新城疫病毒和减蛋综合征病毒同鸭胚增殖的研究

鸭源新城疫病毒 ( NDV D10 )和减蛋综合征病毒 ( EDSV)可以在鸭胚中同时良好增殖 ,两种病毒的血凝价分别与单独接种时一致 ;同胚增殖病毒对鸭胚或 SPF鸡胚的感染能力和致病性分别与单独接种组无显著差异 ;利用同鸭胚增殖病毒的尿囊液制备的二联油乳剂灭活疫苗接种免疫鸡可以分别抵抗 NDV强毒和 EDSV强毒的攻击     

核酸探针检测禽呼肠孤病毒传播动态

用地高辛标记禽呼肠孤病毒(ARV)S1基因中编码σC蛋白的基因片段作为核酸探针,在斑点分子杂交中可检测到1.6pgARV的RNA。利用该核酸探针,通过检测鸡羽毛囊及体内病毒繁殖情况,比较研究了ARV感染后在鸡体内及鸡群中的传播动态。结果显示,研究建立的核酸探针检测方法灵敏度高、特异性强和操作简便,适于批量样品的检测。同时,用此方法检测发现,ARV感染24h后可侵染大部分器官,并且很快传播到同群未攻毒的鸡中,羽毛囊中的病毒检出率与鸡内脏器官中病毒的检出率一致。用核酸探针检测鸡羽毛囊中ARV的方法检测ARV的感染与流行情况,成本低,不影响鸡群生产。     

鸭微卫星标记的筛选及其遗传多样性分析

采用磁珠富集法从AFLP片段中筛选微卫星标记,并对5个福建地方鸭品种的遗传多样性进行检测。基因组DNA用EcoRⅠ/MseⅠ内切酶酶切的同时与接头连接,酶切连接产物与用生物素标记的探针杂交,应用磁珠捕获100~2000bp含有微卫星序列的DNA片段并通过pGEM-T载体转化到大肠杆菌DH5a感受态细胞中,构建富集微卫星序列的基因组文库。随机挑选46个阳性克隆,测序获得14条含有微卫星的DNA序列并递交到GenBank（登录号：FJ599499~FJ599512）。设计合成14对微卫星引物,通过PCR优化从中选择5对引物用于5个福建地方鸭品种的遗传多样性分析。结果显示,5对微卫星引物共检测到31个等位基因,各微卫星基因座的有效等位基因数为1.969 7~2.834 4,多态信息含量和杂合度的平均值分别为0.5133和0.7480,遗传多样性丰富,说明磁珠富集法适合用于鸭微卫星标记的分离与筛选,筛选得到的5个微卫星位点可作为有效的遗传标记用于福建省地方鸭品种遗传多样性的研究。     

用核酸探针检测鸡脏器及羽毛囊中禽呼肠孤病毒

摘要：用地高辛标记禽呼肠孤病毒（ARV）S1基因中编码σ3蛋白的基因片段作为核酸探针，在斑点分子杂交中可检测到1.6pg的ARV RNA。利用该核酸探针，通过检测鸡羽毛囊及体内病毒繁殖情况，比较研究了 ARV感染后在鸡体内及鸡群中的传播动态。结果显示，ARV感染后在24小时时就可侵染大部分器官，并且很快传播到同群未攻毒的鸡中。羽毛囊中的病毒检出率与鸡内脏器官中病毒的检出率一致。该研究证明用核酸探针检测鸡羽毛囊中ARV的方法检测ARV的感染与流行情况，具有灵敏、特异性高，操作方便的特点，尤其适用于大量样品的同时检测。     

用DNA-RNA杂交检测新城疫病毒的研究

用光敏生物素标记鸡新城疫病毒ｃＤＮＡ，制备核酸探针，经斑点杂交和碱性磷酸酶显色后，探针同该ｃＤＮＡ的ＰＣＲ产物、ＰＣＲ产物重组子和新城疫强弱毒株呈现阳性反应，与ＩＢＶ、ＩＬＴＶ、ＭＧ和正常尿囊液等均呈阴性杂交反应，田间病料的杂交试验也表明，该ｃＤＮＡ探针是敏感且特异的检测方法     

花生抗青枯病种质微卫星DNA的分离

提取抗青枯病花生材料基因组DNA,分别经Hae Ⅲ、Rsa Ⅰ酶切后,与生物素标记的6条微卫星探针杂交,富集微卫星DNA。测序分析后,得到非冗余序列180条,其中微卫星序列133条、小卫星序列47条,成功设计出141对引物。其中40对引物用于检测29份花生材料,发现5对为多态性引物,能很好地揭示野生种与栽培种四大类型的...     

检测四种鸡呼吸道疫病病毒可视化芯片技术的优化及初步应用

可视化芯片技术是在传统芯片技术的基础上发展起来的一项新的疾病诊断和基因分析技术,对于临床疾病检测和诊断具有重要意义。本研究针对禽流感病毒(Avian influenza virus,AIV)的核蛋白(nucleprotein,NP)基因设计一对特异性引物,以H9亚型禽流感病毒分离株c DNA为模板,经PCR扩增、连接、转化和核酸序列鉴定后,得到含有AIV-NP基因的重组质粒。同时复苏本实验室保存的3株分别含有新城疫病毒(Newcastle disease virus,NDV)的融合(fusion,F)蛋白基因、鸡传染性喉气管炎病毒(Infectious laryngotracheitis virus,ILTV)胸苷激酶(haemaggluttinin-neuraminidase,TK)基因和鸡传染性支气管炎病毒(Infectious bronchitis virus,IBV)核衣壳(nucleocapsid,N)蛋白基因的重组菌。以上述4种病原的靶基因核酸序列片段的正义链为模板,设计寡核苷酸探针,喷样到尼龙膜上制备成芯片,利用不对称PCR技术扩增生物素标记的靶基因,与芯片进行杂交后,检测结果直接用肉眼就可以判定。本研究对芯片制备流程和检测过程中主要条件进行优化。结果表明当寡核苷酸探针喷样浓度为25μmol/L、芯片杂交反应的时间为1 h、杂交温度为50℃、Streptavidin HRP Conjugate(1.0 mg/m L)稀释2 000倍、二氨基联苯胺(diaminobenzidine,DAB)显色时间为5 min时,芯片检测技术的结果最佳。用该方法与PCR/RT-PCR技术同时对临床采集的96份疑似病料进行检测,两种方法的检测结果一致。本研究构建的检测4种禽呼吸道疾病病毒可视化基因芯片技术具有高通量、快速、准确等优点,为鸡病的临床诊断提供了新的技术。     

~(125)I标记斜纹夜蛾(Spodoptera litura)病毒DNA探针的制备

用~(125)I碘化法标记斜纹夜蛾病毒DNA(SLNPV-DNA),所制备的~(125)I-DNA用于膜上DNA-DNA分子杂交实验。结果证明,~(125)I-SLNPV-DNA探针具有特异性,可供应用。     

结合不对称PCR技术构建鸡NDV-IBV-ILTV联合检测基因芯片

基因芯片(gene chip)是依据核酸杂交原理发展的一种生物新技术,在生命科学研究领域具有重要的应用价值.本研究将不对称PCR和基因芯片两种技术相结合,构建了同步检测鸡(Gallus gallus)传染性喉气管炎病毒(Infectious laryngotracheitis virus,ILTV)、新城疫病毒(Newcastle disease virus,NDV)、传染性支气管炎病毒(Infectious bronchitis virus,IBV)的共检基因芯片.分别选取ILTV的胸苷激酶(thymidine kinase,TK)和糖蛋白B(glycoproteins B,gB)基因、NDV的融合蛋白(fusion,F)和血凝素-神经氨酸酶蛋白(haemagglutinin-neuraminidase,HN)基因以及IBV的膜蛋白(membrane,M)和核衣壳(nucleocapsid,N)基因设计引物,从重组质粒菌中扩增制备探针基因,用乙醇沉淀法纯化后点制于氨基修饰的载玻片上,制备基因芯片;靶基因用cy3标记引物,进行不对称PCR扩增,扩增的荧光标记单链产物与芯片杂交.不对称PCR结果显示,当限制性引物与非限制性引物浓度比例在1:10时ILTV-TK、NDV-HN和IBV-N的单链产物增加最多,当浓度比在1:20时,ILTV-gB、NDV-F和IBV-M的单链产物增加最多;相应的标记样品与3种病毒检测芯片杂交后,均出现较强的杂交信号,而阴性对照检测不到荧光信号,灵敏性实验表明,当DNA浓度为1.8x 104拷贝时杂交仍为阳性.本研究构建的诊断基因芯片对12份临床样品进行初步应用检测,与PCR检测技术检出率基本一致.本实验所建立的联合检测基因芯片能够快速、准确、高通量的诊断NDV-IBV-ILTV,可以应用于集约化养殖业中对多种鸡疫病病毒的检测.     

地高辛(DIG)和碱性磷酸酶标记cDNA探针检测PRSV

根据GenBank发表的PRSV(番木瓜环斑病毒)中国Sm株系的外壳蛋白基因序列,设计特异引物,以美中红(Caricapapaya L.cv.Meizhonghong)带病样品的RNA为模板进行RT-PCR扩增,将其目的cDNA片段克隆到pGEM-Teasy质粒载体上。以重组质粒作模板,用PCR-DIG标记方法制备cDNA探针,另一种非放射性探针是经凝胶电泳分离、纯化cDNA片段,用碱性磷酸酶直接进行标记。利用地高辛(DIG)标记和碱性磷酸酶直接标记的cDNA探针核酸分子杂交及RT-PCR方法对PRSV进行了检测。结果表明,(1)测序结果表明美中红No.2序列与中国优势株系Sm的同源性为94.7%;(2)DIG标记的3种cDNA探针(861,455和215bp)对样品的总RNA检测结果一致,且861bp的探针杂交斑点最为清晰,上述核酸杂交结果与RT-PCR检测结果相符,核酸分子杂交检测的灵敏度和特异性能满足常规检测的需要;(3)碱性磷酸酶直接标记861bp的cDNA探针可进行PRSV的斑点杂交检测,可获得有效的杂交结果;(4)DIG标记探针对叶脉进行了印迹杂交检测,结果与RT-PCR检测结果相符。     

Endogenous organic acid variations in different chickpea (Cicer arietinum L.) genotypes

Abstract

Changes in cropping systems during the past century have led to selective pressure on weed flora. Species and ecotypes with characteristics enabling them to survive in high-input farmland have increased in numbers, at the cost of plants lacking these characters. Since the 1950s, the perennial weed species Cirsium arvense (L.) Scop. and Sonchus arvensis L. have mainly been controlled by the herbicide group synthetic auxins like MCPA. During recent decades, C. arvense seems to have become less susceptible to MCPA in both Europe and North America but the reasons are unclear.

To study the importance of selective pressure on weed ecotypes, both short- and long-term studies were carried out in Uppsala, Sweden. The first consisted of two growth-characteristic greenhouse experiments. The hypothesis was that ecotypes of C. arvense and S. arvensis from high-input farmland were different and displayed a more competitive growth pattern than did ecotypes from low-input farmland.

The second study was a field experiment with four ecotypes of C. arvense from low-input farmland to study if selective pressure was in force, over a period of six years. The four ecotypes had different growth characteristics and herbicide sensitivity and they were exposed to crop competition and MCPA treatments during the experimental period. The hypothesis was that ecotypes with a more competitive growth pattern and MCPA tolerance would survive to a greater extent than would other ecotypes. For C. arvense, the results from the growth-characteristic experiment showed that the growth pattern of ecotypes from high-input farmland differed, showing a more directly elongated growth pattern with fewer spines on the leaves compared with ecotypes from low-input farmland, which usually were of rosette-type. Results from the field experiment with C. arvense showed that after six years MCPA-sensitive and/or rosette-type ecotypes had almost disappeared while ecotypes with a more directly elongated growth pattern and less sensitive to MCPA survived to a much greater extent. The conclusion was therefore that when exposed to selective pressure like crop competition and herbicide treatments, ecotypes of C. arvense with a more directly elongated growth pattern and less sensitive to herbicide treatment survived to a greater extent compared with ecotypes missing these traits. Ecotypes from high-input farmland had generally fewer leaf spines than did ecotypes from low-input farmland. This may suggest a trade-off between spine formation and rapid competitive growth. In the growth-characteristic experiment with S. arvensis, no differences between ecotypes from high- and low-input farmland regarding growth characteristics or leaf spines could be detected. This might partly be due to a lower exposure of S. arvensis to selective pressure compared with C. arvense, since S. arvensis generally is less sensitive to MCPA.     

有机酸处理条件对玉米秸秆热解特性的影响

酸洗预处理能有效改善K^+对生物质热解的影响,该文利用热重分析仪和裂解-气相色谱质谱联用仪进行了玉米秸秆的热解试验,研究了不同有机酸酸洗浓度(3%、5%和7%)、酸洗温度(25、50和75℃)和酸洗时间(1、2和3 h)对玉米秸秆热解特性的影响。结果表明:酸洗能显著降低玉米秸秆内在K^+的含量;经过不同条件的有机酸洗预处理后,玉米秸秆的TG/DTG(thermogravimetry/differential thermogravimetry)曲线均向高温段移动,最大热解速率随着酸洗浓度和酸洗温度的增加逐渐增大,随着酸洗时间的增加先增大后降低,在酸洗温度为75℃时,最大热解速率达到最大值15.49%/min;与此同时,玉米秸秆热解主要产物为酚类、酮类和呋喃类化合物,酸洗后,其酚类物质产率明显增加,在酸洗浓度为7%时达到最大值16.75%,而酮类和呋喃类化合物产率减少,分别在酸洗时间为1 h和酸洗浓度为7%时达到最小值0.10%和7.13%。酸洗后,焦炭产率减少,在酸洗浓度为3%时达到最小值18.79%。通过研究不同处理条件下有机酸对玉米秸秆热解特性的影响,为生物质预处理中酸溶液的选择提供了参考。     

Organic Acids and Diffusive Flux of Organic and Inorganic Phosphorus in Sandy-Loam and Clayey Latosols

This study evaluated the effects of organic acids on the diffusive flux of phosphorus (DFP) along time. Treatments were a factorial 2?×?2?×?2?×?2?×?5 design: two soils (clayey Red Latosol and a sandy-loam Red-Yellow Latosol), two organic acids (citric acid, CA, and humic acid, HA), and two phosphorus (P) sources [monopotassium phosphate (KH₂PO₄) and inositol hexaphosphate (IHP)] either with or without heating (100 °C) of the soil. The soil plus treatments, in all combinations, were placed in contact with an anion exchange resin and the P in the resin was measured after 2, 4, 6, 10, and 15 days. The DFP was greater when the soils were treated with CA than when treated with HA. Citric acid was more effective in increasing the DFP from KH₂PO₄, whereas HA was the organic ligand that promoted a larger DFP when the source was IHP.     

Interactive effects of organic acids in the rhizosphere

Organic acids released into the rhizosphere may perform many beneficial functions to the plant including metal detoxification and enhancement of nutrient acquisition. Typically, these organic acids are studied in isolation; however, roots simultaneously exude a cocktail of organic acids and other substances, and their combined impact on rhizosphere processes may be quite different. It has been hypothesized that some exudates may play secondary roles (e.g. inhibitors of microbial activity, blockage of sorption sites), which might enhance the longevity and nutrient-mobilization capacity of others. Here we investigated how the decomposition, sorption and P-solubilizing effects of citrate, malate and oxalate are affected by the presence of malonate and shikimate. We found that in a range of agricultural soils the decomposition of citrate, malate and oxalate was rapid, but not influenced by the presence of large quantities of shikimate or malonate. This suggests that the individual organic acids are taken up by different transport mechanisms or components of the microbial community. At large concentrations, malonate decreased sorption of citrate, malate and oxalate on the soil, whilst shikimate had little effect. The capacity of citrate, malate and oxalate to desorb P was significantly greater in cocktails containing malonate compared with the single organic acid; no effect was seen with shikimate. We conclude that neither malonate nor shikimate at realistic concentrations will significantly affect the biodegradation of citrate, malate or oxalate in the rhizosphere, and while malonate did enhance P desorption, this effect is additive rather than synergistic. Overall, we found little evidence that malonate and shikimate act as secondary regulators of citrate, malate and oxalate behavior in soil.     

Low pH and aluminum tolerance of Bradyrhizobium strains isolated from acid soils in Indonesia

Sixty-seven strains of Bradyrhizobium isolated from soybean plants growing on acid soils in West Java and Sumatra, Indonesia, were examined for the effect of the pH and aluminum concentration on their growth in nutrient media, compared with 61 strains of Bradyrhizobium from soils in Japan. The results in this study indicated that the indigenous population of Bradyrhizobium in the soils of Indonesia showed a large difference in acid- and Al-tolerance from that of Japan. Eighty-five and 48% of the isolates from Japanese soils and Indonesian soils, respectively, were unable to grow in YEM broth at pH below 4.5. The acid-tolerance was correlated with AI-tolerance of the isolates on YEM agar plates at pH 4.4. Seventy-five percent of the isolates that grew in YEM broth at pH 4.5 were also resistant to 400 µM Al on the YEM plates. Acetylene reduction assay of the root nodules revealed that 3 of the acid- and Al-tolerant isolates from Indonesian soils showed a significantly high nitrogen fixation activity.     

Effect of humic acid and fulvic acid based liquid and foliar fertilizers on the yield of wheat crop

Abstract

An organic-mineral liquid fertigation fertilizer containing humic acid, nitrogen (N)–phosphorus (P)–potassium (K), zinc (Zn), sulfur (S), manganese (Mn) and a liquid foliar fertilizer consisting of fulvic acid and gibberellic acids were formulated and applied to wheat. The purpose of this study was to develop an efficient and cost effective liquid and foliar fertilizer and compare their potential with commercially available urea and DAP. The fulvic acid and humic acids were obtained by alkaline extraction of lignite coal. All the treatments were significant over the control. The application of the liquid fertigation fertilizer and liquid foliar fertilizer along with 50?kg of urea per acre showed the best results biological yield (grain plus straw yield). It was found that the highest yield was obtained in T₆ treatment and the second best results were obtained with T₂ treatment, but has a high cost, while treatment T₆ was the most significant economical and yielded a high income when compared to other treatments.     

Plant defence inducers rapidly influence the diversity of bacterial communities in a potting mix

Plant hormones and their functional analogues are used in agriculture to combat plant diseases and pests. These chemicals are generally targeted at shoots, but soils inevitably receive a dose during application. In this study, we used 16S rRNA gene amplicon pyrosequencing to determine whether the diversity of bacterial communities in a potting mix (referred here as ‘model soil’) is influenced by exposure to salicylic acid (SA), methyl jasmonate (MeJA), ethylene (ET) and abscisic acid (ABA). The number of operational taxonomic units (OTUs; richness) and Simpson's Diversity Index values (evenness) associated with each sample was not influenced by the hormone treatments, but changes in community composition were observed. Relative to the control, all hormones more than doubled the abundance of a Limnobacter population, ABA led to a four-fold increase in a Cellvibrio population, ET led to a c. 70% decrease in an Algoriphagus population, and MeJA and ABA applications halved the abundance of a Massilia population. These changes may influence plant productivity.     

不同来源腐殖酸的组成和性质的研究

本研究从三种有机物料草炭、褐煤、风化煤中提取腐殖酸,对其组成和性质进行了测定,同时对胡敏酸进行酒精沉淀分级。研究结果表明:褐煤胡敏酸氧化度和芳香度最高,其次为风化煤胡敏酸,最后是草炭胡敏酸;风化煤富里酸的氧化度和芳香度最高,其次为褐煤富里酸和草炭富里酸。胡敏酸各级分的回收率均随着级分数的升高而增加。回收率以风化煤胡敏酸最高,依次为褐煤胡敏酸、草炭胡敏酸。     

Soil Humic and Fulvic Acids from Different Land-Use Systems Evaluated By E4/E6 Ratios

Changes in land-use systems such as the introduction of trees on agricultural land can give rise to changes in the physicochemical properties of the soils, also affecting the quantity and quality of organic matter incorporated into the system. The present study assesses humic substances (HS) in the soil such as humic acid (HA) and fulvic acid (FA) by looking at the relationship between the optic densities determined at 465 and 665 nm (E4/E6 ratio). Topsoil samples (0–20 cm) from pine woodlands 60 years of age were compared with agricultural soils of similar age in the central-south region of the Province of Buenos Aires, Argentina. The pH of the topsoil from beneath the pine trees was highly acidic (5.0 vs. 6.2) and a significant increase in the level of organic carbon (OC) was registered. The carbon to nitrogen (C/N) ratio was also higher (by one order of magnitude) beneath the pine trees, although the humification conditions of the soil organic matter (SOM) were good in the soils of both studied land-use systems. The E4/E6 ratio was higher in the HA and FA 2º (second extraction) beneath the pines, indicating a smaller molecular size of the HS bound to the clay minerals. This fact can be attributed to the higher concentration of hydrogen ions beneath the pines and consequently the loss of polyvalent cations, mainly calcium. The most labile organic molecules (FA 1º – first extraction) were of a larger size in soils beneath the pines, most likely owing to a specific characteristic of the Pinus genus, although the fraction in question constitutes a minority fraction among the HS. Clear differences were established between the E4/E6 ratios in HA and FA, making this a highly useful method for determining molecular changes in HS as a result of changes in land use.     

Chemical Composition of Ten Blackcurrant (Ribes nigrum L.) Cultivars

Abstract

Ten blackcurrant cultivars were evaluated for some quality components. On average, the sugar content was 9.0%, of which fructose made up 45%, glucose 40% and sucrose 15%. Soluble solids were 15.4% as an average for the ten cultivars in three years. The acid content was 5.1% of fresh weight, 88% was citric acid and 12% malic acid. The ascorbic acid content varied from 67 mg per 100 g fresh fruit in Hedda, to 204 mg per 100 g in Blackdown. Except for Hedda and Øjebyn, which both had low values of ascorbic acid and colour properties, the quality components of the cultivars were found to be within an acceptable range.