Gene expression of fatty acid-binding proteins, fatty acid transport proteins (cd36 and FATP) and β-oxidation-related genes in Atlantic salmon (Salmo salar L.) fed fish oil or vegetable oil

Relative gene expression pattern of fatty acid transport proteins (FATP and cd36), intracellular fatty acid-binding proteins (FABP3, FABP10 and FABP11), β-oxidation-related genes [carnitine palmitoyl transferase II (CPTII), peroxisome proliferator-activated receptor β (PPARβ), acyl-CoA oxidase (AOX), long-chain fatty acyl-CoA synthetase (FACS), acyl-CoA dehydrogenase (dehydrogenase)] and uncoupling protein 2 (UCP2) was assessed by RT-qPCR in Atlantic salmon muscle (red and white), liver, heart, myosepta and visceral fat. FABP11, a FABP isoform not previously described in Atlantic salmon, was highly expressed in visceral fat and myosepta and at the lower level in red muscle, white muscle, myosepta and heart. Furthermore, Atlantic salmon were fed either a diet containing fish oil (FO) or a complete replacement of FO with a vegetable oil blend (55% rapeseed oil, 30% palm oil and 15% linseed oil; VO) for the production cycle (27 months from start of feeding and until ∼4.5 kg mean weight). The expression of genes related to β-oxidation, fatty acid uptake and transport in the white muscle indicate ( n  = 3) significant down-regulation in VO fed Atlantic salmon and correlated with previously reported white muscle triacylglycerol stores and β-oxidation. FABP11 in visceral fat and myosepta was also down-regulated in VO fed fish.     

猪瘟病毒DNA疫苗的构建及动物免疫试验

利用DNA重组技术将猪瘟病毒（CSFV） C株E2囊膜蛋白全长基因克隆到真核表达载体pcDNA4.0的CMV启动子下游,采用磷酸钙转染法将重组质粒转入293T细胞,流式细胞仪(FACS)检测293T细胞瞬时表达了E2囊膜蛋白。将构建的重组质粒肌肉注射BALB/c小鼠,用流式细胞仪和酶联免疫吸附试验（ELISA）检测证明成功诱导小鼠产生了抗E2蛋白的抗体,为下一步利用DNA疫苗免疫小鼠研制抗猪瘟病毒单克隆抗体打下基础。     

逆转录病毒载体介导的猪瘟病毒E2基因在真核细胞中表达及动物免疫试验

利用DNA重组技术将猪瘟病毒(CSFV)石门株囊膜蛋白E2基因插入逆转录病毒载体pBABE-puro中构建成重组逆转录病毒载体pBABE-puro-E2,该重组逆转录病毒表达载体与pVSVg质粒经磷酸钙共转染法将其转入293GP细胞中包装逆转录病毒假病毒。用包装的假病毒感染SP2/0细胞,经嘌呤霉素筛选阳性细胞后进行流式细胞技术(FACS)分析,结果表明CSFVE2基因在SP2/0细胞膜上成功表达。将表达E2蛋白的SP2/0细胞腹腔免疫BALB/c小鼠,用流式细胞仪检测证明,成功诱导小鼠产生了抗E2蛋白的抗体。为下一步利用细胞免疫小鼠研制抗猪瘟病毒单克隆抗体打下基础。     

蛋白质定向进化的高通量筛选方法

定向进化是改造蛋白质分子的有效新策略。创建突变库的方法已经有很多而且比较通用,而建立有效的高通量的筛选方法是蛋白质定向进化成功的关键。本文综述了近几年发展起来的用于定向进化的高通量的文库筛选方法,介绍了各种展示技术及流式细胞分选技术的原理及其在文库筛选中的应用,分析了存在的问题及发展趋势。     

猪瘟病毒DNA疫苗的构建及动物免疫试验

利用DNA重组技术将猪瘟病毒(CSFV)C株E2囊膜蛋白全长基因克隆到真核表达载体pcDNA4.0的CMV启动子下游,采用磷酸钙转染法将重组质粒转入293T细胞,流式细胞仪(FACS)检测293T细胞瞬时表达了E2囊膜蛋白。将构建的重组质粒肌肉注射BALB/c小鼠,用流式细胞仪和酶联免疫吸附试验(ELISA)检测证明成功诱导小鼠产生了抗E2蛋白的抗体,为下一步利用DNA疫苗免疫小鼠研制抗猪瘟病毒单克隆抗体打下基础。     

Bovine Immunodeficiency Virus Expression in Vitro is Reduced in the Presence of β-Chemokines,MIP-1α, MIP-1β and RANTES

The inhibition of HIV expression in vitro by a cocktail of the -chemokines MIP-1, MIP-1 and RANTES provided the initial evidence that HIV utilizes chemokine receptors as co-receptors for infection of cells. Bovine immunodeficiency virus (BIV), a lentivirus, infects a wide variety of leukocyte populations, but the cellular receptor(s) utilized by this virus for infection of cells is not known. The purpose of this study was to determine whether MIP-1, MIP-1 and RANTES affect BIV expression in vitro, as a prelude to identifying the cellular receptors utilized by this virus. Fetal bovine lung (FBL) cells were pretreated with serial dilutions of a cocktail of the chemokines, and then the cells were infected with BIV. Virus expression in these cells was determined by counting the syncytia that had developed in the cultures by five days after infection. A significant decrease in syncytium formation, corresponding to increasing concentrations of the chemokines, was the result. Reacting the chemokines with chemokine-specific neutralizing antibodies prior to treatment of the cells neutralized the effect of the chemokines on virus replication in a dose-dependent manner, restoring viral expression to a level similar to that of untreated cells. The presence of a CCR5 homologue on the surface of FBL cells was confirmed using an anti-CCR5 monoclonal antibody and FACS analysis. Collectively, these data provide preliminary evidence that BIV may utilize the CCR5 receptor for infection of cells in vitro, but additional studies are necessary to confirm this.     

逆转录病毒载体介导的猪瘟病毒E2基因在真核细胞中表达及动物免疫试验初报

利用DNA重组技术将猪瘟病毒(CSFV)石门株囊膜蛋白E2基因插入逆转录病毒载体pBABE-puro中构建成重组逆转录病毒载体pBABE-puro-E2,该重组逆转录病毒表达载体与pVSVg质粒经磷酸钙共转染法将其转入293GP细胞中包装逆转录病毒假病毒。用包装的假病毒感染SP2/0细胞,经嘌呤霉素筛选阳性细胞后进行流式细胞技术(FACS)分析,结果表明CSFVE2基因在SP2/0细胞膜上成功表达。将表达E2蛋白的SP2/0细胞腹腔免疫BALB/c小鼠,用流式细胞仪检测证明,成功诱导小鼠产生了抗E2蛋白的抗体。为下一步利用细胞免疫小鼠研制抗猪瘟病毒单克隆抗体打下基础。     

猪瘟病毒E2基因在真核细胞中表达及抗E2蛋白单克隆抗体的制备

利用DNA重组技术将猪瘟病毒（CSFV）石门株囊膜蛋白E2基因插入逆转录病毒载体pBABE-puro中构建重组逆转录病毒载体pBABE-puro-E2,该重组逆转录病毒载体与pVSVg质粒经磷酸钙共转染法转入293T细胞中包装逆转录病毒假病毒.用包装的假病毒感染SP2/0细胞,经嘌呤霉素筛选阳性细胞后进行流式细胞术（FACS）分析,结果表明CSFV E2基因在SP2/0细胞膜上成功表达.将表达E2蛋白的SP2/0细胞腹腔免疫BALB/c小鼠,成功诱导小鼠产生了抗E2蛋白的抗体.取免疫小鼠脾细胞与SP2/0骨髓瘤细胞融合,经克隆和筛选获得了4株稳定分泌抗猪瘟病毒E2蛋白单克隆抗体的杂交瘤细胞株,所分泌的单抗可与CSFV产生特异性反应并具有中和活性.