Survivin—siRNA真核重组质粒的构建及对前列腺癌DU145细胞增殖抑制作用

目的应用RNA干扰技术（RNAi）,构建针对凋亡抑制因子survivin的siRNA,并检测其对DU145细胞增殖抑制的影响．方法构建重组质粒shRNA-sur1和shRNA-sur2并分别转染DU145细胞株,通过MTT法和流式细胞术观察其对细胞的增殖抑制作用及凋亡的影响．结果成功构建针对survivin干扰的两种真核表达质粒,转染72h后,两个实验组细胞的生长速度分别是脂质体组的（37．2％±5．8％）（n=3）和（43．5％±7．2％）（n=3）．流式细胞术发现：实验组细胞的G1期细胞比例明显高于对照组,而G2期和S期细胞比例减少,细胞出现凋亡,与脂质体组比较,两实验组细胞的凋亡率分别为（21．70％±6．45％）,（14．835±5．65％）．结论成功构建针对survivin siRNA的真核重组质粒,两种质粒体外均可抑制DU145增殖并诱导其凋亡,为survivin介导的肿瘤基因沉默提供良好的基础．     

Marc-145细胞CD163基因克隆及其真核表达质粒构建

猪繁殖与呼吸综合征是危害养猪业重要传染病之一,CD163是猪繁殖与呼吸综合征感染易感细胞受体之一.从Marc-145细胞克隆出CD163基因并测序,再根据得到序列,应用Primer5.0软件设计引物,成功地扩增出带酶切位点CD163 ORF基因,通过Hind Ⅲ和Xho Ⅰ双酶切位点将CD163基因克隆入真核表达载体pcDNA3.1-myc-His中,获得重组质粒pcDNA3.1-CD163,并将重组质粒pcDNA3.1-CD163在BHK细胞上进行表达鉴定.结果表明,Marc-145细胞CD163蛋白在BHK细胞中得到了表达.这为进一步研究CD163受体在PRRSV感染Marc-145细胞过程中作用以及与其他受体之间相互作用提供试验数据.     

Effects of lupeol and miR-145-5p on proliferation and apoptosis of prostate carcinoma cells

AIM To investigate the effect of lupeol combined with microRNA-145-5p (miR-145-5p) on the proliferation and apoptosis of prostate carcinoma LNCaP cells. METHODS After hsa-miR-145-5p and lupeol were applied to LNCaP cells for 24, 48 and 72 h, the cell viability inhibitory rate was detected by MTT assay. PI single staining plus flow cytometry was used to detect the cycle distribution. The flow cytometry with annexin V/PI double staining and TUNEL experiment were used to detect apoptosis. Transwell method was used to detect cell migration and invasion abilities. Wound healing experiment was used to detect cell migration ability. The cell colony formation assay was used to calculate the colony formation inhibitory rate. RESULTS The cells in cell control group, non-specific control group and solvent group did not show effective LNCaP cell viability inhibition, migration inhibition and invasion inhibition, while the viability, migration and invasion abilities were significantly inhibited, and early apoptosis in vitro were induced in hsa-miR-145-5p group and lupeol group. In particular, the combination (hsa-miR-145-5p+lupeol) group showed more effective proliferation inhibition than the single-drug groups. CONCLUSION Both hsa-miR-145-5p and lupeol inhibit the proliferation, migration and invasion of LNCaP cells, and induce early apoptosis in vitro. Lupeol enhances the proliferation-inhibitory and apoptosis-inducing effects of hsa-miR-145-5p on LNCaP cells.     

Marc-145细胞接种PRRSV后多次收毒的探索

为提高单个Marc-145细胞的产毒数量,减少原材料的损耗,提高生产效率,对接种猪繁殖与呼吸综合征病毒(PRRSV)的Marc-145细胞在24h或36h收毒,随后补入不含病毒的维持液,12h或24h后再收毒,反复收毒4次。结果表明,数次收获毒液的TCID_(50)差异不显著,且与常规接毒细胞所产毒液的TCID_(50)基本相同,说明多次收毒在实际生产中具有应用前景。     

10kV柱上开关电器的分类与应用探讨

随着国民经济的高速发展、用电量不断增加的同时,客户对供电的可靠性及供电质量提出了更高的要求;10kV柱上开关电器在配网中分段和支线的合理应用,为提高供电可靠性作出应有的贡献.但是由于我国各地区发展极不平衡,配电网的结构与布局日趋复杂,各种技术水平的开关设备有着不同的应用.     

兔抗猪繁殖与呼吸综合征病毒血清在Marc-145细胞上对病毒复制影响

制备兔抗猪繁殖与呼吸综合征病毒抗血清,探索其对病毒复制影响,通过大量培养猪繁殖与呼吸综合征病毒XH-GD株病毒,高速离心后蔗糖梯度纯化浓缩并免疫新西兰大白兔,4免后采集抗血清,ELISA法测定效价。同时将抗血清与病毒共同孵育接种Marc-145细胞,荧光定量PCR和TCID50测定病毒复制情况。成功制备兔抗猪繁殖与呼吸综合征病毒抗血清,效价达1640,抗血清可以在m RNA水平和TCID50上降低病毒滴度。结果表明,兔抗猪繁殖与呼吸综合征病毒抗血清可以在Marc-145细胞上抑制猪繁殖与呼吸综合征病毒复制。     

马尾松扦插育苗造林配套技术试验

马尾松扦插造林配套技术试验结果表明：选择优良家系建立采穗圃，采取去梢促萌措施，能使采穗母树平均产穗条２０８条／株；地膜覆盖可使扦插苗生根率提高到８９．７６％，比无地膜覆盖提高１３．４７％；采用切根扦插苗造林，其成活率可达９５．３３％；扦插苗造林比实生苗造林的幼林树高增加２９．６２％，地径增加２８．５７％；通过无性系选优的幼林，１．５年生平均树高达１．３３ｍ。     

胚抢救技术在果树上的应用

胚抢救技术在果树早熟、无籽、三倍体类型育种及获得远缘杂种、阻止胚早期退化和缩短育种周期等方面具有重要作用。对胚抢救技术在果树作物育种中的应用及取得的成绩作了综述，并对胚抢救中应注意的几个问题，如适宜的培养时期、营养及环境条件作了简介，同时初步提出了果树胚抢救技术存在的不足及今后的研究方向。     

Establishment and Identification of One Marc-145 Cell Line Stably and Highly Expressing Porcine CD163

This study was attempted to generate one Marc-145 cell line stably and highly expressing porcine CD163 (pCD163) and set the foundation for PRRSV isolation and vaccine production.CD163 was shown to be a cellular receptor capable of mediating infection of PRRSV non-permissive cell lines.The pCD163 gene was amplified by RT-PCR from porcine alveolar macrophages and cloned into the eukaryotic expression vector pCI-neo, then the positive plasmid pCI-pCD163 was transfected into Marc-145 cells.After selecting with G418 and subcloning for 3 times, Marc-145 cell line expressing pCD163 was established.IFA results indicated that the fluorescence of pCD163-Marc cells was significantly brighter than Marc-145 cells;Western blotting results indicated that the pCD163-Marc cells could express higher levels of CD163 and the expression level was 8.7 times higher than Marc-145 cells.The pCD163-Marc cell line could be stably passaged for 20 passages and the expression level of CD163 was similar with different passages, which would be a valuable tool for facilitating virus propagation and vaccine production.     

探讨电力10kV电缆线路故障预防方法

电缆线路由于受到电缆及其附件质量、电缆施工质量及维护检修的影响,导致各种故障的产生,严重影响城市电网的安全运行。笔者结合自身的实际经验,分析电缆线路故障的原因,提出治理电缆线路故障的方法,保证电网的安全运行。