黄瓜上甜瓜黄斑病毒寿光分离物的初步鉴定及序列分析

在山东寿光黄瓜保护地种植区采集到疑似感染甜瓜黄斑病毒(Melon yellow spot virus,MYSV)的黄瓜病叶,采用RT-PCR对其进行检测,将扩增产物连接到pEASY-T1 Simple克隆载体后进行测序,结果显示目的片段的大小为505bp,与预期结果一致;序列同源性比对分析结果表明,黄瓜上MYSV寿光分离物与中国三亚的MYSV-Sanya分离物(甜瓜-GQ397254)和中国台湾MYSV-TW(西瓜-FJ386391)亲缘关系较近,同源性可达到98%。     

Purification and characterization of glucose 6-phosphate dehydrogenase (G6PD) from grass carp (Ctenopharyngodon idella) and inhibition effects of several metal ions on G6PD activity in vitro

Glucose 6-phosphate dehydrogenase (G6PD) is a key enzyme catalyzing the first step of the pentose phosphate pathway which generates NADPH for anabolic pathways and protection systems in various organisms, including fish. In the present study, G6PD was purified from grass carp (Ctenopharyngodon idella) hepatopancreas using the methods of 2′,5′-ADP-Sepharose 4B affinity chromatography followed by DEAE Sepharose Fast Flow ion exchange chromatography. The characterization of G6PD and inhibition effects of several metal ions on G6PD activity in vitro were also determined. Grass carp hepatopancreas G6PD, with a specific activity of 18 U/mg protein, was purified 1,066-fold with a yield of 19.5 % and Mr of 71.85 kDa. The enzyme had a temperature optimum of 42 °C, pH optimum of 7.5 and 9.0. The K _m values for G6-P and NADP⁺ were determined to be 0.026, 0.0068 mM, respectively. The V _max values for G6-P and NADP⁺ were 2.20 and 2.27 μM min^?1 mg protein^?1, respectively. The catalytic efficiency for G6-P and NADP as the substrates was 0.085 and 0.334 × 10^?6 min^?1 mg protein^?1, respectively. Inhibition effects of metal ions on the purified G6PD activity indicated that IC₅₀ values of Zn⁺², Mn⁺², Al⁺³, Cu⁺², and Cd⁺² were 0.42, 0.54, 0.94, 1.20, and 4.17 mM, respectively. The Ki constants of Zn⁺², Al⁺³, Cu⁺², and Cd⁺² were 0.52, 1.12, 0.26, and 4.8 mM, respectively. Zn⁺², Al⁺³, and Cd⁺² showed competitive inhibition, while Cu⁺² inhibited the G6PD in a noncompetitive inhibition manner. Our study provided important information about the control of the grass carp liver PPP, the biosynthesis of several important related biomolecules, and the status of detoxification systems in grass carp liver in relation to metabolism.     

凤阳山常绿阔叶林土壤养分特性

对凤阳山自然保护区常绿阔叶林的不同坡向（东坡、西坡）与不同层次（0～10cm,10～30cm,30～60cm）土壤养分进行了测定分析。结果显示：东坡和西坡只在pH、铵态氮、有效磷方面表现出较强的差异性,其余差异均不显著,西坡和山谷在pH、铵态氮、全磷上存在差异显著;各养分因子中只有pH随着土层加深而增大,有机质、全氮、全磷和速效养分指标均表现出A层〉B层〉C层的趋势,且AB层、AC层差异显著;各营养元素间的相关性表现为,pH与多数养分均表现为极显著的负相关,有机质、全氮、水解氮、有效磷、速效钾之间多表现为极显著的正相关,全磷与全氮、C／N与有机质也表现为极显著的正相关。综合土壤主要肥力因子发现,东坡土壤质量优于西坡,能较好地满足植物生长对养分的需求。     

番茄内生拮抗细菌的分离鉴定及培养条件研究

针对番茄生产上灰霉病和叶霉病两大瘸害,为寻找安全、高效无污染的生防菌株及其最佳培养条件,本试验采用组织分离法从健康的番茄植株中分离出642个内生细菌菌株,并采用平板对峙法筛选出对番茄灰霉病菌和叶霉病菌拮抗作用强且稳定的两个菌株Thyy1和Jcxy8。通过形态学观察及生理生化特征测定,初步鉴定Jcxy8属环状芽孢杆菌（Bacillus circulans）,菌株Thhy1属枯草芽孢杆菌（Bacillus subtilis）。内生拮抗细菌在以豆饼粉为原料的6号培养基中生长速度快,发酵滤液对两种病原菌的抑制作用强。培养基初始pH值、培养时间、温度、通气量等对菌株生长及其抗菌物质的分泌有明显影响。以豆饼粉培养基、初始pH6．7、培养时问48h、温度30qc、并尽量增大培养通气量为菌株的最佳培养条件。     

小麦谷氨酰胺合成酶同工酶转录特点及其启动子序列分析

谷氨酰胺合成酶是小麦氮同化关键酶,分为胞液型和质体型(TaGS2)两类,其中胞液型TaGS又分为TaGS1、TaGSr和TaGSe。为了研究异源六倍体小麦A、B、D染色体组TaGS同工酶表达差异及调控机制,本项目利用三代测序技术测定了TaGS同工酶基因转录水平,依据中国春基因组序列克隆了豫麦49的12个TaGS同工酶启动子,并对其序列进行了分析。结果表明,TaGS1主要由6B染色体基因转录,TaGSe和TaGSr主要由4D染色体基因转录,TaGS2主要由2D染色体基因转录,不同TaGS同工酶转录起始位点距起始密码子ATG的距离不同。启动子元件分析显示,6B染色体上的TaGS1启动子有较多W-box、AC-I、ABRE、as-1和茉莉酸甲酯等响应元件,4D染色体上的TaGSe启动子有较多胁迫响应转录因子(MYB、MBS、LTR等)结合元件和植物生长素响应元件,4D染色体上的TaGSr启动子有较多WRE3等转录因子结合元件,2D染色体上的TaGS2启动子有较多A-box、WRE3、ARE及AT富集区。不同TaGS同工酶启动子顺式元件种类、数目及排列顺序均不同,为进一步研究GS同工酶调控机制奠定了基础。     

内生环状芽孢杆菌Jcxy8对番茄灰霉病的防病机制研究

为明确从番茄植株体内筛选出的内生环状芽孢杆菌(Bacillus circulan)Jcxy8对番茄灰霉病菌的抑菌作用及防病的生理生化机制,采用平板打孔法测定了菌株Jcxy8对灰霉病菌(Botrytiscinerea)的拮抗力。结果表明:菌株Jcxy8对灰霉病菌的抑菌圈直径为35.6mm,抑菌圈边缘的产孢抑制率达到66.9%。当菌株培养滤液浓度为40%时,病菌孢子萌发完全被抑制。镜检发现抑菌圈周围的菌丝(或芽管)细胞消融,生长扭曲,中间或顶端膨大成泡囊状。Jcxy8菌株与灰霉病菌同时处理的番茄果体内可溶性蛋白含量比清水对照处理高12.7%,比单独接种灰霉病菌处理高39.1%;SOD、POD、CAT活性均较只经病菌处理低;O2-产生速率比清水对照和病菌处理低,而比菌株Jcxy8处理高。说明菌株Jcxy8对番茄果实有明显的诱导抗病作用。     

In vitro exposure to copper influences lipid metabolism in hepatocytes from grass carp (Ctenopharyngodon idellus)

In the present study, three different copper (Cu) concentrations (control, 10 and 100 μM, respectively) and three incubation times (24, 48 and 96 h) were chosen to assess in vitro effect of Cu on lipid metabolism in hepatocytes of grass carp Ctenopharyngodon idellus. Increased glucose 6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase and carnitine palmitoyltransferase I activities were observed in hepatocytes with increasing Cu concentration and exposure duration. Cu decreased mRNA levels of several lipogenic and lipolytic genes at 24 h. However, at 48 h, Cu down-regulated the process of lipogenesis but up-regulated that of lipolysis. The Cu-driven up-regulation of lipolytic genes was maintained after 96 h and accompanied by a decreased intracellular triglyceride accumulation, while no effect on lipogenic genes was shown. Thus, 96-h Cu exposure induced lipid depletion, possibly due to the up-regulation of lipolysis. Although in this process, lipogenesis might be up-regulated, it was not enough to compensate lipid consumption. Our study represents the first approach to concentration- and time-dependent in vitro effects of Cu on lipid metabolism of fish hepatocytes and provides new insights into Cu toxicity in fish at both enzymatic and molecular levels.