异源表达番茄LeMPK3基因提高烟草的抗低温胁迫能力

促分裂原活化蛋白激酶(MAPK)信号系统在植物应对环境胁迫中发挥重要作用。为了探讨LeMPK3基因在低温胁迫中的功能,我们从番茄中分离了LeMPK3基因并将其转化到烟草中,获得了异源表达LeMPK3基因的转基因烟草。RT-PCR分析结果表明,番茄幼苗在低温和高温胁迫及外源物质——过氧化氢(H2O2)、茉莉酸甲酯(MeJA)和水杨酸(SA)处理下LeMPK3基因的表达量提高。在烟草中异源表达LeMPK3基因可以提高超氧化物歧化酶(SOD)、过氧化物酶(POD)、过氧化氢酶(CAT)和抗坏血酸过氧化物酶(APX)活性,增加脯氨酸和可溶性糖的含量,增强逆境相关基因的表达。说明异源表达番茄LeMPK3基因提高烟草对低温的抗性。     

Induction of UDP-Glucose:Salicylic Acid Glucosyltransferase Activity in Tobacco Mosaic Virus-Inoculated Tobacco (Nicotiana tabacum) Leaves

Salicylic acid (SA) is a putative signal that activates plant resistance to pathogens. SA levels increase systemically following the hypersensitive response produced by tobacco mosaic virus (TMV) inoculation of tobacco (Nicotiana tabacum L. cv Xanthi-nc) leaves. The SA increase in the inoculated leaf coincided with the appearance of a [beta]-glucosidase-hydrolyzable SA conjugate identified as [beta]-O-D-glucosylsalicylic acid (GSA). SA and GSA accumulation in the TMV-inoculated leaf paralleled the increase in the activity of a UDP-glucose:salicylic acid 3-O-glucosyltransferase (EC 2.4.1.35) ([beta]-GTase) capable of converting SA to GSA. Healthy tissues had constitutive [beta]-GTase activity of 0.076 milliunits g-1 fresh weight. This activity started to increase 48 h after TMV inoculation, reaching its maximum (6.7-fold induction over the basal levels) 72 h after TMV inoculation. No significant GSA or elevated [beta]-Gtase activity could be detected in the healthy leaf immediately above the TMV-inoculated leaf. The effect of TMV inoculation on the [beta]-GTase and GSA accumulation could be duplicated by infiltrating tobacco leaf discs with SA at the levels naturally produced in TMV-inoculated leaves (2.7-27.0 [mu]g g-1 fresh weight). Pretreatment of leaf discs with the protein synthesis inhibitor cycloheximide inhibited the induction of [beta]-GTase by SA and prevented the formation of GSA. Of 12 analogs of SA tested, only 2,6-dihydroxybenzoic acid induced [beta]-GTase activity.     

Ethylene Production by Tobacco (Nicotiana tabacum) Callus

Tobacco callus cultures grown on defined agar-solidified media produced ethylene in differing amounts, which were related to cultural treatment and age of the callus. There was a close correlation between the rate of ethylene production and growth. In darkness, maximal rates occurred in the third week of growth with ethylene production in the range of 750 nl (callus piece)^?1 d^?1 (fr. wt. = 1.5 g), and in the light, maximal rates occurred in the first week of growth, 200 nl (callus piece)^?1 d^?1 (fr. wt. = 200 mg). Growth was also correlated with ethylene production when the latter was altered by exposure of the callus to inhibitors of ethylene synthesis, L-canaline, benzyl isothiocyanate, and 3,5-diiodo-4-hydroxy-benzoic acid. No correlation was found following treatment with AgNO₃, a presumptive inhibitor of ethylene action. The inhibition of growth and ethylene production by L-canaline was partially reversed by gassing the cultures with ethylene (1 μl/1). A mercuric perchlorate sink had no significant effect on growth. A possible relationship between ethylene evolution and growth is discussed.     

烟草的历史

1　吸烟习俗的起源和传播当今遍布于全世界各地的烟草原产于美洲 ,最早栽培并吸食的是当地印第安人。美洲印第安人从什么时候开始吸烟 ,迄今还不很清楚。目前已发现最早的资料是在墨西哥恰帕斯州的帕伦克 ,有在公元 4 32年修建的玛雅文化的古典神庙中的一块浮雕 ,可以看到古代玛雅人在宗教的仪式中有人用管状烟斗来吸烟 ,而且还在喷出烟雾。在美国阿利桑那州的帕布罗城发现公元 6 5 0年前后印第安人居住的洞穴遗址 ,其中有烟叶实物和烟斗 ,烟斗中还存有烟灰。在墨西哥马德雷山上海拔 12 0 0ｍ处的一个山洞中 ,发现一个塞有烟草的空心草管 ,…     

土壤pH值对烤烟叶片生理生化特性的影响

烟草生长初期,超氧化物歧化酶(SOD)活性及丙二醛(MDA)含量与pH增长呈正相关,而过氧化氢酶(CAT)和过氧化物酶(POD)活性则随着pH的增加而下降,pH值6.5和7.5时叶绿素含量和净光合速率(Pn)较高,旺长期达到高峰;生长后期,各处理的MDA含量和POD活性最高,SOD、CAT活性最低.各pH值处理的叶片中,叶绿素含量、Pn下降,比叶重达到高峰,但pH 8.5下的烟叶中叶绿素含量高,Pn大,比叶重较小.pH 6.5和7.5下的烟叶中蛋白质和可溶性糖总体含量高于其它pH的.烟碱含量在pH 5.5时最高,pH 8.5时最低.     

Glyphosate Tolerance in Tobacco (Nicotiana tabacum L.)

A glyphosate-tolerant tobacco cell line, Nicotiana tabacum L. Indiana (I7), was selected from the glyphosate-sensitive Wisconsin 38 (W38) line through a single step exposure to the herbicide. Tolerance and growth characteristics of I7 cells were the same for cells maintained for more than 1 year in the presence or absence of glyphosate. Glyphosate tolerance levels were constant through the growth cycle. Tolerance is not due to reduced uptake of glyphosate. Shikimate levels in I7 and W38 cells maintained in glyphosate-free medium were similar, whereas W38 cells accumulated 46 times more shikimate than I7 cells, when cells of both lines were exposed to the herbicide. Glyphosate treatment caused increased levels of aromatic amino acids in W38 cells and slightly lower levels in I7 cells. Specific activities of dehydroquinate synthase, shikimate dehydrogenase, and shikimate kinase were similar in the two cell types, whereas DAHP synthase and EPSP synthase specific activities were elevated in I7 cells. Plants regenerated from I7 cells retained tolerance to glyphosate.     

烟草ESTs资源的SSR信息分析

烟草ESTs数量迅速增加为开发新的SSR标记提供了宝贵的资源.经过软件分析,对242 683条烟草ESTs序列剔除冗余序列,在211 728条非冗余烟草ESTs序列中,共检索出9 339个SSR,SSR之间的距离约为14.21 kb,检出率为4.41%,包括216种重复基元.其中三核苷酸重复类型的SSR占主导地位,占总SSR的50.34%,其次为二核苷酸和单核苷酸,分别为23.00%,16.48%,其余重复类型所占比例均不足5%.在所有重复基元中,A/T重复为主要类型,占所有重复14.68%,其次为AT/TA、AG/TC、AAG/TTC,分别为10.49%、9.48%、6.85%.随机设计10对EST-SSR引物,对6个品种烟草进行扩增,10对EST-SSR引物均能扩增出产物,其中1对引物在6个品种有多态性.本研究为烟草EST-SSR标记的建立和进一步应用奠定了基础.     

纳豆激酶基因的优化设计、合成及其在烟草叶片中的瞬时表达

根据植物偏爱密码子优化设计、合成纳豆激酶基因sNK,利用重叠延伸PCR法在其中插入番茄果实特异性表达基因E8的第一内含子构成sNKi基因,通过农杆菌渗透法将这两种基因渗入到烟草NC89叶片中并实现瞬时表达。通过RT-qPCR法将两种基因在烟草叶片中转录水平的表达量进行比较,结果表明两种基因在烟草叶片中均表达,且sNKi基因的表达量显著高于sNK基因;通过纤维蛋白平板法在两种基因的瞬时表达样品中均能检测到纤溶酶活性,表明目的基因在烟草叶片中可正常翻译并表现出溶栓活性,且sNKi基因在翻译水平的表达量显著高于sNK基因。表明内含子对人工合成的纳豆激酶基因的瞬时表达具有显著的促进作用。     

干旱和低温胁迫影响烟草幼苗海藻糖代谢的差异比较

为了明确海藻糖代谢在干旱和低温胁迫下的响应及其差异,以及海藻糖在烟草耐旱和耐冷性中的作用,选用烟草品种云烟203幼苗为实验材料,通过对烟草幼苗进行干旱和4℃低温胁迫处理,研究叶片中海藻糖含量及代谢相关酶(海藻糖-6-磷酸合成酶TPS、海藻糖-6-磷酸磷酸酶TPP、海藻糖酶THase)活性及基因表达变化情况。结果表明,在干旱和低温胁迫下,海藻糖含量表现出先升高后降低趋势,均在处理2 d时海藻糖积累达到最大,且干旱胁迫下海藻糖含量高于低温胁迫。TPS、TPP活性在干旱和低温胁迫下先升高后降低,且干旱高于低温;THase活性在干旱和低温胁迫下均不断升高,低温下的THase活性高于干旱。TPS、TPP、THase基因在干旱下的表达量均高于0 d对照,低温下低于0 d对照。以上结果表明,干旱和低温胁迫均能促使烟草体内海藻糖的积累,干旱更能诱导相关基因的表达、相关酶活性及海藻糖含量的升高,表明海藻糖对干旱响应更敏感。     

低温胁迫下褪黑激素对烟草悬浮细胞精氨酸脱羧酶活性的影响

研究了褪黑激素对烟草(Nicotiana tabacum)悬浮细胞在低温胁迫下精氨酸脱羧酶活性及细胞生存率的影响。发现褪黑激素可以明显提高低温胁迫下烟草悬浮细胞精氨酸脱羧酶的活性, 并明显提高细胞的生存率。表明褪黑激素可能在低温条件下通过调节植物细胞内多胺的合成而提高抵御冷害的能力。     

低温胁迫下褪黑激素对烟草悬浮细胞精氨酸脱羧酶活性的影响

研究了褪黑激素对烟草(Nicotiana tabacum)悬浮细胞在低温胁迫下精氨酸脱羧酶活性及细胞生存率的影响.发现褪黑激素可以明显提高低温胁迫下烟草悬浮细胞精氨酸脱羧酶的活性,并明显提高细胞的生存率.表明褪黑激素可能在低温条件下通过调节植物细胞内多胺的合成而提高抵御冷害的能力.     

不同光质对烟草叶片生长及光合作用的影响

通过对烟草植株覆盖白、红、黄、蓝、紫色滤膜获得不同光质,研究了光质对烟叶生长及光合作用的影响。结果表明,与白膜处理相比,红膜与蓝膜处理下的烟草叶片较厚,比叶面积较小,叶绿素a/b比值、净光合速率、可变荧光强度（Fv）和最大荧光强度（Fm）的比值Fv/Fm（PSⅡ最大光化学量子效率）、PSⅡ实际光化学量子效率（ΦPSⅡ）、光饱和点和CO2饱和点均较高。黄膜处理下的叶片较白膜处理的更薄,净光合速率、Fv/Fm、ΦPSⅡ、光饱和点、CO2饱和点均较低。紫膜处理的叶片比叶面积比白膜处理的小,净光合速率和Fv/Fm比白膜的大。实验结果表明红光、蓝光和紫光促进了烟叶的生长,这种促进作用是与其高光合效率紧密相连的;而黄光对烟叶的生长有一定程度的抑制作用。     

Analysis of Methylated Patterns and Quality-Related Genes in Tobacco (Nicotiana tabacum) Cultivars

Methylation-sensitive amplified polymorphism was used in this study to investigate epigenetic information of four tobacco cultivars: Yunyan 85, NC89, K326, and Yunyan 87. The DNA fragments with methylated information were cloned by reamplified PCR and sequenced. The results of Blast alignments showed that the genes with methylation information included chitinase, nitrate reductase, chloroplast DNA, mitochondrial DNA, ornithine decarboxylase, ribulose carboxylase, and promoter sequences. Homologous comparison in three cloned gene sequences (nitrate reductase, ornithine decarboxylase, and ribulose decarboxylase) indicated that geographic factors had significant influence on the whole genome methylation. Introns also contained different information in different tobacco cultivars. These findings suggest that synthetic mechanisms for tobacco aromatic components could be affected by different environmental factors leading to variation of noncoding regions in the genome, which finally results in different fragrance and taste in different tobacco cultivars.     

烤烟种质资源差异性分析

对700多份烤烟种质资源的形态特征、主要性状、抗病性和烟叶化学成分等方面的差异性进行研究和分析,结果表明：烤烟种质资源形态差异主要表现在株型、叶形、叶耳、叶尖、茎叶角度和花序等特征上;主要性状差异表现在株高、打顸株高、叶数、节距、茎围、腰叶的长和宽等性状,其变幅分别为97．0—326．0cm、75.3～133.7cm、8．6—54片、2．6—10．1cm、6.3～13．8cm、41．6-99．0cm、16.6—47．8cm;脚叶、腰叶和项叶的长宽比变幅分别为1．3～3,7、1．5—4．9、1．6～5．8;移栽至现蕾的天数变幅为30-100d;各性状指标的变异系数在7．8％-30．3％之间。不同种质间烟叶化学成分差异较大,变异系数在16．4％-69．2％之间。在702份种质中,抗病、中抗、感病和中感病的种质分别占18．2％、36．0％、6.6％、39．0％,可提供丰富的种质资源供育种选择利用。     

烟草EST-SSR位点分析

利用MISA软件对烟草EST公共数据库中的简单重复序列(SSRs)进行了分析。结果表明,在133523条EST序列中,共获得81757条SSR序列,SSRs之间的距离约为0.92 kb。其中,六碱基重复丰度最大,占60.3%,而单碱基、三碱基、四碱基、二碱基和五碱基重复丰度分别为20.0%、11.0%、4.2%、2.8%和1.7%。在单碱基、二碱基、三碱基和四碱基重复模体中,丰度最大的分别是A/T、AG、AAG和AAAT,而CG在编码区内丰度很低。用CAP3软件进行冗余分析表明,在这6种类型的重复模体中,冗余与非冗余的烟草EST之间没有显著差异。在得到的SSR序列中随机选择10个序列设计引物,在7个烟草品种中进行PCR扩增。结果表明,10对引物全部扩增出PCR产物,其中8对引物扩增出预期片段。用这8组扩增出预期片段的PCR产物进行变性PAGE凝胶电泳检测,结果表明,其中有4对引物(EB4、EB5、EB6和EB8)扩增出多态性条带。     

Steryl Glycoside Formation in Seedlings of Nicotiana tabacum L

Particulate enzyme preparations from tobacco seedlings (Nicotiana tabacum L.) were used in the synthesis of steryl glycoside. The data obtained by measuring cholesterol-4-¹⁴C incorporation generally agree with results obtained with UDP-glucose-¹⁴C. The in vitro reaction was linear for the first 10 minutes and had a pH optimum of 7.0 to 7.4. Addition of ATP activated while UDP-glucose inhibited slightly the reaction. In short term experiments, the percentage disappearance of endogenous and added sterol was about the same.     

An Arabinogalactan-protein from the Leaves of Nicotiana tabacum

The signals of fatty acids in the form of triglycerides were observed in the ¹³C NMR spectrum of an intact soybean seed. The major fatty acid component composition of triglycerides in a soybean seed, which includes linoleic acid, oleic acid and palmitic acid, was estimated by subtracting the spectra of authentic fatty acids from the spectrum of the intact soybean seeds. The fatty acid compositions of seeds of 11 Japanese soybean cultivars and 5 lines bred at the Asian Vegetable Research and Development Center (AVRDC) were estimated by this rapid (within lhr for one seed) and nondestructive analytical method.     

烟草雌、雄配子DNA 含量变化

采用显微分光光度法测定了烟草( Nicotiana tabacum) 精细胞和卵细胞的DNA 含量。烟草是二胞花粉, 花粉萌发后生殖细胞在花粉管中分裂形成精细胞。授粉后45 h 花粉管到达子房, 在花粉管内的精细胞DNA 含量为1C。当花粉管在退化助细胞中破裂, 释放出的两个精细胞开始合成DNA。在与卵细胞融合前,两个精细胞DNA 含量接近2C。随着精细胞的到达及合成DNA, 卵细胞也开始合成DNA, 融合前的卵细胞DNA 含量也接近2C。精、卵细胞融合后, 合子DNA 含量为4C。烟草雌、雄配子是在细胞周期的G2 期发生融合, 属于G2 型。     

烟草雌、雄配子DNA含量变化

采用显微分光光度法测定了烟草（Nieotiana tabacum）精细胞和卵细胞的DNA含量。烟草是二胞花粉,花粉萌发后生殖细胞在花粉管中分裂形成精细胞。授粉后45h花粉管到达子房,在花粉管内的精细胞DNA含量为1C。当花粉管在退化助细胞中破裂,释放出的两个精细胞开始合成DNA。在与卵细胞融合前,两个精细胞DNA含量接近2C。随着精细胞的到达及合成DNA,卵细胞也开始合成DNA,融合前的卵细胞DNA含量也接近2C。精、卵细胞融合后,合子DNA含量为4C。烟草雌、雄配子是在细胞周期的G2期发生融合,属于G2型。     

Further Characterization of Expression of Auxin-Induced Genes in Tobacco (Nicotiana tabacum) Cell-Suspension Cultures

We have described the modulation of four auxin-regulated genes during the growth cycle of suspension-cultured tobacco (Nicotiana tabacum [L.] var White Burley) cells. The genes were transiently expressed 2 to 8 h after transfer of stationary phase cells to fresh medium, during the transition from the quiescent phase of cells leaving the mitotic cycle to the synthesis phase of the cell cycle. After this transient induction, the cells showed a decreased sensitivity to auxin. Although the expression pattern suggests that induction of these genes might be important for cell division, over-production of antisense mRNA for one of these genes (pCNT103) did not influence cell division in transgenic tobacco cells. Furthermore, stimuli such as salicylic acid were capable of inducing gene expression but were unable to restore cell division. Although these data do not conclusively exclude a role for these genes in cell division, their significance in this process is discussed in view of their homology with other auxin-induced genes and in view of the specificity of hormone-induced early responses.