H5N1亚型AIVHA抗原表位的高效表达及其抗原活性分析

血凝素（hemagglutinin,HA）蛋白是禽流感病毒（avian influenza virus,AIV）的一个重要表面抗原性蛋白,在疾病诊断和防治上有重要意义。本研究为了探讨一种更为简便有效的HA重组蛋白表达途径,利用生物信息学软件,对H5N1亚型AIVHA基因编码的氨基酸序列进行分析,在分析其在大肠杆菌中的密码子偏好性、稀有密码子分布情况及有关蛋白的抗原性等重要特性后,构建了HA抗原表位重组表达质粒pET-32a（＋）-HA。经测试,该重组质粒在1mmol/LIPTG诱导剂作用下诱导过夜,能在大肠杆菌Rosetta-gami B（DE3）中高效表达,并得到48.1kD大小的目的重组表达蛋白。重组蛋白用6×His-tagged protein纯化试剂盒纯化后,与福氏佐剂等量混合制备成抗原,以200μg/鸡的剂量皮下注射2月龄SPF鸡3次,采血分离血清。Western-Blot试验结果表明,该重组表达蛋白能分别与所制备的高免鸡血清及H5N1亚型AIV阳性血清发生特异性反应,在硝酸纤维素膜上出现特异性杂交带。说明本试验研究的HA抗原重组表达蛋白具有良好的免疫原性和反应原性,保留了HA蛋白的抗原活性,提示该重组蛋白在H5亚型AIV的防治技术研究中具有重要的实际应用价值。     

苜蓿秸秆配施玉米秸秆还田对氮矿化、微生物生物量和酶活性的影响

秸秆的质量是影响秸秆还田后氮素矿化的重要因子.玉米秸秆C/N比较高,还田后往往造成对土壤氮素的固持,因此秸秆直接还田要补施氮肥,而在如今的农业生产中为满足对粮食产量的需求,氮肥的施用越来越多,不仅增加了生产成本还危害了土壤质量.在秸秆还田条件下,如何能够更加科学合理地施用氮肥是一个关键问题.该试验在可控条件下,研究苜蓿秸秆和玉米秸秆共同还田对氮矿化、土壤微生物生物量碳氮和酶活性的影响,比较豆科作物秸秆或者氮肥配施玉米秸秆在相同C/N下(25:1)还田对土壤供氮、土壤微生物生物量和酶活性的影响.室内实验共设置4个处理:(1)对照(CK);(2)玉米秸秆单施(M);(3)苜蓿秸秆配施玉米秸秆调节C/N为25:1 (MM);(4)无机氮肥配施玉米秸秆调节C/N为25:1(MF),在25℃下共培养270 d.玉米秸秆单施在整个培养过程中均表现为对土壤氮素的固持作用,苜蓿秸秆与氮肥配施米秸秆均缓解了土壤微生物对土壤氮素固持作用,提高了土壤无机氮含量,苜蓿秸秆配施相比于氮肥配施延长了氮素的可利用性.苜蓿秸秆与氮肥配施提高了土壤微生物生物量碳氮含量以及转化酶和β-葡萄糖苷酶活性,并且以苜蓿配施处理最高.与玉米秸秆单施相比,苜蓿秸秆配施对脲酶活性影响没有差异,氮肥配施显著降低了脲酶活性.相关性分析表明,氮输入量显著影响了土壤无机氮含量、土壤微生物生物量和酶活性.相比于氮肥配施玉米秸秆还田,苜蓿秸秆配施玉米秸秆还田具有较高的土壤无机氮含量、土壤微生物量和酶活性.因此,我们认为相比于氮肥配施玉米秸秆还田,苜蓿秸秆配施玉米秸秆还田是一个比较好的农业生产实践模式.     

在灰潮土-芹菜系统中包膜尿素和尿素氮的利用和去向

通过盆栽试验研究了包膜尿素(自制)和尿素等肥料在灰潮土中对芹菜生长、氮吸收、产量和品质等的影响。结果表明：在2种质地的灰潮土中施包膜尿素较尿素使芹菜增产11．5％～15．2％,N吸收增加5．9％～9．5％,土壤氨挥发损失N减少12．2％～14．4％,淋失和反硝化N损失减少25．5％～28．3％,N素利用率提高20．1％～27．9％,土壤持留N量占施氮量的32．0％～37．3％;芹菜产品中NO3^--N含量降低44．2％～60．3％,维生素C含量显著提高,品质明显得到改善。     

丝状霉形体丝状亚种SC型脂蛋白LppQ N末端基因克隆与序列分析

脂蛋白LPPQ是丝状霉形体丝状亚种SC型(MmmSC)非洲株、欧洲株和疫苗株所特有的。LPPQ N末端域具有良好的免疫原性，在牛体内可诱导产生强大、特异、早期、持续的免疫反应。本研究根据已发表的LPPQ基因序列设计引物，用Pyrobest^TM高保真DNA聚合酶从MmmSC HVRI X株中扩增出了LPPQ N末端基因序列，并进行了克隆与序列测定。核苷酸序列比较结果显示，HVRI X株的LPPQ N末端基因序列与国外发表的序列同源性为99．7％，由其推导的氨基酸序列同源性为99，1％，为脂蛋白LPPQ N末端基因体外表达奠定了基础。     

西安蔬菜主产区土壤硝态氮累积现状与蔬菜产品、浅层地下水氮素污染调查研究

以西安郊区主要周边蔬菜产区为研究地点,通过分析蔬菜鲜样、菜田土壤2 m垂直分层土样、浅层地下水的水样,研究蔬菜及菜田生态系统氮肥污染现状.结果表明,按我国的地下水硝酸盐含量标准,在选点采样中1.5～3 m深的地下水硝酸盐含量超标,菜田土层土质疏松与浇水次数较多的菜田硝酸盐向下淋洗现象明显,温室较露地2 m深土层硝态氮与铵态氮分布浓度高,菜田2 m深土层中常有硝态氮与铵态氮的聚集峰,不同深度地下水的菜田韭菜硝酸盐含量差异明显.     

鸡传染性支气管炎病毒地方分离株S1、N和M基因的克隆与表达

摘要：将IBVSX16株的S1、N和M基因克隆入真核表达载体pVAX1,构建IBV真核表达载体pVAX1-16S1、pVAX1-16N和pVAX1-16M。体外转染BHK-21细胞,用间接ELISA检测到目的蛋白表达,用构建的3个表达载体免疫小鼠,从小鼠血清中检测到了抗IBV抗体,初步证实用IBVS1、N和M基因作为核酸疫苗免疫动物,可以激活机体免疫应答。     

氮肥对杂交小麦果聚糖积累与转运及其杂种优势的影响

氮肥影响作物的碳氮代谢及生长发育。选用６个杂交小麦及其7个亲本材料在施氮（200 kg /hm²）和不施氮（0 kg /hm²）条件下，比较研究了不同生育期不同器官果聚糖的积累与转运及其杂种优势。无论施氮与否茎鞘和总麦草中果聚糖的积累模式基本相似。茎鞘是贮存果聚糖的主要营养器官，分配了总麦草果聚糖积累量的63%～87%，并在开花期其积累量达最大值。总麦草转运的85%以上的果聚糖来源于茎鞘，其对籽粒产量的贡献率最大（9.42%~19.36%）。氮缺乏减少叶片中果聚糖的积累量，但增加茎鞘和总麦草中果聚糖的积累量。穗轴及颖壳、茎鞘和总麦草的果聚糖积累在挑旗期、抽穗期和开花期存在一定的杂种优势（4.7%~51.7%），叶片、茎鞘和总麦草的果聚糖转运量、转运率也具有较强的杂种优势，施氮增强叶片中果聚糖的转运优势，但减弱茎鞘和总麦草中的转运优势。结果说明，杂交品种比其亲本具有更强的果聚糖转运能力，施氮对茎鞘果聚糖的积累与转运及其杂种优势均有抑制作用。     

Effects of land-use type and nitrogen addition on nitrous oxide and carbon dioxide production potentials in Japanese Andosols

Land-use type and nitrogen (N) addition strongly affect nitrous oxide (N₂O) and carbon dioxide (CO₂) production, but the impacts of their interaction and the controlling factors remain unclear. The aim of this study was to evaluate the effect of both factors simultaneously on N₂O and CO₂ production and associated soil chemical and biological properties. Surface soils (0–10 cm) from three adjacent lands (apple orchard, grassland and deciduous forest) in central Japan were selected and incubated aerobically for 12 weeks with addition of 0, 30 or 150 kg N ha^–1 yr^–1. Land-use type had a significant (p < 0.001) impact on the cumulative N₂O and CO₂ production. Soils from the apple orchard had higher N₂O and CO₂ production potentials than those from the grassland and forest soils. Soil net N mineralization rate had a positive correlation with both soil N₂O and CO₂ production rates. Furthermore, the N₂O production rate was positively correlated with the CO₂ production rate. In the soils with no N addition, the dominant soil properties influencing N₂O production were found to be the ammonium-N content and the ratio of soil microbial biomass carbon to nitrogen (MBC/MBN), while those for CO₂ production were the content of nitrate-N and soluble organic carbon. N₂O production increased with the increase in added N doses for the three land-use types and depended on the status of the initial soil available N. The effect of N addition on CO₂ production varied with land use type; with the increase of N addition doses, it decreased for the apple orchard and forest soils but increased for the grassland soils. This difference might be due to the differences in microbial flora as indicated by the MBC/MBN ratio. Soil N mineralization was the major process controlling N₂O and CO₂ production in the examined soils under aerobic incubation conditions.     

A 15N tracing technique-based analysis of the fate of fertilizer N: a 4-year case study in eastern China

The fate of fertilizer N applied with different irrigation amounts in tobacco fields was quantitatively studied by applying ¹⁵N double-labelled NH₄NO₃ in lysimeters. The ¹⁵N (fertilizer N originating from the fertilizer applied in 2011) in tobacco plants, ¹⁵N in soils and ¹⁵N loss were observed continuously from 2011 to 2014. The results showed that 21.6% of ¹⁵N was utilized by tobacco plants, 72.1% remained in the 0–60 cm soil layer and 6.3% was lost from the soil–plant system after the first season’s harvest (2011) of flue-cured tobacco. During the four seasons from 2011 to 2014, cumulative utilization of ¹⁵N by tobacco plants was 34.3%, while 54.2% remained in the 0–60 cm soil layer, and 11.5% was lost via mechanisms such as leaching and volatilization. The fate of ¹⁵N in terms of accumulation in plants and soils or losses from the soil–plant system from 2012 to 2014 was greatly affected by the fertilizer and irrigation management strategies in 2011. The results of this investigation suggest that the major amount of fertilizer N applied during the first season remains available in the soil for utilization by tobacco plants after 4 years.     

氮肥后移对土壤氮素供应和冬小麦氮素吸收利用的影响

采用田间试验研究了氮肥后移对土壤氮素供应和冬小麦氮素吸收利用的影响。结果表明,与农民习惯施氮（N 300 kg/hm2,基肥和拔节肥各占1/2）比较,氮肥后移处理（N210kg/hm2,基肥、拔节肥和孕穗肥各占1/3）在不降低小麦产量的同时,大大提高了氮肥利用率,且全生育期氮素表观损失极低。过量施用氮肥(N 300 kg/hm2)明显提高了60 cm以下土层硝态氮含量,增加了其向地下水淋溶迁移的风险。氮肥后移可提高小麦成熟期0-20cm土层硝态氮积累量,降低其在20-100cm土层的积累。基于冬小麦不同生育阶段的氮素吸收量而进行氮肥后移是可行的,氮肥后移可节省氮肥30%,是较为理想的施氮方式。