECs showed significantly different lectin binding profile for few lectins compared to BIEC-c4 cells. BIEC-c4 cells were studied for infectivity to E. coli, Salmonella enterica, bovine rotavirus, bovine coronavirus, and bovine viral diarrhea virus. Bovine strain E. coli B41 adhered to BIEC-c4 cells and Salmonella strains S. Dublin and S. Mbandaka showed strong cell invasion. BIEC-c4 cells were susceptible to bovine rotavirus. LPS stimulation upregulated IL-10, IL-8, and IL-6 expression and Poly I:C upregulated TLR 8 and TLR 9 expression. This study provides important knowledge on the glycoconjugate expression profile of primary and immortalized BIECs and infectivity and immune responses of primary BIECs to bacterial and viral pathogens or ligands.     

刺槐种子中凝集素的纯化及性质研究

从刺槐(RobiniapseudoacaciaL.)的种子中分离纯化出2种凝集素RPsAⅠ和RPsAⅡ。种子经磨碎、过滤、离心、硫酸铵分级、透析、Sepharose4B离子交换和SephadexG 150分子筛层析,即可获得在PAGE和SDS PAGE上均呈现单一蛋白染色带的凝集素纯品,分子筛层析测得RPsAⅠ和RPsAⅡ的分子量分别为132000和130000,SDS PAGE测得亚基分子量分别为38000和31000。当2种凝集素的质量浓度分别达到7 80μg/mL和15 60μg/mL时能凝集兔血红细胞,凝集活性均可被L 阿拉伯糖、D 果糖、D 半乳糖、麦芽糖抑制;热稳定性实验结果表明2种凝集素对热处理均具有一定的稳定性;当pH>9和pH<5时,它们的活性都明显降低。     

一个重要的精子受体—猪透明带糖蛋白β-D-半乳糖残基

 【目的】证明末端半乳糖残基在猪透明带糖蛋白中的分布以及它在精子结合和侵入中的作用。【方法】体外成熟的猪卵母细胞去除周围卵丘细胞，用异硫氰酸荧光素(fluorescein isothiocyanate，FITC) 络合西非单叶豆凝集素(Bandeiraea simplicifolia lectin-I，BS-I) 或蓖麻凝集素(Ricinus communis agglutinin，RCA-I)处理30min，分别观察末端-D-半乳糖残基或-D-半乳糖残基在透明带中的定位分布。荧光显微镜观察显示，BS-I和RCA-I都标记于整个透明带上且比较集中于透明带的外层，其中BS-I显示微弱荧光反应，而RCA-I呈强荧光反应且在透明带的外层反应尤为强烈。植物凝集素处理的卵母细胞用于体外受精，分别在受精2h或12h观察精-卵结合和精子入卵情况。【结果】RCA-I处理组卵母细胞平均精子结合数显著低于对照组（18 vs 95），而精子入卵则完全被阻断；可是BS-I处理组的卵母细胞的精子结合数少量被抑制（59 vs 95），而且精子入卵率与对照组无显著差异。【结论】猪透明带末端-D-半乳糖残基作为精子受体的重要组成成分，参与精-卵结合和受精。     

南美藜种子蛋白质研究

以西藏产南美藜Ｍ１为材料，对其种子蛋白质进行了研究，结果表明，南美藜种子蛋白质含量较高，达１５．５７％，富含赖氨酸（５．７１％），精氨酸（１０．０６％）等碱性氨基酸及天冬氨酸（７．６３％），谷氨酸（１１．６３％）等酸性氨基酸，种子蛋白质各组中心清蛋白（８．３６％）为主，占蛋白质总量的５３．６９％，南美藜种子总蛋白主要由６９，６６，５５，５２，５０，４１，３９，３６，３５，３２，２３，１８，１４，１     

柱状田头菇凝集素的分离纯化及其部分性质

经硫酸铵分级沉淀、活性炭柱吸附、DEAE-Sepharose FF离子交换层析及Sephadex G-100分子筛层析,从柱状田头菇子实体浸出液中分离到在PAGE上显示1条蛋白质染色带的凝集素ACL(Agrocybe cylindracea lectin)。经Sephadex G-100测得ACL分子量为67890,经SDS-PAGE测定,其亚基分子量分别为34673、20000和13183。ACL的中性糖含量为5．87％,氨基酸中天冬氨酸含量最高,半胱氨酸含量最低。ACL能凝集多种动物和人的A、B或O型红细胞,对鼠红细胞的血凝活性可被7种单糖和3种双糖抑制。ACL是一种对热较为稳定的凝集素。     

转基因大豆内外源DNA在少孢根霉发酵中的降解

试验旨在研究转基因大豆内、外源基因在少孢根霉发酵过程中的降解变化规律,并探讨其降解规律与相关酶活性的关系。试验采用定性和定量PCR检测各基因的降解变化,同时测定相关酶活。结果表明:内源基因Lectin和外源基因EPSPS在高温蒸煮下降解至800 bp以下,在发酵4 d后降解至200 bp以下;发酵过程中外源基因相对含量的变化与核酸酶、脂肪酶、蛋白酶活性的变化相一致。该研究为我国转基因食品加工过程中外源基因的降解及其机理研究提供一定的理论基础。     

Biochemical comparison of lectins among three different color strains of the red alga Kappaphycus?alvarezii

The red alga Kappaphycus alvarezii is economically important as an edible species and as a source of carrageenan, and has been extensively cultivated in many tropical countries. For this species, different color strains, which differ from each another in growth rate and carrageenan content, have been reported for decades. In this study, lectins from brown, red, and green strains of K. alvarezii cultivated in Vietnam were isolated and characterized for evaluation of their biochemical properties and contents. The results showed that each color strain contained in common the three lectins, named KAA-1, KAA-2, and KAA-3, which shared the hapten-inhibition profile of hemagglutination, 20 N-terminal amino acid sequence, and equivalent molecular mass within a range of 28,016 ± 1.2 to 28,021 ± 1.8 Da, but differed in their yields, with the highest yield of KAA-2. These properties of the three isolectins were also comparable among the three different color strains. However, the sum of the yields of the three isolectins decreased in the order: red (21.4 mg) to green (15.9 mg) to brown strains (15.1 mg), from 500 g fresh alga. Thus, this algal species can be a good source of useful lectins, irrespective of color strain.     

抗病虫基因新资源:海洋绿藻孔石莼凝集素基因

植物凝集素(Lecin)具有抵抗植物病原菌和农业害虫的活性以及抗病毒活性,在植物保护上具有广阔的应用前景。从海洋绿藻孔石莼中分离到一种凝集素基因序列,为培育抗病虫作物新品种提供了新资源。     

Taqman定量PCR技术检测转基因大豆中外源基因拷贝数

[目的]采用Taqman定量PCR技术检测转基因杂交大豆中外源,Ⅲ终止子基因的拷贝数。[方法]以大豆凝集素基因为内参照基因,以非转基因大豆基因组DNA为内参照基因标准品,通过梯度稀释法分别求取了内参照基因和质粒DNA的cz值与拷贝数对数值的相关性标准曲线方程,并通过将得到的0值代入标准曲线方程求取了样品的拷贝数。[结果]内参照基因标准曲线方程为Y=-3．422x＋35．201,R^2=O．998;外源基因标准曲线方程为Y=-3．348x＋34．890,R^2=0．999。nos终止子基因及其下游边界序列在转基因杂交大豆中为单拷贝。[结论]为确定转基因大豆外源基因拷贝数提供了理论依据。     

Hemagglutinins in vegetative tissues of Bengal gram

Hemagglutinating activity was identified in the roots and vegetative tissues of Bengal gram using rabbit erythrocytes. The activity in the roots appears to be similar to that of the seeds in respect to their sugar inhibition property. Seed and root extracts were found to have fetuin inhibitory agglutinating activity, whereas cotyledons showed different sugar specificity. However, 7-day cotyledons and roots from 3 and 7-day plants were also found to have galactose and N-acetyl-D-galactosamine inhibitory activity. Protein profile in different tissues during their developmental stages was analyzed by SDS-PAGE. A major band at 66 kDa was common to all tissues at different stages of their development except 3-day cotyledons where 2 major bands at 22 kDa and 74 kDa were observed. Two prominent bands at 74 kDa and 86 kDa were found along with 60 kDa protein in 3-day old roots whereas a major band at 33 kDa was found in 15-day old roots. Total protein content and specific activity of hemagglutinin were measured. Increase in specific activity with lower protein content was observed in cotyledons of 15-day old plant. Specific activity was eight times higher in the roots and four times more in leaf tissues of 7-day old plants in comparison to 15-day old plants. Stems also showed a significant increase of specific activity (p = 0.013) in 7-day old plants without any significant alteration in the protein content.