实验室培养条件下绒泡菌科四种黏菌个体发育比较研究

本文应用悬滴培养、燕麦-琼脂培养等方法及显微成像技术对绒泡菌科Physaraceae 4种黏菌（黄头绒泡菌Physarum flavicomum、淡黄绒泡菌Physarum melleum、垂头绒泡菌Physarum album、针箍菌Physarella oblonga）的个体发育特征进行比较研究,首次实现了垂头绒泡菌在实验室条件下的完整生活循环。4种黏菌孢子萌发均为V-型开裂,但萌发时间有所不同,黄头绒泡菌萌发所需时间最短,仅需5h,而针箍菌萌发所需时间最长,需要2d;针箍菌和淡黄绒泡菌形成的黏变形体均可转变为游动胞,而相同条件下黄头绒泡菌和垂头绒泡菌并未有游动胞形成;4种黏菌原生质团的生长速率不同,在光照刺激下针箍菌、淡黄绒泡菌、黄头绒泡菌和垂头绒泡菌的原生质团分别经过2-3d、2-3d、7d和13d,发育成子实体;完成孢子到孢子的整个生活史,针箍菌和淡黄绒泡菌需要25-30d,黄头绒泡菌和垂头绒泡菌需要40-45d。     

猪繁殖与呼吸综合征病毒M+N基因重组腺病毒载体的构建

本研究构建了M+N基因的重组腺病毒载体。首先用反转录聚合酶链反应(RT-PCR)的方法分别扩增出猪繁殖与呼吸综合征病毒M基因和N基因,将两者用口蹄疫病毒2A序列串联起来,将连接好的M+N基因插入腺病毒穿梭载体pAdTrack-CMV,经筛选获得了重组质粒pAdTrack-CMV/M+N;然后在大肠杆菌BJ5183内将此重组质粒和腺病毒骨架载体pAdEasy-1进行同源重组,获得了插有外源基因的重组腺病毒质粒pAd/M+N;最后,经PacⅠ酶切线性化后转染HEK-293细胞,成功获得含有M+N基因的重组腺病毒,为重组腺病毒活载体疫苗的研究奠定基础。     

Binding aspects of baicalein to HIV-1 integrase

Human immunodeficiency virus type 1 (HIV-1) integrase is an essential enzyme in the life cycle of the virus. It is responsible for catalyzing the insertion of the viral genome into the host cell chromosome. This integrase is an attractive target for the design of a HIV antiviral drug, because integrase has no human counterpart. In order to know the interaction mode of HIV-1 integrase with its inhibitor, we investigated the effect of the inhibitor, baicalein, on the conformation of the HIV-1 integrase catalytic domain [IN-(50-212/F185K)] using fluorescence and circular dichroism (CD) spectroscopy. We found that baicalein binds to the hydrophobic region of the HIV-1 integrase catalytic core domain. This binding of baicalein induces the conformational change of the enzyme. We also found that the binding ratio of baicalein to the HIV-1 integrase catalytic domain is 2:1.     

CAS agar diffusion assay for the measurement of siderophores in biological fluids

We developed a simple and universal method, by modifying the universal CAS (Chrome azurol S) assay, measuring siderophores in various biological fluids. We named the assay as CAS agar diffusion (CASAD) assay. CAS plate devoid of nutrients was prepared by using Bacto-agar (1.5%, w/v) as a matrix. Holes with 5-mm-diameter were punched on the CAS agar plate. Each hole was added by 35 microl of the test fluids containing Desferal that was twofold serially diluted. After incubating at 37 degrees C or room temperature for 4-8 h, the size of orange haloes formed around the holes was measured. The size of orange haloes correlated well with the concentration of Desferal in all the biological fluids tested in this study. CASAD assay showed consistent results in wide pH range from 5 to 9. Addition of iron to the test fluids containing Desferal decreased the size of orange haloes in a dose-dependent manner, which suggests that the CASAD assay detects only iron non-bound siderophore. These results suggest that CASAD assay would serve as a simple, stable, and highly reproducible test for screening and quantitative siderophore analysis in biological fluids.     

水稻转导外源总DNA变异后代盐胁迫下株高及生理指标分析

通过田间筛选的G系列水稻耐盐新品系生理生化指标和形态学指标的测定,进一步鉴定筛选的耐盐水稻的耐盐性.采用沙培法培养水稻幼苗,在水稻幼芽期和两叶一心期进行盐处理,设清水、0.3％ NaCl、0.6％ NaCl、0.9％ NaCl 4个浓度盐溶液浇灌植株.前者,待两叶一心期,测定丙二醛(MDA)含量、脯氨酸(Pro)含量、超氧化物歧化酶(SOD)活性和株高;后者,从浇盐水当天开始算起,分别测定第3天和第6天水稻的上述4个指标.结果显示,G16、G20这两个新品系在两个处理下都是随着盐浓度的增加,MDA含量逐渐降低,但在同一浇灌条件下均低于对照G6;Pro含量逐渐升高,但在同一浇灌条件下均高于对照G6;SOD活性先增强后减弱,但在同一浇灌条件下均高于对照G6;随着盐浓度的增加株高逐渐减小,但在同一浇灌条件下均高于对照G6.通过MDA含量、PRO含量、SOD活性和株高的显著变化,由此得出田间筛选出的水稻G16、G20的耐盐性都强于对照G6,从生理角度和形态学角度证实了田间筛选耐盐水稻结果的正确性,也证实了丙二醛含量、脯氨酸含量、SOD活性和株高可以做为水稻耐盐性鉴定的生理生化指标和形态学指标.     

凤蝶亚科(凤蝶科,鳞翅目)16S rRNA基因的分子系统发生分析

对15种凤蝶亚科蝶类线粒体16S rRNA基因部分序列进行了测定,并结合GenBank中其它相关类群的序列,采用邻接法(NJ)、最大简约法(MP)、最大似然法(ML)和贝叶斯法构建凤蝶亚科的分子系统树,探讨该亚科各类群间的系统发生关系.结果表明,燕凤蝶族构成凤蝶亚科蝶类系统树基部的一个独立分支;燕凤蝶族和裳凤蝶族为单系发生,且裳凤蝶族聚在凤蝶族内部;喙凤蝶族的单系性尚不能确定.综合分子系统学、形态学及寄主植物等相关证据,推测斑凤蝶类为凤蝶族中早期分化的一支;较之裳凤蝶类,斑凤蝶类可能更早从二者最近的共同祖先中分化出来.     

双歧杆菌质粒聚合酶基因对质粒载体稳定性的影响

目的探讨双歧杆菌天然质粒的聚合酶基因（Bifidobacterium plasmid polymerase,BPP）对人工构建的大肠埃希菌～双歧杆菌穿梭质粒载体（shuttle vector）在长双歧杆菌中稳定性的影响。方法首先电转化质粒pBADs—A和pBADs-BPP至长双歧杆菌,培养鉴定后,接种含质粒pBADs—A和pBADs-BPP的双歧杆菌于AMP^-和AMP^＋的MRS培养液中。厌氧培养后,将样品涂布于AMP^＋的MRS固体培养板上计数菌落数。结果相同条件下质粒pBADs·BPP组菌落数高于质粒pBADs—A组（P〈0．05）。结论BPP可以增加质粒载体的稳定性。     

Effect of germination temperature on characteristics of phytase production from barley

The effects of germination temperature on the growth of barley seedlings for phytase production were studied at 15, 20 and 25 degrees C for 6-10 days. The growth rate of the barley seedlings was increased as the germination temperature was increased. The initial rate of total protein production was closely coupled to that of the barley growth, and the rate of total protein production tended to increase as the germination temperature was increased. SDS-PAGE analysis of total protein from the barley seedlings showed time-dependent appearance and disappearance of protein bands. Although no significant phytase activity was detected at zero time of germination, a significant increase in phytase activity up to 7.9-fold occurred during the first several days of germination then decreased. Phosphate production (viz. phytate degradation) in the barley seedlings occurred rapidly at the beginning of germination. However, the rate of production continued to decrease with further germination. A time lag of about 1-2 days between the rate of total protein production and that of phytase production was observed. Unlike the extent of total protein production, that of phytase production was similar irrespective of germination temperature. Partial purification of a crude enzyme extract by hydrophobic interaction chromatography resulted in two phytase fractions (PI and PII). Zymogram analysis demonstrated that PI had two bands with molecular masses of about 66 and 123 kDa while PII had one band corresponding to a molecular mass of about 96 kDa. The optimal temperature for PI was found to be 55 degrees C, while it was 50 degrees C for PII. The enzyme fraction PI had a pH optimum at 6.0, whereas the optimum pH for PII was found to be 5.0. Addition of 0.1% (v/v) Tween 80 was found to increase enzyme activity significantly (i.e., 167% for PI and 137% for PII). Phytate in cereals including barley, rice, corn and soybean degraded effectively by the treatment of the barley phytases.     

多甲氧基黄酮对人胶质母细胞瘤细胞系生物学特性的影响

目的：多甲氧基黄酮（PMFs）因其抗氧化、抗癌、抗炎及抗动脉粥样硬化等多样的生物学活性受到国内外学者的广泛关注,但其在神经胶质瘤治疗中的潜在作用尚未见报道。本研究对多甲氧基黄酮处理人胶质母细胞瘤细胞系对其生物学特性的影响,初步探讨PMFs在神经胶质瘤治疗中的潜在应用。方法：不同浓度（0,20,40,60,80,100μg／mL）的PMFs处理人胶质母细胞瘤细胞系U251不同时间后．分别用Annexinv／PI双染法检测细胞凋亡的变化,MTT法检细胞活力的变化,Transwell小室实验检测细胞侵袭能力的变化。结果：随着多甲氧基黄酮的浓度及处理时间的增加,细胞的增殖明显受到抑制,同时诱导细胞大量凋亡,促使细胞生长停滞于G2／M期;此外,多甲氧基黄酮剂量依赖性的抑制胶质瘤细胞的侵袭。结论：PMFs能剂量和时间依赖性降低人胶质母细胞瘤细胞系U251的,同时显著诱导细胞瘤的凋亡和侵袭能力,提示PMFs可能对神经胶质瘤的高增殖性和侵袭性有一定的抑制作用,因此可能具有成为治疗神经胶质瘤药物的潜在应用价值。     

灵芝两个无孢品种中水提物成分分析及免疫活性比较

龙芝2号和鹿角灵芝均为赤芝的栽培品种,且在生长发育过程中均不产生孢子。目前,尚缺乏对两者化学成分和药理活性的系统比较研究。本研究以灵芝两个无孢品种——龙芝2号和鹿角灵芝为原料,研究对比两者子实体水提物成分差异及免疫活性的强弱。采用化学及仪器分析相结合的方法,分析两者水提物中的多糖得率、含量、重均分子量分布特征及核苷、蛋白质、氨基酸含量差异,并研究了灵芝两个无孢品种水提物样品刺激RAW 264.7细胞释放NO活性。结果表明,两者多糖得率及含量差异不大,但龙芝2号中重均分子量分布范围较广,多糖分子量较大,其含有3种多糖,分子量分别为2.021×10⁶、1.802×10⁶和4.825×10⁵,而鹿角灵芝中仅含有1种多糖,分子量为1.589×10⁴;两者中含有的核苷种类相似,但各核苷的含量存在差异;鹿角灵芝和龙芝2号中蛋白质含量分别为10.70%和10.32%,两者均不含有组氨酸,蛋氨酸在两者中含量均较高,分别达到2.556%和2.591%。从鹿角灵芝和龙芝2号中得到的水提物样品均具有体外刺激巨噬细胞释放NO的活性。