作物籽粒中NO-3-N和NO-2-N的积累与氮肥种类和数量的关系

１９９６～１９９７在芸豆（云南）、红小豆（河北）、大豆（黑龙江）和花生（山东）绿色食品产地,研究了施用有机肥和化肥对这些作物籽粒中ＮＯ＾－３、ＮＯ＾－２的累积特点。结果表明,芸豆和红小豆不论施用有机肥还是尿素,土壤中的速效氮与籽粒中ＮＯ＾－２的灰色关联最好;施用有机肥的最大关联度分别出现在土壤库内部（全氮与速效氮之间）或籽粒库内部（ＮＯ＾－３、ＮＯ＾－２之间）;而施用化肥的最大关联度分别出现在土壤     

NH+4-N和NO-3-N肥对大棚黄瓜根际土线虫群落组成及其多样性影响

研究了两种形态氮肥,硝态氮(NO-3-N)和铵态氮(NH+4-N),对大棚黄瓜整个生长期内土壤线虫的群落组成及其多样性的影响差异.结果表明:在黄瓜的整个生育期内,NO-3-N和NH+4-N抑制了土壤线虫总量.在黄瓜各生长阶段,土壤线虫总量在NO-3-N和NH+4-N处理下,均无显著性差异.两种形态氮肥处理后,垫刃属和伪垫刃属仍为优势属,而小杆科从常见科变为优势科,其平均相对丰度为NO-3-N处理高于NH+4-N处理.土壤线虫群落中各营养类群较稳定,但植物寄生线虫与非植物寄生线虫出现频率的变化趋势相反.NO-3-N和NH+4-N处理后,植物寄生线虫出现频率的变化趋势由高到低;非植物寄生线虫出现频率的变化趋势则都是由低到高.在度量土壤线虫群落多样性的指标中,多样性指数(H′)、丰富度指数(SR)较好地反映了不同形态氮肥的施用对大棚黄瓜土壤线虫多样性的影响.与NH+4-N处理相比,NO-3-N更有利于提高土壤线虫的多样性和稳定性.     

蓝藻抗病毒蛋白-N衍生物的克隆、发酵与纯化

蓝藻抗病毒蛋白-N( Cyanovirin-N,CVN)能特异性与病毒表面糖蛋白结合,抑制病毒进入宿主细胞及抑制病毒感染细胞与未感染细胞的融合.通过分子设计及优化,在CVN的N-末端连接了5个氨基酸的柔性多肽(GGGGS),构建了SUMO-L5-CVN融合表达系统.SUMO-L5-CVN在大肠杆菌BL21中呈可溶性表达;通过表达条件优化,以0.5 mmol/L IPTG在20℃诱导24h 是最佳的诱导表达条件;SUMO-L5-CVN表达量占菌体总蛋白的30％;经Ni-NTA亲和层析获得融合蛋白SUMO-L5-CVN,SUMO蛋白酶酶切,以及进一步从Ni-NTA亲和层析获得的目的蛋白L5-CVN蛋白纯度＞98％.结果表明,低浓度的L5-CVN与流感病毒表面糖蛋白gp120就有较高的亲和力.     

原绿球藻固定化培养去除NH_4~+-N的效果

为了探究固定化微藻去除污水中NH4+-N的效果,以原绿球藻(Prochlorococcus)为藻种,采用褐藻胶包埋技术,进行了不同藻球密度(0、100×104、300×104、500×104、700×104、900×104cells·ball-1)、不同藻球用量(固定化藻球与人工污水的体积比V水/V藻球为4∶1、3∶1、2∶1、1∶1)及藻球加固工艺(Ca Cl2加固)等对比试验。结果表明:固定化藻比悬浮藻生长缓慢,去除NH4+-N的速率小于悬浮藻;固定化藻球藻细胞密度越高,去除NH4+-N的效果越好,但考虑单位藻细胞去除率,包埋密度以700×104cells·ball-1为宜;经N饥饿处理的藻细胞去除NH4+-N效果显著优于未饥饿组;固定化藻球用量大,去除NH4+-N越快,V水/V藻球为1∶1培养3 d后去除率可达100%;固定化藻球定期加固,有利于延长其使用寿命,增加对NH4+-N的吸收量。     

米曲霉中蓝藻抗病毒蛋白-N基因的克隆与表达

[目的]提高蓝藻抗病毒蛋白-N(CVN)的异源表达量,获得大量高纯度可溶性蛋白。[方法]采用RT-PCR从酱油发酵酱醪宏基因组中克隆获得CVN基因cvn-SF,构建了重组表达载体p ET32a-cvn-SF,转化E.coli BL21菌株,获得工程菌株,优化表达条件,通过Ni-NTA凝胶亲和层析对CVN-SF蛋白进行纯化。[结果]工程菌株在温度30℃、添加1 mmol/L的IPTG诱导剂、培养时间12 h的条件下获得最高表达量的可溶性蛋白。Ni-NTA凝胶亲和层析纯化得到了230.8 mg/L的CVN蛋白,占提取总蛋白含量的33.37%。[结论]所得CVN蛋白高于目前报道的CVN在大肠杆菌的最高表达量140 mg/L~([11])。     

酚-次氯酸钠显色法测定大鼠脑中游离氨-N含量

本文介绍一种用酚-次氯酸钠试剂直接测定动物组织样品中游离氨-N含量的方法,此法简便、灵敏、省时,最小检测量为0.05微克氨-N。酚-亚硝基铁氰化钠试剂:取酚0.6克、亚硝基铁氰化钠5毫克、加水至50毫升,放置24—48小时后使用。次氯酸钠试剂:Na_2HPO_4·12H_2O 1.79克、Na_3PO_4·12H_2O 1.9克、氢氧化钠0.5克、次氯酸钠原液1毫升,加水至50毫升,贮于棕色瓶内。硫酸铵标准液(1毫升=0.05     

Aerobic denitrification by novel isolated strain using NO-₂-N as nitrogen source

Biological denitrification reaction can be achieved under aerobic environment. Few aerobic denitrifiers using nitrite as sole nitrogen source were identified. Using nitrite as the sole nitrogen source, this work assessed the denitrification activity of yy7, an aerobic heterotrophic denitrifier identified as Pseudomonas sp. (94% similarity) by 16S rRNA sequencing analysis. The logistic equation describes the cell growth curve, yielding a generation time of 2.9h at an initial 18 mg l(-1)NO(-)?-N. Reduction of NO(-)?-N was primarily achieved during its logarithmic growth phase, and was accompanied by an increase in suspension pH and near complete consumption of dissolved oxygen. Three genes relating to nirK, norB, and nosZ were noted to involve in isolate strain. Isolate yy7 can survive and remove up to 40 mg l(-1)NO(-)?-N and, hence, can be applied as an effective aerobic denitrifier during simultaneous nitrification and denitrification via nitrite processes.     

钙网蛋白-N58在毕赤酵母中的表达及活性分析

目的：利用毕赤酵母表达系统表达Calreticulin-N58蛋白。方法：利用重叠延伸PCR方法合成Calreticulin-N58基因,构建成分泌型的重组表达载体pPIC9K-CRT-N58。该重组表达载体经SacI线性化后,电激转化入毕赤酵母GSll5,经MD板和不同浓度G418筛选得到多拷贝重组菌株。经甲醇诱导表达目的蛋白,SDS-PAGE分析重组蛋白的表达情况,在鸡胚中进行活性分析。结果：重组质粒pPIC9K-CRT-N58在毕赤酵母中经甲醇诱导能分泌表达CRT-N58蛋白,摇瓶表达量大约为60mg／L,纯化的目的蛋白有显著的血管生成抑制作用。结论：实验成功地在毕赤酵母表达系统中实现了Calreticulin-N58的分泌表达。为Calreticulin-N58蛋白的进一步药用研究奠定了基础。     

蓝藻抗病毒蛋白-N基因的克隆、表达、纯化及活性鉴定

蓝藻抗病毒蛋白-N(Cyanovirin-N,CVN)具有强效抗HIV及其他包膜病毒活性,该蛋白序列特殊,难以重组制备,在大肠杆菌细胞质中形成包涵体。本研究根据大肠杆菌密码子偏好性对CVN原始核苷酸序列进行优化,通过多次PCR合成SUMO-CVN的全长DNA序列,构建pET3c-SUMO-CVN重组表达质粒,重组质粒转化大肠杆菌BL21(DE3),获得表达菌株。通过对诱导剂浓度和诱导时间的优化,发现以0.5mmol/LIPTG在20℃诱导24h可获得最高表达,SDS-PAGE结果显示,SUMO-CVN为可溶性表达,表达量占菌体总蛋白的28%;经特异性的SUMO蛋白酶对融合蛋白进行酶切及两步Ni-NTA凝胶亲和层析可以得到纯度较高的重组CVN蛋白。ELISA结果表明,重组蛋白CVN与gp120蛋白有较高的亲和力。体外抗病毒活性实验表明,重组蛋白CVN在纳摩尔浓度具有很好的抗HSV-1和HIV-1/ⅢB活性;这为开发基于CVN的新型、高效抗病毒药物打下了基础。     

猪肺炎支原体Mhp366-N蛋白多克隆抗体的制备及应用

【背景】猪肺炎支原体是猪的一种重要的病原。该菌的研究工具较少,特别是缺少开展其致病机制研究需要的抗体。【目的】制备猪肺炎支原体Mhp366-N蛋白抗体并确定其应用范围和使用时的最佳稀释倍数。【方法】Escherichia coli BL21(DE3)-pET28a(+)-mhp366-N重组菌诱导表达Mhp366-N蛋白并纯化。纯化的蛋白免疫小鼠制备多克隆抗体。用免疫印迹和免疫荧光方法检测猪肺炎支原体AH株感染3D4/21细胞后的Mhp366蛋白,确定2种方法中Mhp366-N多克隆抗体的最佳稀释倍数;之后检测临床采集的猪肺泡巨噬细胞中的猪肺炎支原体;最后以免疫组化试验检测猪肺炎支原体感染的肺细胞。【结果】纯化的Mhp366-N蛋白纯度超过85%,免疫小鼠制备的抗血清效价在1:128 000-1:512 000之间。在免疫印迹试验中Mhp366-N多克隆抗体的最佳工作浓度为1:100 000稀释,免疫荧光试验中Mhp366-N多克隆抗体的工作浓度范围在1:1 000-1:10 000 000,其可用于临床采集的猪肺泡巨噬细胞和细胞系中猪肺炎支原体的检测。免疫组化试验结果显示猪肺炎支原体能够进入猪肺泡巨噬细胞、Ⅰ型和Ⅱ型肺泡上皮细胞。【结论】制备的Mhp366-N多克隆抗体为猪肺炎支原体致病机制研究提供了良好的研究工具。     

Influence of different mineral nitrogen sources (NO 3 − -N vs. NH 4 + -N) on arbuscular mycorrhiza development and N transfer in a Glomus intraradices–cowpea symbiosis

Labeled nitrogen (¹⁵?N) was applied to a soil-based substrate in order to study the uptake of N by Glomus intraradices extraradical mycelium (ERM) from different mineral N (NO ₃ ^? vs. NH ₄ ⁺ ) sources and the subsequent transfer to cowpea plants. Fungal compartments (FCs) were placed within the plant growth substrate to simulate soil patches containing root-inaccessible, but mycorrhiza-accessible, N. The fungus was able to take up both N-forms, NO ₃ ^? and NH ₄ ⁺ . However, the amount of N transferred from the FC to the plant was higher when NO ₃ ^? was applied to the FC. In contrast, analysis of ERM harvested from the FC showed a higher ¹⁵?N enrichment when the FC was supplied with ¹⁵NH ₄ ⁺ compared with ¹⁵NO ₃ ^? . The ¹⁵?N shoot/root ratio of plants supplied with ¹⁵NO ₃ ^? was much higher than that of plants supplied with ¹⁵NH ₄ ⁺ , indicative of a faster transfer of ¹⁵NO ₃ ^? from the root to the shoot and a higher accumulation of ¹⁵NH ₄ ⁺ in the root and/or intraradical mycelium. It is concluded that hyphae of the arbuscular mycorrhizal fungus may absorb NH ₄ ⁺ preferentially over NO ₃ ^? but that export of N from the hyphae to the root and shoot may be greater following NO ₃ ^? uptake. The need for NH ₄ ⁺ to be assimilated into organically bound N prior to transport into the plant is discussed.     

Could 5′-N and S ProTide analogues work as prodrugs of antiviral agents?

The nucleoside/nucleotide derived antiviral agents have been the most important components of antiviral therapy used in clinics. Recently, the focus of the medicinal chemists within this exciting research field has been affected mainly by the lack of effective therapies for the Hepatitis C virus (HCV) infection and several other “neglected” diseases caused by viruses such as Zika or Dengue. 2′-Methyl modified nucleosides and their monophosphate prodrugs (ProTides) have revolutionized the therapies for HCV in the last few years and, according to the latest research efforts, have also brought a promise for treatment of diseases caused by other members of Flaviviridae family. Here, we report on the design and synthesis of 5’-N and S modified ProTides derived from 2′-methyladenosine. We studied potential applicability of these derivatives as prodrugs of this archetypal antiviral compound.     

安普霉素的生物合成:辛二糖C7′-N上甲基的甘氨酸来源

安普霉素是一类结构特殊的氨基糖苷类抗生素,其分子中含一个稀有的辛二糖结构单元.迄今为止,尚无关于安普霉素生物合成途径及其前体化合物的完整报道.研究发现,向发酵培养基中添加甘氨酸和丝氨酸均可以出现在菌体生长保持基本不变的情况下促进抗生素产量的现象,表明甘氨酸和/或丝氨酸可能参与了安普霉素的合成.[2-13C]甘氨酸示踪实验的核磁共振(NMR)检测结果表明,甘氨酸专一地掺入安普霉素辛二糖环C7′-N甲基.同时发现,菌体胞内的S腺苷甲硫氨酸(SAM)水平上升与外加甘氨酸的量呈正比,但是甲硫氨酸的加入会抑制安普霉素的合成.据报道,甲硫氨酸对结构中含有甲基取代基的抗生素,如对rapamycin的生物合成中转甲基过程有抑制作用.由此推测,尽管甲硫氨酸本身对抗生素产量呈抑制作用,甘氨酸仍然可能通过甲硫氨酸循环提供甲基.此外,[2-13C,15N]丝氨酸的示踪实验结果表明丝氨酸也可能作为安普霉素的限制性前体物参与了NH2的合成.     

安普霉素的生物合成: 辛二糖C7′-N上甲基的甘氨酸来源

安普霉素是一类结构特殊的氨基糖苷类抗生素, 其分子中含一个稀有的辛二糖结构单元. 迄今为止, 尚无关于安普霉素生物合成途径及其前体化合物的完整报道. 研究发现, 向发酵培养基中添加甘氨酸和丝氨酸均可以出现在菌体生长保持基本不变的情况下促进抗生素产量的现象, 表明甘氨酸和/或丝氨酸可能参与了安普霉素的合成. [2-¹³C]甘氨酸示踪实验的核磁共振(NMR)检测结果表明, 甘氨酸专一地掺入安普霉素辛二糖环C7′-N甲基. 同时发现, 菌体胞内的S腺苷甲硫氨酸(SAM)水平上升与外加甘氨酸的量呈正比, 但是甲硫氨酸的加入会抑制安普霉素的合成. 据报道, 甲硫氨酸对结构中含有甲基取代基的抗生素, 如对rapamycin的 生物合成中转甲基过程有抑制作用. 由此推测, 尽管甲硫氨酸本身对抗生素产量呈抑制作用, 甘氨酸仍然可能通过甲硫氨酸循环提供甲基. 此外, [2-¹³C,¹⁵N]丝氨酸的示踪实验结果表明丝氨酸也可能作为安普霉素的限制性前体物参与了NH₂的合成.     

Design,synthesis and evaluation of carbamate-modified (−)-N1-phenethylnorphysostigmine derivatives as selective butyrylcholinesterase inhibitors

We synthesized carbamate-modified (?)-N¹-phenethylnorphysostigmine derivatives 3a–u and evaluated their anti-cholinesterase activities. In vitro evaluation showed that cyclohexylmethylcarbamate derivative 3u potently and selectively inhibits butyrylcholinesterase.     

71例Ⅲ-N2期非小细胞肺癌综合治疗病例的生存分析

目的:对71例Ⅲ-N2期非小细胞肺癌综合治疗的病例进行生存分析。方法:我院2007年3月至2014年3月诊断为Ⅲ-N2期的非小细胞肺癌患者71例,所有患者均接受手术、诱导或辅助化疗及术后放疗。采用Kaplan-Meier法对以上患者的总生存期(overall survival,OS)和无病生存期(disease free survival,DFS)进行分析。结果:71例患者的中位生存期为32个月,1年、3年和5年生存率分别为84.5%、49.6%和35.5%;1年、3年和5年无病生存率分别为70.4%、41.8%和27.4%;9例患者(12.6%)出现局部复发;鳞状细胞癌患者的中位生存期为43个月,而腺癌患者的中位生存期为21个月。鳞癌患者1年、3年、5年的OS和DFS分别为85%、60%、42.5%和80.0%、55.0%、34.2%;腺癌患者1年、3年、5年的OS和DFS分别为77.1%、42.0%、36.0%和60.0%、28.6%、28.6%,两组OS和DFS均无显著性差异。结论:不同细胞类型非小细胞肺癌综合治疗生存期存在差异     

Effects of NO 3 − -N on the growth and salinity tolerance of Tamarix laxa Willd

The influence of NO ₃ ^? -N on growth and osmotic adjustment was studied in Tamarix laxa Willd., a halophyte with salt glands on its twigs. Seedlings of T. laxa Willd. were exposed to 1 mM (control) or 300 mM NaCl, with 0.05, 1 or 10 mM NO ₃ ^? -N for 24 days. The relative growth rate of seedlings at 300 mM NaCl was lower than that of control plants at all NO ₃ ^? -N levels, but the concentrations of organic N and total N in the twigs did not differ between the two NaCl treatments. Increasing NO ₃ ^? supply under 300 mM NaCl improved the growth of T. laxa, indicating that NO ₃ ^? played positive roles in improving salt resistance of the plant. The twigs of T. laxa Willd. accumulated mainly inorganic ions, especially Na⁺ and Cl^?, to lower osmotic potential (Ψs): the contributions of Na⁺ and Cl^? to Ψs were estimated at 31% and 27% respectively, at the highest levels of supply of both NaCl and NO ₃ ^? -N. The estimated contribution of NO ₃ ^? -N to Ψs was as high as 20% in the twigs in these conditions, indicating that NO ₃ ^? was also involved in osmotic adjustment in the twigs. Furthermore, increases in tissue NO ₃ ^? were accompanied by decreases in tissue Cl^? and proline under 300 mM NaCl. The estimated contribution of proline to Ψs declined as with NO ₃ ^? -N supply increased from 1 to 10 mM, while the contributions of nitrate to Ψs were enhanced under 300 mM NaCl. This suggested that higher accumulation of nitrate in the vacuole alleviated the effects of salinity stress on the plant by balancing the osmotic potential. In conclusion, NO ₃ ^? -N played both nutritional and osmotic roles in T. laxa Willd. in saline conditions.     

烟草花叶病毒及其弱毒疫苗-N14基因组cDNA克隆的构建与侵染性分析

植物病毒弱毒疫苗在防治植物病毒病害中发挥着重要的作用,但对于其致弱的根本原因和机理仍不十分清楚。弱病毒能够在植物体内生存和繁殖,却不严重危害植物的生长、开花和种子生产,而对外来病毒具有杀灭和防治作用,这种共生和保护宿主的现象是大多数植物弱病毒所具有的共同…     

The effect of 17-N substituents on the activity of the opioid κ receptor in nalfurafine derivatives

We have previously reported the essential structure of the opioid κ receptor agonist nalfurafine hydrochloride (TRK-820) for binding to the κ receptor. In the course of this study, we focused on the effect of the substituent at 17-N in nalfurafine on the binding affinity for the κ receptor. The exchange of the 17-N substituent in nalfurafine from cyclopropylmethyl to fluoro-substituted alkyl groups, which are strong electron withdrawing substituents, almost completely diminished the binding affinities for the μ and δ opioid receptors, but the binding affinity for the κ receptor was still maintained. As a result, nalfurafine derivatives with 17-fluoro-substituted alkyl groups showed higher selectivities for the κ receptor than did nalfurafine itself. With regard to the κ agonistic activities, the conversion of the 17-N substituent in nalfurafine from cyclopropylmethyl to fluoro-substituted alkyl groups led to the gradual decrease of the agonistic activities in the order corresponding to their binding affinities for the κ receptor. In contrast, the derivative with the bulky 17-isobutyl group showed lower affinity and agonistic activity for the κ receptor than the derivatives with the smaller functional groups. This research suggested that both the electronic property and the steric characteristics of the 17-N substituent would have a great influence on the binding property for the κ receptor.     

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