为了明确成熟度对豫西地区烤烟上部烟叶质量的影响,以河南三门峡地区主栽品种云烟99为试验材料,分析了不同成熟度(M1～M4)鲜烟叶的质体色素含量、碳氮代谢相关酶活性等生理指标以及烤后烟叶的内在化学成分、中性致香物质含量和感官质量的差异.结果表明,随着成熟度的增加,烟叶质体色素含量逐渐降低,至M4成熟度时叶绿素总降解量达0.69 mg/g.与氮代谢相关的硝酸还原酶、谷氨酰胺合成酶活性均呈先升高后降低的变化趋势,与碳代谢相关的淀粉酶活性呈逐渐下降的趋势,蔗糖转化酶活性整体呈升高的趋势.鲜烟叶碳代谢产物中总糖、还原糖和淀粉的含量逐渐增加,达到最大值后开始下降;氮代谢产物中总氮和游离氨基酸含量逐渐降低,至M4成熟度时分别下降了17.30％和50.27％.随着成熟度的提升,烤后烟叶化学成分的协调性更好,在M3成熟度时烤后烟叶的中性致香物质含量最高,感官质量评价最佳.综合来看,在一定范围内随着成熟度的提升,烤后烟叶质量有升高的趋势,超过一定范围后,烟叶质量出现下降,豫西地区上部烟叶以M3成熟度采收时烟叶质量最佳.

目前我国部分烟区上部烟叶仍存在成熟度不够等问题,成为限制上部烟叶品质提高的重要因素.为揭示上部烟叶的成熟机理,从烟叶生理生化角度对烤烟上部烟叶成熟过程中的碳水化合物、含氮化合物、致香物质和质体色素等物质代谢规律进行了总结,同时重点阐述了上部烟叶成熟过程中生物酶活性、代谢相关基因和蛋白功能等的变化规律.在上部烟叶成熟度研究和应用现状分析基础上,对未来上部烟叶成熟机理、成熟特征判断和生长调节剂使用等热点研究方向进行了展望.

Carbon nanomaterials (CNMs) have attracted a great deal of research interest for their potential application in plants. Some types of CNMs have exhibited the ability on regulating plant growth of plants, which showed a promising future in agriculture. However, detailed mechanism of their regulation on plant growth is still not well characterized especially at the molecular level. In this study, our objective is to study the regulation mechanism of carbon nanoparticles (CNPs) on tobacco callus growth at protein level. During the incubation period, proteomic profiling of tobacco callus was investigated by using the isobaric tags for relative and absolute quantitation labelling (iTRAQ) coupled with 2D-LC MS/MS. The function of differentially expressed proteins was achieved by using a gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) analysis with InterproScan, and qRT-PCR was used to support the results of the proteomic data. The results showed that the growth of tobacco callus was enhanced by CNPs, with the optimal concentration at 50 mg/L. The expression levels of 198 cellular proteins were significantly up- or down-regulated after CNPs treatment. Some differentially expressed proteins were located in mitochondria and involved in calcium-mediated signaling pathway. The proteomic profile changes provided more deeper understanding of the highly complex regulatory mechanisms in tobacco callus exposed to CNPs.

为解析碳纳米溶胶促进烟草钾素吸收的生理机制,利用砂培试验和非损伤微测技术(Non-invasive micro-test technology,NMT),研究了碳纳米溶胶对4个烟草品种K+通道基因表达、根系K+流速、植株钾素吸收和积累的影响.结果表明:碳纳米溶胶能促进烟草K+内流通道基因的表达,促进K+内流,抑制K+外流,且不同品种、不同部位的K+通道基因对碳纳米溶胶的响应各不相同.在碳纳米溶胶处理下,RG17、鄂烟1号、云烟87和中烟100的K+外流通道基因NTORK1表达量相比对照分别降低31.1％、59.9％、25.6％和32.0％;K+内流通道基因NKT1在鄂烟1号和云烟87根系中的表达显著增加,增幅分别为210.6％和130.9％.碳纳米溶胶处理增加了各烟草品种根系的K+流速,与对照相比,RG17、鄂烟1号、云烟87和中烟100经碳纳米溶胶处理后K+内流速率分别增加38.17、39.71、13.96、8.61 pmol·cm-2·s-1.此外,经碳纳米溶胶处理后,鄂烟1号、云烟87和中烟100植株钾素积累量较对照分别增加了18.6％、15.6％和46.6％.可见,碳纳米溶胶通过调节根系K+通道基因的表达可增强根系对钾素的吸收并促进烟草体内钾素的积累,但不同烟草品种对碳纳米溶胶的响应存在明显差异.

To investigate the relationship between the characteristics of soil organic carbon components and the quality of tobacco leaves, the samples of soils and tobacco leaves were collected from 10 tobacco-planting areas in Liangshan and analyzed. The results showed that: 1) The contents of total soil organic carbon and high, medium and low active organic carbon in Liangshan tobacco-planting areas were 2.24-27.84, 0.16-1.43, 0.51-3.46 and 0.65-4.64 g·kg-1, respectively. The carbon/nitrogen ratio in soils ranged from 2.52 to 23.75 with a large variation. 2)The contents of total soil organic carbon and high, medium and low active organic carbon extremely significantly positively correlated to the contents of alkaline nitrogen, available phosphorus, available potassium and total nitrogen, which indicated that soil organic carbon was an important indicator for soil fertility. 3)When organic carbon contents in soils were higher, the proportion of tobacco samples with higher total alkaloid and total nitrogen contents increased significantly. When soil carbon/nitrogen ratio was lower, the proportion of tobacco samples with higher total alkaloid content decreased remarkably, which suggested that soil organic carbon had important effects on the contents of total alkaloid and total nitrogen in tobacco leaves.

In order to investigate any promoting effect and associated physiology by nano-carbon sol, culture experiments were carried out under solid and liquid culture conditions. The effects of nano-carbon sol application on the expression of related genes including water channel protein gene and cell division gene, biomass, nutrient accumulation and suspension cell growth characteristics of tobacco BY-2 cells were studied. The results showed that the addition of nano-carbon sol enhanced the growth of tobacco callus and increased the accumulation of N, P and K nutrients. In all the treatments, 50 mg/L concentration of nano-carbon sol had the largest increase rates of nutrient accumulation in callus, namely 41.3％, 27.6％ and 47.0％, respectively. At the same time, nano-carbon sol significantly increased the expression levels of aquaporin gene NtPIP1 and cyclin gene CycB at early culture stage, which played an important role for promoting callus growth. In suspension culture experiments, nano-carbon sol presented a good biocompatibility at the early and middle culture stages without significantly affecting the density, packed cell volume and mortality of the suspension cells. At the late culture stage, nano-carbon sol had an inhibitory effect at the concentration of 10 mg/L, which decreased the cell density and cell viability, which is likely to be associated with the aging of cultured cells. In conclusion, the effects of nano-carbon sol on the growth of tobacco cells were influenced by the culture conditions and application concentration.

为揭示青枯雷尔氏菌的遗传多样性,进而有效防治烟草青枯病,从江西省抚州市宜黄、黎川、乐安、广昌、资溪和南丰等烟叶产区采集青枯病发病烟株进行病菌分离,并以青枯雷尔氏菌种特异引物759/760进行PCR鉴定,获得了青枯雷尔氏菌33株.同时采用RAPD熔解曲线分析法(McRAPD)及内切葡聚糖酶基因(egl)测序法进行遗传多样性分析,确定其序列变种.McRAPD结果显示,多数青枯雷尔氏菌菌株遗传分化与地理来源相关性不强.egl基因测序结果表明,33株菌属于亚洲分支(演化型Ⅰ)的序列变种13、15、17和34,序列变种和地理来源具有较强相关性.序列变种15菌株占比最高,为57.6％,19株分离自广昌、黎川、乐安、南丰、宜黄和资溪等6县10个乡镇,是优势序列变种.序列变种17占比最少,为9.1％,3株全部分离自宜黄县黄陂镇.而4株序列变种34主要分离自乐安县增田镇,黎川县潭溪乡和资溪县高阜镇,占比12.1％.广昌县尖峰乡和头陂镇,南丰县傅坊乡和太源乡等产区在云烟87上分离到序列变种13,占分离菌株的21.2％,在江西省抚州市为首次发现.

In order to study the effect of micro morphological characteristics in quality evaluation,flue-cured tobacco leaves of different stalk positions were prepared and investigation was made into differences of microscopic morphology and its relationship with apparentquality and physical index based on SEM combined with image analysis.Results showed that:(1) Cell outline of flue-cured leaves was clear with central part bulged and wrinkled,and the wrinkle declined with the rising of stalk position.(2) Micro-morphological characteristics of different leaf positions showed obvious difference.Lower position leaves had large cell area and irregular cell shapes,while middle position leaves had relatively small cell area and irregular cell shape,and upper position leaves had regular cell shape.(3) Cell shape showed difference among leaves of different leaf structure and thickness,in which cell shape was relatively irregular in leaves with relatively loose structure and thin thickness.(4) The degree of irregularity of cell shape had a direct and indirect (by cell circumference) positive effect on leaf structure.The cell shape factor which characterized the degree of irregularity of cell shape had a better application prospect for evaluating leaf structure and maturity of flue-cured tobacco leaves.

To investigate the effects of soil texture on the physicochemical properties, enzyme activities and microbial communities of soil, soil samples collected from central and south Henan tobacco-planting areas, i.e. Xiangxian, Jiaxian and Biyang, were analyzed. The results showed that: 1)Main soil texture in the three counties was clay loam soil, and the proportions of clay soil and loam soil were small. The field capacity of loam soil was higher than that of clay soil, however the subsurface hardness of clay soil was higher than that of loam soil and clay loam soil. 2)pH values and contents of organic matter and available K were higher in clay soil, while available P contents in loam soil and chlorine contents in clay loam soil were higher. 3)The phosphatase activity of loam soil was higher than that of clay soil, however its invertase and catalase activities were lower than those of clay loam soil and clay soil; among the three kinds of soils, the urease activity of clay soil was higher; while the quantity and abundance index of bacteria in clay soil and the diversity index of bacteria in clay loam soil was higher than those in loam soil. 4)The bacterial community compositions in loam, clay loam and clay soils were similar on phylum level, but there was some difference in relative abundance. The relative abundances of Acidobacteria were the highest both in loam soil and clay soil, and the proportion of Proteobacteria were the highest in clay loam soil. The fungal community compositions and relative abundances were not significantly different among the three kinds of soils.

There has been great interest in potential of carbon nano-materials in regulating plant growth and development.Carbon nano-materials can promote the growth and development of plants.It can promote seed germination,root growth and biomass accumulation of some plants,which promise a wide application prospects in agriculture.However,carbon nano-materials also have negative effects on plant to some extent,such as tissue damage,growth inhibition,and it may create some environmental risks.In this paper,the types of carbon nano-materials,the effects of carbon nano-materials on the plant growth and development,the interaction mechanism and safety evaluation of plant production were reviewed.We also discussed the research progress,aiming to provide useful references for the in-depth research.

Soil column leaching experiment and soil pot experiment were conducted in this study in order to clarify the effects of nano carbon sols (NCS) on pH, EC and nutrient contents of alkaline soil. The results showed that NCS obviously reduced soil pH and EC values, the maximal reduced value were 0.49 units for pH and 0.14 mS/cm for EC, respectively, and the reducing effect was more obvious under higher NCS concentration. NCS significantly increased nitrate contents in 0-80 cm soil, averagely increased by 21.99%, 27.34% and 25.57% under different treatments, respectively, but it influenced little on ammonium nitrogen content. NCS increased available phosphorus contents in different soil layers, averagely increased by 10.10%, 13.72% and 12.30% respectively under different treatments, but the effects were different in different soil layers. NCS had higher activation effect on soil potassium, could increase soil available potassium contents significantly, averagely increased by 6.62%, 12.54% and 13.18% under different treatments. Therefore, NCS can obviously reduce pH and EC value, improve the available nutrient contents in alkaline soil and thus is effective in improving alkaline soil.

To detect the amount of Ralstonia solanacearum in tobacco planting soil before bacterial wilt occurrence and effectively control tobacco bacterial wilt, locked nucleic acid-based (LNA-based) quantitative real-time PCR assay was established and used in this study. Taking cytochrome c gene (NCBI Genbank accession number: WP_011002214.1) from Ralstonia solanacearum as the target, its primer sequence covering the target fragment was synthesized with LNA technique. Quantitative real-time PCR was used to quickly detect the amount of Ralstonia solanacearum in tobacco planting soil. The results showed that the standard curve of cytochrome c gene had a good correlation with R2> 0.99. Comparing with conventional DNA primers, LNA primer of cytochrome c gene could increase annealing temperature of PCR reaction and amplification efficiency. The DNA of 18 soil samples from tobacco fields (paddy soil) infected with bacterial wilt were tested and the amount of Ralstonia solanacearum in per gram of soil was 2.52×104-1.88×107. Therefore, LNA-based quantitative real-time PCR assay is suitable for the quantitative determination of Ralstonia solanacearum in paddy soil,and the early detection of tobacco soil will contribute to the forecast of tobacco bacterial wilt.

To investigate the contents, annual variation and regional difference of mineral nutrient elements in flue-cured tobacco and the relationships with cigarette smoke sensory quality indexes, 821 tobacco leaves samples (C3F) were collected from main tobacco planting areas in China from 2011 to 2015. The mineral nutrient elements including N, K, Ca, Mg, Cl, Mn, Fe, Zn, Cu and Ni in the samples were analyzed. The results showed that: 1) From 2011 to 2015, the N content in the samples was generally suitable; K and Cl contents presented great regional differences. Mg and Mn contents were relatively low in leaves from some areas, and Zn content was low in most samples. By planting areas, Cl and Zn contents were lower in leaves from some areas in the upper and middle reaches of the Yangtze River, and the K and Zn contents in leaves from Huanghuai and North tobacco planting areas should be increased properly. 2) From 2011 to 2015, the annual variations in contents of the 10 mineral elements were different in different years. By comparing with the published data, it was discovered that N, K and Mn contents were increased, Ca and Cu contents were decreased, and Mg, Cl and Fe contents in flue-cured tobacco did not change obviously within recent 30 years in China. 3) Ca, Ni and Cu contents negatively correlated to smoke sensory quality to a certain extent, and the order of gray correlation between 10 mineral elements and sensory quality was N>K>Ca>Mg>Fe>Zn>Ni>Mn>Cl>Cu.

To develop a method for rapid recognition of the producing area of flue-cured tobacco based on its appearance characteristics, the distribution of appearance regional characteristics of cured middle leaf samples of grade C3F sampled from different planting areas were studied,and the relationships with the appearance quality and physical characteristic of flue-cured tobacco were analyzed by quantitative assignment method. The results showed that: 1) Leaves from Huanghuai and North planting areas were much similar in appearance regional characteristics, which displayed a slightly grey background color of leaves, moderate or thicker thickness and relatively hard and brittle flexibility. Leaves from Southwest and the upper and middle reaches of the Yangtze River planting areas were similar, having a white background color, relatively bright luster and relatively smooth leaf surface tissue. The background color of leaves from southeast planting areas was slightly red, leaf luster was relatively dark and leaf surface tissue was slightly rough. 2)The background color of leaves from the secondary areas in the same planting areas was not significantly different, while the difference of waxiness and flexibility was larger. 3) The cluster analysis based on different regional scales gave identical results, and the secondary areas in the same planting area were clustered in the same category. 4) The correlation of appearance regional characteristic with appearance quality (except for the index of color difference between front and back) was weaker, while that with leaf thickness and weight of leaf per area (physical properties) was relatively strong. Therefore, the appearance regional characteristics of tobacco leaf could be used to recognize its producing area.

为满足“真龙”品牌香烟原料风格特色的需求,以奉节基地主要收购点烟叶为材料,研究烟叶化学成分指标与外观和评吸质量的关系.结果表明:奉节基地烟叶烟碱和蛋白质等含氮相关指标含量较高(尤其是上部叶),还原糖、总糖和淀粉等碳水化合物含量略低;总氮和蛋白质含量与烟叶感官质量呈显著负相关,还原糖和总糖含量与烟叶感官质量呈显著正相关;总糖、还原糖对烟叶的柔和性和甜度等评吸指标与成熟度和色度等外观指标呈显著正相关,蛋白质和两糖比对杂气、余味等评吸指标与上部烟叶外观指标呈显著负相关.生产中应加强碳氮代谢和烘烤方面的调控,适当提高烟叶总糖含量,降低蛋白质含量.

In order to clarify the adsorption characteristics of nano-carbon on heavy metals, the adsorption characteristics of nano-carbon on Cd and Cu were investigated in a liquid-liquid system. The effects of nano-carbon on the available Cd content in soil and the Cd absorption of tobacco cultured in soil were discussed. The results showed that the Cd and Cu isothermal adsorption curves of nano-carbon could be fitted by Freundlich and Langmuir equations. The Cd and Cu adsorption by nano-carbon agreed with a quasi-two stage kinetic equation, and its adsorption rate was fast. At the initial solution concentration of 2 000 mg/L, the Cd adsorption amount was more than 80% of the equilibrium adsorption amount after 10 min, while it only took 3 min to reach that level for Cu adsorption. With the rise of temperature, the adsorption amounts decreased, which indicated that the adsorption by nano-carbon was an exothermic reaction. With the increase of pH, the adsorption amount by nano-carbon raised. Under the condition of this experiment, both the available Cd content in soil and the Cd content in tobacco leaves decreased with the increase amount of nano-carbon, and the optimal nano-carbon addition rate was 4.0 gram/pot.

[目的]明确石墨纳米溶胶对钾的吸附特性及其对烟草钾吸收和钾离子流速的影响.[方法]采用等温吸附试验和水培试验,分析石墨纳米溶胶对钾的吸附特性及对烟草钾吸收的影响.[结果]l)石墨纳米溶胶对钾的等温吸附线用Langmuir方程能更好的拟合;石墨纳米溶胶对钾的吸附可分为快慢两个阶段,反应符合准二级动力学方程.随着温度升高,石墨纳米溶胶对钾的吸附量降低.在pH为7.0时,石墨纳米溶胶对钾的吸附量达到最大.2)适宜浓度石墨纳米溶胶能促进烟草对钾的吸收积累,当浓度为40 mg/L时,地上部钾含量达到最高;当浓度为5 mg/L时,地上部钾素积累量达到最高.3)随着浓度升高,石墨纳米溶胶在烟草根系表面的富集增强.在10 mg/L浓度下,烟草根系的钾离子流速由外流变为内流.[结论]石墨纳米溶胶有较强的吸附特性,能吸附在根系表面,影响根系钾离子吸收.

To investigate the differences of physicochemical characteristics among flue-cured tobacco of difference genotypes and their influences on sensory quality, the main physicochemical characteristics and sensory quality of flue-cured tobacco leaves of several varieties were analyzed via field and smoking experiments. The results showed that: 1) K/Cl ratio, reducing sugar/total sugar ratio, single leaf weight and stem content differed among the varieties. The K/Cl ratio and stem content of cv. Yunyan 100 were relatively higher;the reducing sugar/total sugar ratio of cv. Honghuadajinyuan was relatively higher, while its single leaf weight was relatively lower. 2) The differences in leaf appearance quality among the varieties were closely correlated to the physical characteristics(especially weight per unit leaf area)of tobacco, and the different sensory quality was correlated more closely to the chemical characteristics. 3) Based on six physicochemical indexes, including K/Cl ratio, nitrogen/nicotine ratio, nicotine, reducing sugar/total sugar ratio, starch and single leaf weight, a prediction model (with a determination coefficient of 0.93) for comprehensive smoke sensory quality of tobacco was established, the model could be applied to evaluate flue-cured tobacco varieties and tobacco leaf quality after further experimental validation.

To promote rational fertilization and improve quality of flue-cured tobacco leaves,the soil samples and corresponding flue-cured tobacco leaves were collected to investigate the soil nutrient status and its relationship with chemical components of flue-cured tobacco leaves in Fengjie production area.Results:The soil pH was appropriate for tobacco planting in mail area of Fengjie,and the main features of soil nutrients were high content of organic matter,available N,available P and medium trace element,and low content of available K.The main features of chemical components in tobacco leaves were as follows,nicotine content was relatively low,and protein content was relatively high,and the ratio of reducing sugar to total sugar was low.There was a relationship between available N,organic matter,Mn,Zn content in soil and chemical components in leaf tobacco,especially the relationship between available N in soil and protein content in leaf tobacco.A fertilizing scheme of controlling N,maintaining P and increasing K was recommended in Fengjie production area.