Expression of Anti-Lipopolysaccharide Factor Isoform 3 in Chlamydomonas reinhardtii Showing High Antimicrobial Activity     

Anguo Li  Ruihao Huang  Chaogang Wang  Qunju Hu  Hui Li  Xiao Li 《Marine drugs》2021,19(5)

Antimicrobial peptides are a class of proteins with antibacterial functions. In this study, the anti-lipopolysaccharide factor isoform 3 gene (ALFPm3), encoding an antimicrobial peptide from Penaeus monodon with a super activity was expressed in Chlamydomonas reinhardtii, which would develop a microalga strain that can be used for the antimicrobial peptide production. To construct the expression cluster, namely pH2A-Pm3, the codon optimized ALFPm3 gene was fused with the ble reporter by 2A peptide and inserted into pH124 vector. The glass-bead method was performed to transform pH2A-Pm3 into C. reinhardtii CC-849. In addition to 8 μg/mL zeocin resistance selection, the C. reinhardtii transformants were further confirmed by genomic PCR and RT-PCR. Western blot analysis showed that the C. reinhardtii-derived ALFPm3 (cALFPm3) was successfully expressed in C. reinhardtii transformants and accounted for 0.35% of the total soluble protein (TSP). Furthermore, the results of antibacterial assay revealed that the cALFPm3 could significantly inhibit the growth of a variety of bacteria, including both Gram-negative bacteria and Gram-positive bacteria at a concentration of 0.77 μM. Especially, the inhibition could last longer than 24 h, which performed better than ampicillin. Hence, this study successfully developed a transgenic C. reinhardtii strain, which can produce the active ALFPm3 driven from P. monodon, providing a potential strategy to use C. reinhardtii as the cell factory to produce antimicrobial peptides.  相似文献   

马传贫驴白细胞弱毒疫苗株跨膜蛋白主要免疫决定区基因的克隆与表达   总被引：3，自引：0，他引：3  

薛飞  贾斌  朱远茂  杨建德  王晓钧  相文华  赵立平  吕晓玲  沈荣显 《中国预防兽医学报》2003,25(5):321-325

从感染驴白细胞的马传贫驴白细胞弱毒疫苗株前病毒DNA中克隆了编码跨膜蛋白主要免疫决定区(TMIR)的基因，并在大肠杆菌中进行了表达。所表达的融合蛋白有一部分是可溶的，其氨基端带有6个组氨酸的标签，因此可以用固定化金属离子亲和层析法在非变性条件下进行纯化。在间接酶联免疫吸附试验(ELISA)和免疫印迹试验中，重组的TMIR蛋白可与马传贫阳性血清样品发生反应，而与健康马血清无任何反应。这表明该重组蛋白具有良好的抗原性和特异性，可用于马传贫弱毒疫苗株在体内外复制、接种马体内免疫应答及马传贫诊断的研究。  相似文献   

水飞蓟对动物机体免疫功能的影响研究   总被引：2，自引：0，他引：2  

何维明  薛振芳 《畜牧兽医学报》1995,26(2):147-153

将小鼠分为四组，分别饲喂１０％水飞蓟、２０％水飞蓟、环磷酰胺（０．０１ｇ／ｋｇ）和空白对照（常规饲料），研究对小鼠免疫功能的影响。实验结果证明，水飞蓟各剂量组能增高小鼠外周血白细胞数目（Ｐ＜０．０１、０．００１）；提高小鼠腹腔巨噬细胞的吞噬率（Ｐ＜０．００１，０．００１）及吞噬指数（Ｐ＜０．０５、０．００１）并随剂量加大有增强的趋势；能促进小鼠淋巴细胞转化，提高淋巴细胞转化百分率（Ｐ＜０．０１、０  相似文献   

PRRSV弱毒细胞疫苗与灭活疫苗生殖道黏膜免疫效果观察     

金光明  王珏  宁康建  吕锦芳  李升和  应如海 《勤云标准版测试》2005,(3)

应用组织学方法比较观察了猪繁殖与呼吸综合征病毒(PRRSV)弱毒细胞疫苗与灭活疫苗生殖道黏膜免疫的效果。试验选用5月龄长白×大约克夏二元杂交母猪18头。PRRSV弱毒细胞疫苗和灭活疫苗通过生殖道黏膜免疫接种。结果表明PRRSV弱毒细胞疫苗生殖道黏膜接种组母猪的子宫黏膜上皮内淋巴细胞数较灭活疫苗生殖道黏膜免疫接种组和对照组母猪显著减少(P<0.01);而且子宫黏膜上皮变矮变薄,变得有些不规则。灭活疫苗生殖道黏膜接种组母猪子宫黏膜上皮内淋巴细胞数与对照组母猪间差异不显著(P>0.05)。本试验结果显示PRRSV弱毒细胞疫苗经生殖道黏膜免疫接种则对生殖道黏膜上皮内淋巴细胞产生抑制作用。说明PRRSV弱毒细胞疫苗和灭活疫苗经生殖道黏膜免疫效果不理想  相似文献   

用单克隆抗体识别出一种与马立克病肿瘤细胞相关的淋巴细胞表面抗原     

《畜牧兽医学报》1995,(2)

  相似文献   

用酶联免疫吸附试验检测鸡白血病以及控制、根除鸡白血病措施的探讨     

李厚达 《扬州大学学报(农业与生命科学版)》1987,(4)

用酶联免疫吸附试验(ELISA),对2群鸡的鸡蛋清和1株鸡的马立克氏病疫苗进行检测,发现2群鸡的鸡蛋清中,鸡白血病病毒的阳性率分别是11％和29％,鸡马立克氏病冻干苗隐藏鸡白血病病毒群体特异性(gs)抗原的阳性率为100％。本文指出我国禽苗可能带有鸡的白血病病毒,分析讨论了鸡白血病病毒的垂直传递和水平传播的规律,提出在曾祖代和祖代鸡群中,采用ELISA试验,检测鸡蛋清,能减少以至根除鸡的白血病。  相似文献   

Development of a semi-selective medium and an immunofluorescence colony-staining procedure for the detection of Clavibacter michiganensis subsp. sepedonicus in cattle manure slurry     

N. J. M. Roozen  J. W. L. Van Vuurde 《European journal of plant pathology / European Foundation for Plant Pathology》1991,97(5):321-334

Various compounds and basal media were tested for their suitability to create a semi-selective medium for isolation ofClavibacter michiganensis subsp.sepedonicus (Cms) from cattle manure slurry containing c. 10⁸ colony forming units (cfu) per ml.Plating efficiency of Cms in yeast glucose mineral medium (YGM) was 104% compared with yeast peptone glucose medium. Nalidixic acid, polymyxin B sulphate and the experimental disinfectant S-0208 inhibited colony growth of cattle slurry bacteria as compared with Cms in YGM. The optimal concentration of these inhibitors in combination was determined by modified agar diffusion tests and by pour plating in 24-well tissue culture plates. The semi-selective medium YGMI consisted of YGM supplemented with nalidixic acid (2 mg/l), polymyxin B sulphate (30 mg/l) and S-0208 (125 mg/l). Plating efficiency varied for Cms between 50.9 and 69.6%, for cattle slurry bacteria between 1.8 and 2.5% and for saprophytes from potato heel end extracts between 11.5 and 27.4%.Differentiation of Cms colonies from other colonies was based on their small and bluish colony morphology in pour plates and on immunofluorescence colony-staining (IFC). IFC of a pure culture of micro colonies of Cms in YGM was possible after one day incubation (colonies c. 5 cells). Green background fluorescence in the agar gels was prevented by addition of Tween 20 (0.1%) to the washing buffer and the use of 1% agar gels. IFC of macro colonies of Cms in YGMI, visible with 4x objective magnification, was possible after 4 days. The detection level of the target organism in artificially inoculated cattle slurry in YGMI based on colony morphology varied between 1.4×10³ and 2.3×10⁴ cfu per ml of cattle slurry. Miniaturized plating combined with IFC, using wells in tissue culture plates (=16 mm), proved suitable for detection, but was c. 30 times les sensitive. The recovery of Cms was negatively correlated with the number of saprophytic colonies in the agar plates (R ²=0.74).  相似文献   

Biochemical properties of the pathogenesis-related proteins from tobacco     

J. F. Antoniw  R. F. White 《European journal of plant pathology / European Foundation for Plant Pathology》1983,89(6):255-264

This review describes the discovery and identification of the pathogenesis-related proteins (PRs) from tobacco. In crude leaf extracts the PRs are distinguished from the proteins in uninfected plants by their solubility at pH 3, resistance to a range of proteases, and mobility in polyacrylamide gels upon electrophoresis (PAGE) in non-denaturing conditions. PAGE has been used as a qualitative and semi-quantitative assay for PRs, and their migration in gels made from different acrylamide concentrations has been used to identify charge and size isomers and electrophoretically identical PRs in different tobacco cultivars. The subunit composition and molecular weight (mol. wt) of the four PRs identified first in Xanthi-nc were determined by SDS-PAGE; staining the gels has shown that these same four proteins in Samsun NN did not contain carbohydrate, lipid or nucleic acid, nor were they isozymic forms of twenty five enzymes known to increase in activity following infection with TMV. Evidence suggests that most of the PRs in Xanthi-nc and Samsun NN are extracellular.The purification of several PRs from Xanthi-nc, Samsun NN and other tobaccos is described, as well as their mol. wt, subunit and amino acid composition. PRs 1a, b and c consist of a single polypeptide and have similar mol. wt and amino acid compositions. Antisera prepared against purified Xanthi-nc b₁ protein have been used to determine serological relationships between PRs and form the basis of a very sensitive quantitative assay using ELISA. The regulation of synthesis of some PRs has been shown to involve translational control.  相似文献   

An improved ferritin assay for canine sera     

Andrews GA  Smith JE  Gray M  Chavey PS  Weeks BR 《Veterinary clinical pathology / American Society for Veterinary Clinical Pathology》1992,21(2):57-60

An improved serum ferritin assay for canine serum has been developed. It uses two monoclonal antibodies in a sandwich arrangement. Serum ferritin can be determined on undiluted canine sera with this assay. The recovery of ferritin added to canine serum ranged from 98 to 106%, the within-assay coefficient of variability was 3.3 to 4.5%, and the assay-to-assay variability was 9.8 to 10.2%. Serum ferritin from 61 apparently healthy dogs had a geometric mean of 252 ng/ml, with a range of 80 ng/ml to 800 ng/ml.  相似文献   

Serum lipase determination in the dog: a comparison of a titrimetric method with an automated kinetic method     

Walter GL  McGraw P  Tvedten HW 《Veterinary clinical pathology / American Society for Veterinary Clinical Pathology》1992,21(1):23-27

An enzymatic, kinetic method for determining serum lipase activity was evaluated and compared to a standard manual method for use in dogs. The kinetic method was a commercial kit adapted for use on a tandem access clinical chemistry analyzer and utilized a series of coupled enzymatic reactions based on the hydrolysis of 1,2-diglyceride by lipase. The manual method was the Cherry-Crandall technique based on the titration of base against the acid formed by hydrolysis of an olive oil substrate by lipase. The correlation between the two methods was very good (r = 0.94). The reference range for 56 clinically healthy dogs assayed by the kinetic method was 90 to 527 U/L. Diseases associated with a greater than twofold elevation in serum lipase activity as determined by the kinetic method included pancreatitis, gastritis with liver disease, and oliguric renal failure with metabolic acidosis. In some cases, pancreatitis was seen with other clinical problems, such as gastroenteritis, diabetic ketoacidosis, duodenal mass, disseminated intravascular coagulation, and septic peritonitis. Diseases associated with serum lipase activity within the reference range or elevated less than twofold included gastritis, gastric ulcer, cholestasis, phenobarbital-induced hepatopathy, colitis, copper hepatopathy, abdominal hematoma, apocrine gland adenocarcinoma, and thrombocytopenia with pneumonia.  相似文献