黄河鲤谷胱甘肽对氨氮胁迫的生理响应

[目的]研究氨氮对鲤幼鱼肝组织谷胱甘肽及其酶系的影响,阐析谷胱甘肽对环境胁迫的抗氧化防御机理.[方法]将黄河鲤在对照组(0 mg/L)和低(10 mg/L)、中(20 mg/L)、高(30 mg/L)3个氨氮处理组中染毒,研究氨氮胁迫6、24和48h后对肝组织中谷胱甘肽(GSH)含量、谷胱甘肽过氧化物酶(GSH-Px)和谷胱甘肽转移酶(GST)活性的影响.[结果]氨氮胁迫诱导黄河鲤幼鱼肝组织中GSH含量、GSH-Px和GST活性迅速升高,并在氨氮胁迫24h达到最大值,从而抵御氨氮造成的氧化胁迫.随着氨氮胁迫时间的延长,肝组织中GSH含量、GSH-Px和GST活性降低,抗氧化能力减弱.[结论]氨氮胁迫诱导黄河鲤幼鱼肝组织中谷胱甘肽含量及其酶系活性改变,以有效清除细胞中的活性氧和有毒代谢物,提高鱼类对环境的适应性.     

胰岛激活蛋白B聚体的各亚基克隆及表达的初步研究

研究将百日咳杆菌用B-G培养基活化后再在改良的S-S培养基上液体培养,用CTAB-NaCl法提取百日咳杆菌的基因组DNA.经PCR扩增出胰岛激活蛋白 B聚体的四个亚基基因,分别将其克隆于pGEX-6p-1载体上,并探讨了在大肠杆菌BL21(DE3)中可溶性表达IAP B聚体各亚基的可能性.通过控制诱导表达温度,S2,S3均可以实现可溶表达,以GST亲和层析得到的S2终产物0.4 mg/L培养物, S3为0.6 mg/L培养物.S4表达虽是可溶性的,但由于融合蛋白的等电点近于7,多步纯化易损失.S5的表达量太低,有待进一步分析.     

Effect of E-cinnamaldehyde against Sclerotinia sclerotiorum on potato and induction of glutathione S-transferase genes

This research was carried out to assess the effect of E-cinnamaldehyde (EC) against Sclerotinia sclerotiorum, causal agent of potato white mold, under invivo and invitro conditions. Based on the results, EC was able to completely inhibit (P < 0.05) mycelial growth of four isolates in both volatile and contact phases after five days of inoculation at the concentrations 500 μl and 5 μl/ml, respectively. Moreover EC at concentrations 30 and 50 μl EC/ml sterile distilled water significantly reduced white mold on six potato cultivars including Pashandi, Istambouli, Agria, Marfauna, Alpha and Spartaan in greenhouse trials. In next step, induction of glutathione S-transferase (GST)-like genes identified from the pathogen (isolate 3) genome was assessed. After 2 h of exposure to both EC and cinnamom extract, four genes with locus numbers SS1G_07319.1, SS1G_07195.1, SS1G_00703.1 and SS1G_01918.1 were up-regulated. Furthermore, EC and cinnamon extract appeared to have no effect on gene expression in SS1G_10295.1. In this study, enzyme activity of GST was determined with 1-chloro-2, 4-dinitrobenzene as substrate. While enzymatic activity was 42.3 and 45.6 units mg⁻¹ protein in sterilized distilled water and ethyl acetate, respectively, it was recorded as 78.9 and 86.4 units mg⁻¹ protein for cinnamon extract and EC, respectively.     

抗GST单克隆抗体的制备及应用

用纯化的重组GST(谷胱甘肽S-转移酶)蛋白免疫BALB/C小鼠,取其脾细胞与SP2/0骨髓瘤细胞融合,对杂交瘤细胞及时筛选,阳性孔经三次有限稀释法克隆,成功获得4B3和1H10共2株能稳定传代并分泌抗GST单克隆抗体(McAb)的杂交瘤细胞.2株单克隆抗体腹水间接ELISA效价达10-6,抗体类型均为IgG1,κ链,经Western-blot分析表明,2株单抗与GST-IBV-S1-F融合蛋白及重组GST蛋白均具有特异性反应,验证这2株单抗可用于检测GST融合蛋白的表达情况;用1H10单抗制备的亲和凝胶柱可用于GST融合蛋白的纯化.     

Effect of methyl parathion and chlorpyrifos on certain biomarkers in various tissues of guppy fish, Poecilia reticulata

Tests of acute toxicity were performed on the most common species of aquarium fish, Poecilia reticulata. Guppies (P. reticulata) were exposed to progressive concentrations of methyl parathion (MP) and chlorpyrifos (CPF); a semi-static method according to guidelines of OECD was used. Tests of acute toxicity were conducted using 10 fish for each separate concentration and for the control group. The results were subjected to probit analysis to determine the 96 h LC₅₀ values. The 96 h LC₅₀ values of MP and CPF to P. reticulata were 8.48 ppm/L (5.98–10.89) and 0.176 ppm/L (0.313–0.224) respectively. In addition, behavioral changes at each concentration were observed for the individual fish. Fish were exposed for 96 h to different sublethal concentrations of MP and CPF (¼ LC₅₀, 1/8 LC₅₀ and 1/10 LC₅₀) and their oxidative stress-induction potential was estimated in brain, liver and gills of fish. MDA content is induced in all tissues but maximum rise was observed in gills (161% and 153% for MP and CPF respectively). With regard to antioxidant defense system (ADS), GSH level decreased in the brain, liver and gills of tissues of MP treated fishes (22%, 6% and 13% respectively) and showed increase in brain and gills CPF treated (23% and 21% respectively). CAT, GST, GR and SOD levels fluctuated in all treatment groups relative to the control. Brain AChE showed dose-dependent inhibition in fish exposed to the higher concentrations reached 45% and 66% for MP and CPF respectively. Collective findings demonstrated that pesticide exposure of fish induced an increase in MDA and fluctuated ADS along with inhibited AChE. These findings may be used as valuable biomarkers for evaluation of water pollution.     

苯醚甲环唑对大型溞慢性毒性和GST酶活性影响的研究

研究苯醚甲环唑对大型溞两代14d慢性毒性试验和子代恢复试验,观测了大型溞的存活率、生长速度、产卵量、产卵时间等生物学指标.并以苯醚甲环唑EC50浓度处理48 h,测定大型溞体内GST酶活力与时间和剂量的效应关系,探寻敏感性毒性指标.结果表明,苯醚甲环唑对大型溞的产卵时间、产卵量以及卵的孵化有明显的抑制作用,子代恢复试验发现存活下来的后代虽有一定的恢复,但是较高浓度处理组的一些敏感指标如产卵量、产卵时间等与对照组相比差异显著.苯醚甲环唑对GST酶活性影响随处理时间的延长和浓度的提高而逐渐增强,处理48 h后酶活力达到最高值.当苯醚甲环唑浓度为1.00 mg/L时,诱导率最大,达到324.84％,试验结果表明GST酶可以作为苯醚甲环唑对大型溞毒性影响的生物标志物.     

3个湖泊中黄颡鱼（Pelteobagrus fulvidraco）体内CYP1A1和GST的比较及其功效分析

环境中污染物生物标志物的选取和研究是环境监测的重要内容之一,作为污染物肝脏解毒作用的关键酶,多功能氧化酶（CYP1A1）和谷胱甘肽硫转移酶（GST）常被用来作为多种污染物监测的生物标志物。比较了3个湖泊（巢湖、洪泽湖和鄱阳湖）黄颡鱼（Pelteobagrus fulvidraco）体内的CYP1A1和GST的活性,并对其进行功效分析。结果显示,3个湖泊中黄颡鱼体内CYP1A1和GST之间呈现反相关关系;洪泽湖黄颡鱼体内CYP1A1值明显小于其他两个湖,而洪泽湖黄颡鱼体内GST值明显大于巢湖,与鄱阳湖相比差     

塔玛亚历山大藻对中国明对虾肝胰腺和鳃SOD、GST和MDA的影响

本文选择在我国分离得到的一株能典型产生麻痹性贝毒(PSP,Paralytic Shellfish Poison)的有毒赤潮甲藻塔玛亚历山大藻(Alexandrium tamarense,ATHK株),研究其是否能通过引发中国明对虾的脂质过氧化作用, 使机体产生过多活性氧自由基, 导致氧化损伤而发挥其毒性作用。塔玛亚历山大藻粗提液经肌肉注射方式染毒中国明对虾,于染毒后1、3、6、12、24和48 h 测定肝胰腺和鳃SOD,GST活力和MDA含量的影响。结果显示,染毒后 1-6 h, 对虾肝胰腺和鳃组织SOD,GST酶活力均增加,12和48h鳃组织的上述指标受到抑制。对虾肝胰腺MDA含量除1h外未见明显改变,鳃中MDA含量随时间增加呈升高趋势。研究表明：塔玛亚历山大藻粗提液在中国明对虾体内代谢过程中能诱导对虾产生活性氧自由基,塔玛亚历山大藻粗提液能诱导MDA含量增加,降低SOD和GST活力,能够引发鳃的脂质过氧化,对鳃造成氧化损伤。     

Cry8E亚致死浓度对马铃薯甲虫解毒酶和保护酶的影响

苏云金芽胞杆菌Cry8E杀虫晶体蛋白对马铃薯甲虫具有中等毒性,用亚致死剂量9和7μg/m L的Cry8E分别饲喂2、3和4龄的马铃薯甲虫幼虫,然后测定羧酸酯酶(Car E)、乙酰胆碱酯酶(ACh E)、谷胱甘肽-S-转移酶(GST)和过氧化氢酶(CAT)的酶活性。β-羧酸酯酶的活性随Cry8E浓度的升高明显提高,随龄期增长活性降低,酶活性在浓度间和龄期间差异都极显著。谷胱甘肽-S-转移酶活性下降,2龄幼虫体内的乙酰胆碱酯酶的活性明显升高,并且随龄期变化差异显著(P0.01)。3龄幼虫体内的CAT酶活性随Cry8E浓度的增加而显著升高(P0.01)。本研究阐明了Cry8E对马铃薯甲虫解毒酶和保护酶活性的影响,为理论研究奠定基础。