In order to support the optimal cultivation technology of the New Flue-cured Tobacco Variety Yunyan 121 and promote the popularization and application of the variety, Weining County, a representative tobacco area in western Guizhou, which is suitable for the cultivation of Yunyan 121, was selected. Three factors of nitrogen application rate, density and number of leaves were set. The effects of different nitrogen application rate, density and number of leaves on the yield and quality of Yunyan 121 were analyzed by using quadratic saturated D-optimal design, index sum method and regression analysis, The optimum combination of nitrogen application rate, density and leaf number of Yunyan 121 was found out. The results showed that the yield and quality of Yunyan 121 were significantly affected by different nitrogen application rate, density and leaf number. When the nitrogen application rate was 98.80 kg/hm~2, the density was 15 247 plants/hm~2 and the leaf number was 16 pieces/plant, Yunyan 121 had the highest yield per mu（1 941.53 kg/hm~2）; When the nitrogen application rate was109.9 kg/hm~2, the density was 14 993 plants/hm~2 and the number of leaves left was 16 pieces/plant, Yunyan 121 had the highest output value per mu（53 286.36 yuan/hm~2）; When the nitrogen application rate was 85.72 kg/hm~2, the density was 17 571 plants/hm~2 and the number of leaves left was 19 pieces/plant, Yunyan 121 had the highest comprehensive quality value（92.1）. According to the yield, output value and comprehensive quality value of tobacco leaves, Yunyan 121 is planted in the Western tobacco area of Guizhou. When the nitrogen application rate is 98.2 kg/hm~2, the density is 15 937 plants/hm~2 and the number of leaves left is 17 pieces/plant, the yield of tobacco leaves can be1 922.64 kg/hm~2, the output value is 52 258.92 yuan/hm~2 and the comprehensive quality value is 89.78, which can give consideration to the yield and quality of tobacco leaves.

叶片数是烟草产量的重要构成因素,对提高烟农种烟收入和保证卷烟原料的供应有重要意义.低温易引起烤烟早花,导致叶片数减少,而韭菜坪2号可在高海拔地区种植,且叶片数不受影响.为揭示韭菜坪2号烤烟叶片数的遗传规律,本研究以叶片数较少的优质烤烟NC82和叶片数较多的烤烟品种韭菜坪2号为亲本,构建六世代(P1、P2、F1、F2、BC1和BC2)遗传群体,采用"主基因+多基因"混合遗传模型对韭菜坪2号的叶片数进行遗传分析.结果表明,最优模型为2MG-ADI,即受2对加性-显性-上位性主基因控制,2对主基因中加性效应起主导作用,且存在互效作用;主基因在F2、BC1、BC2世代的遗传率分别为62.72％、6.87％、45.51％,遗传率不高,可能受环境或其他因素的影响较大.本研究结果为韭菜坪2号叶片数基因的遗传和利用提供参考依据.

为探究NtMYB4a是否与其他蛋白质发生互作,并共同影响烟草次生代谢产物的合成和响应非生物胁迫,通过构建NtMYB4a基因酵母双杂交诱饵载体,从烟草cDNA文库中筛选出与NtMYB4a互作的候选蛋白,并利用实时荧光定量PCR初步验证低温胁迫下候选蛋白和NtMYB4a的共表达关系.结果表明,从cDNA文库中初步筛选出39个与NtMYB4a互作的蛋白,经回交验证排除假阳性,最终得到6个与NtMYB4a具有互作关系的蛋白,分别是RALF?like 22蛋白、锌指A20/AN1结构域蛋白、液泡分选蛋白、天冬氨酸蛋白酶、B2蛋白和应激增强蛋白.在低温胁迫下,6种互作蛋白的基因表达量变化趋势与NtMYB4a相似,均呈升高—降低—升高的趋势.相关性分析发现,液泡分选蛋白、B2蛋白、RALF?like 22蛋白和应激增强蛋白基因的表达与NtMYB4a的表达呈正相关关系,锌指A20/AN1结构域蛋白和天冬氨酸蛋白酶基因的表达与NtMYB4a的表达呈较弱的负相关关系,推测NtMYB4a可能与这些蛋白互作参与烟草低温胁迫的防御调控.本研究结果为进一步探究NtMYB4a参与响应非生物胁迫的分子调控机制提供了依据.

为改善烟苗生长环境,提高烟苗素质,对现有常规160孔育苗盘(CK)进行优化改进,设计出2种(T1:160孔凸底和T2:160孔平底)新型通气孔育苗盘开展育苗试验,并分析了通气孔育苗盘对育苗关键期烟苗生长发育和生理生化指标的影响.结果表明:(1)改进的2种通气孔育苗盘对烟苗生育期无影响;(2)通气孔育苗盘明显促进了后期烟苗的根系生长和生物量积累,在成苗期(6叶1心)T1和T2处理所育烟苗地下部干重、鲜重、根体积和表面积均显著高于CK,根干鲜重提高了42.90％～84.62％,根体积提高了35.29％～70.59％,根表面积提高了53.52％～80.89％;(3)通气孔育苗盘T1和T2处理所育烟苗在成苗期(6叶1心)的POD活性较CK分别提高了104.80％、202.75％,CAT活性分别提高了513.43％、707.20％,淀粉酶活性分别提高了45.43％、107.67％;(4)T1处理所育烟苗地上部矮小,T2处理烟株生物量积累更多,根系更发达,基质真菌数量更少,有利于烟苗的真菌病害防控.总之,通气孔育苗盘明显促进了成苗期(6叶1心)烟株根系生长和根系生物量积累,以160孔平底通气孔育苗盘培育烟苗素质最好.

钾含量是烟叶重要的品质指标之一,研究烟叶钾含量杂种优势形成的原因,对高钾含量杂交种的选育具有重要意义.本研究选用钾含量差异较大的12个烟草品种(系)为亲本,按照NCⅡ遗传交配设计组配出35个杂交组合,分析了烟叶钾含量杂种优势的表现,并探讨了烟叶钾含量杂种优势相关基因表达差异.结果表明:烟叶钾含量主要受非加性效应基因的影响,具有明显的杂种优势,中亲优势变幅为-38.06％～29.74％.AKT1、NtKC1和NtTPK1基因在烟叶中的中亲相对表达量与烟叶钾含量杂种优势显著相关,相关系数分别为0.86、0.84和0.86;AKT1和KAT1基因在根系中的中亲相对表达量与烟叶钾含量杂种优势的相关性达到了显著水平,相关系数分别为0.80和0.77.与亲本相比,杂种优势相关基因AKT1和NtKC1在烟叶和根系中均呈正向超显性表达模式,tTPK1在烟叶和根系中均呈正向显性表达模式,KAT1基因在烟叶中呈正向显性表达模式,在根系中呈正向超显性表达模式.说明内流型钾离子通道基因AKT1、NtKC1、KAT1和钾离子转运蛋白基因NtTPK1在杂交种根系和叶片中的显性和超显性表达,在杂交种烟叶钾含量优势的形成中起了关键作用.

杂种优势利用是提高作物产量和品质的一种重要育种方法,明确生态条件和打顶措施对杂种优势的影响,对提高烟草杂种优势研究利用效率有重要意义.本试验以烟叶烟碱含量杂种优势相差较大的3个杂交组合及其亲本为材料,选择生态条件差异较大的3个生态试验点,研究了不同生态环境和打顶措施对烟叶烟碱含量、烟叶烟碱含量杂种优势及其相关基因相对表达量的影响.结果表明,不同生态环境间的烟叶烟碱含量差异显著,烟株打顶前后烟叶烟碱含量发生了显著的变化,打顶处理的烟叶烟碱含量显著高于不打顶处理;不同生态环境、打顶和不打顶处理间的烟叶烟碱含量的中亲优势值差异不显著,而超亲优势值、超标优势值差异显著;不同生态环境、打顶和不打顶处理间,烟叶烟碱含量杂种优势相关的PM T、A O、QS基因的中亲优势表达量相差也不显著,与处理间中亲优势值的表现差异一致.结果表明,在选用中亲优势作为度量指标进行杂种优势的遗传研究中,可不考虑生态环境和打顶的影响,在一个试验点对双亲及F1采用相同的农艺措施就能获得较为准确的鉴定结果;而在选用超亲优势、超标优势作为度量指标进行杂交种应用潜力评价中,需在不同生态环境、不同农艺措施下进行鉴定.

为了解NtMYB4a基因的表达是否与烟草多酚化合物的合成有关,本研究以NtMYB4a过表达和CRISPR/Cas9敲除植株及其野生型植株为试验材料,采用实时荧光定量PCR技术分析NtMYB4a和多酚物质合成相关酶基因PAL、C4H、4CL、HCT的表达,使用HPLC方法测定烟叶中绿原酸、芦丁等8种多酚物质的含量,并进行相关性分析.结果 表明,过表达NtMY B4a植株中,除花中的C4H外,茎、叶、花中PAL、C4H、4CL和HCT的表达量均极显著高于野生型,烟株茎、叶、花中绿原酸含量也显著增加;敲除NtMYB4a使烟株花、茎和叶中PAL、C4H、4CL和HCT的表达均下调,其中茎中PAL、HCT的表达极显著下调,使叶和花中的绿原酸、芦丁、槲皮素、莨菪亭含量均显著降低,茎中芦丁含量则显著增加,而山奈酚、芥子酸、对香豆酸和咖啡酸含量无显著性变化;NtMYB4a的表达量与绿原酸含量呈极显著正相关关系,与芦丁、莨菪亭、山奈酚、芥子酸、槲皮素、对香豆酸、咖啡酸含量无显著相关性.因此,NtMYB4基因对烟草绿原酸合成相关酶基因表达及绿原酸合成具有调控作用.本研究结果为进一步阐明烟叶中绿原酸形成的分子机制提供理论依据.

为评价不同烤烟品种的生态适应性和筛选适宜贵州毕节烟区种植的烤烟品种,以云烟87为对照,对贵烟202、湘烟5号、云烟116、CC297和CC27等5个品种的农艺性状、田间自然发病率、经济性状和感官质量进行分析,结合一年两点区域试验分析和综合指数法对各参试品种进行综合性状评价.结果表明,云烟116在两个试验点表现均优于对照,适应环境能力强,属广适性品种;CC27在黔西试验点表现优于对照,但是在七星关试验点表现较差,贵烟202和湘烟5号在黔西试验点表现良好,在七星关试验点表现与对照相当;CC297的均价、上等烟率、一年两点区域试验分析和综合指数评价均差于对照.云烟116的农艺性状、田间自然发病率、经济性状和感官质量等综合表现优于对照,适宜在毕节烟区种植.

MYB转录因子是植物体内最大的一类转录因子,在植物应对非生物胁迫中发挥重要作用.为探讨烟草NtMYB4a基因在响应干旱、低温和茉莉酸甲酯胁迫下的功能,以NtMYB4a基因的过表达和敲除突变体及野生型烟株为材料,采用RT-qPCR技术分析了干旱、低温和茉莉酸甲酯处理0、1、3、6、12、24和48 h后烟草叶片中Nt-MYB4a的表达变化以及脯氨酸和丙二醛含量差异.结果显示:过表达突变体中NtMYB4a的表达量极显著高于野生型烟株,且不易受干旱、低温和茉莉酸甲酯胁迫的影响,烟叶中的脯氨酸含量增高,丙二醛含量降低;而基因敲除突变体中NtMYB4a的表达量显著低于野生型烟株,且易受干旱、低温和茉莉酸甲酯胁迫的影响,烟叶中的脯氨酸含量降低,丙二醛含量增加.此外,利用Plant CARE软件分析了NtMYB4a启动子区域,结果显示该区域含有赤霉素、生长素、脱落酸、茉莉酸甲酯、光、干旱、低温等激素和非生物胁迫相关的顺式作用元件.这些结果表明,Nt-MYB4a基因的表达受干旱、低温胁迫和茉莉酸甲酯的诱导,可能与烟株的抗逆性相关.

The application effect of the floating raising tobacco seedling technique from 1997 to 2011 was summarized to popularize the floating raising tobacco seedling technique in tobacco production in Guizhou.The achievements,patents,system,management mode,technological standard and management standard of the floating raising tobacco seedling technique were reviewed.The authors proposed the countermeasures to further promote application of the floating raising tobacco seedling technique in tobacco production based on analyzing the problems in Guizhou.

Artificial neural network is a powerful tool for modelling and prediction in various fields. Tobacco management specialists need simple and accurate techniques to evaluate tobacco growth and cultivation in the growth period. The objective in this study is to develop a back-propagation neural network (NN) model to accurately predict tobacco growth under various climatic and soil conditions in China. The proposed model used the field test data at 11 locations in main tobacco areas in China. As the output of the NN, the dry weight of tobacco plants was significantly influenced by dry weigh preceding stage, time for sampling, average temperature, relative humidity, soil organic matter and available P value, so these 6 environment factors were taken as the NN input. In the study, an optimal NN model was developed with 6 inputs, 1 output and 19 neurons in the hidden layer. With the same test data, the proposed NN model could perform better than the conventional regression model in terms of the relative error ranges and relative coefficient.

Yunyan 85,a flue-cured tobacco variety,was planted under 1482 plants/667m2 density and applying organic manure in 5 typical ecological areas to study flue-cured tobacco quality and aroma components of different ecological areas for excavating the ecological advantage of flue-cured tobacco in Guizhou.The results showed that the flue-cured tobacco yield,single leaf weight,unit leaf area weight of Yunyan 85 planted under 1482 plants/667m2 density and applying organic manure in 5 typical ecological areas were all lower than those planted under the conventional cultivation pattern,the nicotine content of flue-cured tobacco applied with organic manure was relatively lower,its sugar content was relatively high and other quality indexes of flue-cured tobacco slightly decreased compared with those under the conventional cultivation pattern.In general,the flue-cured tobacco leaf has the characteristics of good harmoniousness,high sugar/nicotine ratio,elegant gas,intensified pleasure,abundant aroma and obvious sweet.There is obvious difference in aroma components among 5 different ecological areas,which reflects regional characteristics.

In order to understand the effects of continuous mono-cropping on the forms of soil nitrogen and plant nutrition and its relative physiological index,we analyzed soil nitrogen,various soil microorganisms and the nature of nitrogen accumulation in the tobacco leaf in a potting experiment using different soil types.It was demonstrated that due to the different soil types,the total N,alkali-hydrolysable N,nitrate N and ammonium N increased with continuous mono-cropping whereas the nitrifying bacteria,ammonium oxygenation bacteria and nitrification decreased.Continuous mono-cropping led to decreased chlorophyll and nitrate reductase levels and increased proline content of the leaf.At the same time,mono-cropping also led to a gradual increase in total N,nicotine and protein in the leaf and their decline in the stem and the root.A quaternary yellow soil which is not suitable for mono-cropping of tobacco had lower amounts of nitrate,nitrifying bacteria,ammonium oxygenation bacteria and nitrification than that of yellow lime soil.Improved crop performance under continuous mono-cropping could be achieved by reducing the soil N content to induce higher levels of beneficial microorganisms.

[Objective] The study aimed to research the occurrence mechanism of soil obstacle factors in continuous cropping tobacco land.[Method] The microbiological community and physico-chemical properties in tobacco-growing soils under different continuous cropping years were studied by variance analysis method.[Result] The significant differences on bacteria quantity were found in tobacco-growing land between continuous cropping years of n10 and n≥10,that on fungus quantity were found between continuous cropping years of 5n10 and n≥10 and no significant differences on actinomycetes quantity were found.The significant differences on number of Ralstonia solanacearum pathogens were found in tobacco-growing land between continuous cropping years of n10 and n≥10 and no significant differences on number of Phytophthora parasitica var nicotiana pathogens.Between continuous cropping years of n10 and n≥10,the contents of total N,total P and organic matters and the value of pH in soil had extremely significant differences,no statistical differences were found on the content of total K,the contents of available N and P in soil had not significant differences and the contents of available K exhibited significant differences.[Conclusion] Along with the increase of continuous cropping years,the bacteria quantity was decreased,fungus quantity was increased,actinomycetes was slightly increased,R.solanacearum pathogen quantity was decreased,and the contents of total N,total P,and organic matters were increased gradually.

The different formula combination of tobacco leavies from Xingyi,Daozhen,Weining and Fuquan in Guizhou and Yunnan,Henan province were tested to evaluate the smoking index by using a complete-diallel crosses model in quantitative genetics.The results showed that the general combining ability(GCA) of five smoking indexes including aroma quality,aroma quantity,flavor,offensive odor,irritation and total scores was at 1% significant level and the special combining ability(SCA) of aroma quality,irritation and total scores was at 5% significant level.The general combining ability of tobacco leavies from yunnan,Xingyi and Weining was higher than that from other producing areas.The aroma quality of Fuquan Henan was better than others.The irritation of Yunnan Henan was the lowest in all tested combinations.The total score of Xingyi Huquan was the highest in all tested combinations.The tobacco smoking characteristic of the single producing area did not accord with the effect in formula combinations of tobacco production.Therefore,the model of combining ability in quantitative genetics should be used in tobacco compatibility tests in tobacco production.

Five nitrogen concentrations were used in culturing tobacco seedlings in discs to investigate the effect on the growth and development.The results show that different nitrogen concentrations have the different effects on growing process,matter accumulation and physiological traits of tobacco seedlings.The growth period of tobacco seedlings is early 1～5 d and the aerial growth and dry matter accumulation is promoted with the increase of nitrogen concentration within 50～250 mg/L of nitrogen concentration.150 mg/L of nitrogen concentration is beneficial to tobacco seedling growth and culturing good quality seedlings with the developed root system and high root activity.

从土壤微生物活动、土壤酶活性、土壤养分的协调和平衡几个方面阐述了作物连作现象的土壤障碍因子。同时分析了连作土壤根际分泌物对作物生长发育的障碍作用,以及对作物产量和品质造成的不利影响。

Uniform experiments were carried out in 11 counties,scattered in 7 provinces,major tobacco growing regions in South and North China,to investigate contents of Pb and Cd in tobacco plants in relation to environment. Results showed that the content of Pb was higher than that of Cd in roots,stems,leaves and flowers of the tobacco plants. The content of Pb in tobacco plants varied in the sequence: roots leaves stems ≈ shoots (flowers),while Cd followed the sequence of leaves stems ≈ shoots (flowers) roots. The contents of Pb and Cd in leaves showed a trend of lower leaves middle leaves upper leaves and decreased gradually with the growth of the plants,but their accumulations increased along an S curve and their accumulation rates appeared in a singlet curve. Moreover,the contents of Pb and Cd were higher in tobacco leaves produced in South China than in North China. The variation coefficients of Pb and Cd concentrations in tobacco leaves were high,indicating significant influence of the environment. In regards to Pb (Cd) in relation to meteorological factors,negative correlations were observed with sunshine hours and positive ones with rainfall (or relative humidity). Thus,it seems quite reasonable to suggest that the higher Pb and Cd contents in tobacco leaves produced in South China than in North China could be caused by less sunshine hours,rich precipitation and high relative humidity. Furthermore,the contents of Pb (Cd) in tobacco leaves were found positively related to soil factors,such as exchangeable Pb (Cd) and available phosphorus,and negatively to soil pH. Therefore,it is possible to decrease effectively Pb and Cd contents in tobacco leaves in cultivation by growing them in low Pb (Cd) soils,liming properly,avoiding application of high rates of organic matter and heavy-metals-containing phosphorus fertilizers.

Four substrates including loam without disease, grass dung, perlite andFire ashes were researched to know these seedling substrates how to control growth of flue-cured tobacco seedling. It was resulted that seedling rate is higher with the growth of loam without disease and fire ashes. Grass dung benefits the growth of seedling, and accumulation of the dry matter of the tobacco seedling. Fitting perlite can also be use to the growth of seedling due to the enlarged ventilation. The formula of loam without disease: grass dung: perlite= 5: 3: 2 is a better one. Using this formula, the better tobacco seedling diathesis consist of higher seedling rate and growth rate, upgrowth roots can be obtained.

根黑腐病Thielariopsis basicola(Berk.et br.)Fer.是世界性的烟草病害,苗期至成株期均可受害[1].通过2001～2005年的调查,该病在贵州省赫章县地膜烤烟种植区已成为主要病害,并有加重发生的趋势,局部烟区由于该病的发生,导致烤烟绝收.地膜烟根黑腐病的发生特点及防治方法报道极少,为此,笔者在赫章县烤烟大田生产中对其发生及防治方法进行了初步研究.现将结果报告如下: