一个与净叶黄抗赤星病基因紧密连锁的SSR标记

为了在分子水平上弄清烟草赤星病抗性的遗传规律,并进行遗传定位,以抗赤星病品种净叶黄和感赤星病品种NC82为亲本,构建了F₁、F₂、BC₁ 代群体。通过对该群体进行赤星病接种鉴定和抗性遗传分析,发现净叶黄对赤星病的抗性由显性多基因控制。通过分子标记群体扩增,在第M号连锁群上,筛选到一个与净叶黄的赤星病抗性基因紧密连锁的SSR标记,它与抗性基因间的遗传距离为4 cM。该标记可用于抗赤星病育种的辅助选择。     

烟草种质的SSR标记遗传多样性及青枯病抗性的关联分析

烟草青枯病是影响烟草产质量的主要病害之一,开发出与烟草青枯病抗病性密切相关的优异等位基因,可加速抗病品种的选育进程。据此,本研究选用94份烟草种质材料作为研究对象,运用236个SSR标记对这些材料进行基因分型和群体结构分析,开展基于连锁不平衡的关联分析探测与青枯病抗性相关的分子标记。结果表明,236个SSR标记在94份烟草种质中检测到904个等位变异位点,平均每个SSR标记为3.83个位点;这些SSR标记的多态性信息量（PIC）值在0.1361~0.8760之间,平均为0.5136。基于SSR基因分型数据,构建了126个标记单倍型;在此基础上进行群体结构分析,将94份种质划分为2个类群,表明本研究烟草种质的群体结构简单,有利于减少伪关联。二年田间抗病性数据与单倍型的关联分析发现7个单倍型与青枯病抗病性密切相关,其中有3个单倍型在二年的田间试验中均表现出与青枯病抗性密切相关,平均解释率超过10%。研究结果可为烟草青枯病抗病材料的分子标记辅助筛选提供参考。      

基于RAD-seq数据开发烟草多态性SSR标记

采用RAD-seq技术对两份烟草材料"118-3"和"粤烟98"进行简化基因组测序,分别获得45 119 346和50 360 738个高质量干净读长（HQ clean reads）。通过序列分析,在22 145个reads覆盖的参考基因组序列中共检测到25 343个SSR位点,其中以二和三核苷酸重复基序最为丰富,占总SSR位点的71.59%;除单核苷酸类型外,SSR基序的重复次数主要集中在4~9之间。根据SSR位点两翼的序列,利用Primer 3.0成功设计出23 346对SSR引物,引物设计的成功率为92.12%。依据测序数据初步发现这些SSR中有323个（1.38%）SSR在两份测序材料间具有多态性,随机选择其中50个SSR在4份烟草材料中进行验证,结果发现在两份测序材料间的多态率为76%,存在24%的假阳性;在另两份烟草材料中多态率为20%。研究结果表明,利用简化基因组测序技术能够开发大量SSR标记和发现样品间具有多态性的SSR标记,可为SSR标记的开发和应用提供基础。     

烟草青枯病抗性基因的遗传分析及RAPD标记

选用高感青枯病品种"红花大金元"与高抗青枯病品种"Ti448A"配制杂交组合,采用温室苗期叶片人工注射法接种鉴定两亲本及其杂交后代F₁、F₂、BC₁P₁代的青枯病抗性表现,结果表明抗病亲本材料Ti448A的抗病性由一对显性基因控制。依据混合群体分组分析法(BSA)建立抗青枯病基因池和感青枯病基因池,通过RAPD试验,从近200个随机引物中筛选出一个在两池间具有多态性的引物S503,用双亲、F₁、F₂单株DNA进行RAPD试验证明了该引物扩增出的特异性片段S503-750是一个与抗青枯病基因连锁的RAPD标记,利用JOINMAP (version 1.4)软件计算得此标记与抗青枯病基因间的重组率为5.984%,连锁距离为6.261 cM。      

烟草SSR荧光标记与毛细管电泳检测技术研究

将毛细管电泳荧光检测技术应用于烟草SSR标记分析,以期借助自动化、智能化的仪器设备加速对烟草遗传分析的研究进程。采用毛细管电泳荧光检测技术和聚丙烯酰胺凝胶电泳技术,对9份烟草材料进行SSR位点分析,用相同的材料和引物,对这两种方法的检测结果进行比较,建立并优化了毛细管电泳荧光检测法应用于烟草SSR标记分析的技术体系。结果表明,荧光检测法克服了银染法的不足,具有简便、可靠及高通量的优点。同时对微卫星荧光标记技术中低成本、高通量多重PCR体系建立的一些技术问题进行了探讨。     

普通烟草种质资源的SSR标记与指纹图谱分析

利用SSR标记构建了烟草核心种质中80份普通烟草种质的指纹图谱。从286对SSR引物中筛选出8对多态性较好的引物,对这些烟草种质进行分析,共检测到85个多态性位点。研究表明,这8对多态性SSR引物可以将80份烟草种质完全区分开,每份种质都有各自独特的指纹图谱,表明SSR标记完全适用于普通烟草种质鉴定和指纹图谱构建工作。     

大豆对SMV1号株系的抗性遗传分析及抗病基因的SSR标记

利用杂交组合合丰25（S）×东农93-046（R）的F1、F2和F2∶3群体进行了抗病性鉴定,结果表明：F1在接种后表现抗病,F2群体分离比例为3（抗）∶1（感）,对F2衍生的F2∶3家系接种鉴定,纯合抗病家系、抗感分离家系和纯合感病家系的比率符合1∶2∶1,表明东农93-046对SMV1号株系的抗性由一对显性基因（RSMV1）控制。并利用东农93-046（R）×Conrad（S）的正反交组合F2进行抗性鉴定,结果正反交后代均表现为3∶1的分离比例,无细胞质效应,进一步证明了东农93-046是受一对基因控制的抗性种质。经改良的分离群体组群分析法研究发现,东农93-046抗SMV1的位点（RSMV1）位于F连锁群上,与SSR标记的连锁顺序为HSP176、Satt114、RSMV1、Satt510、Sct_033、Satt334、Satt362,标记与RSMV1之间的遗传距离分别为10.9cM、4.3cM、6.6cM、11.7cM、18.0cM和35.3cM。根据标记HSP176选择基因型纯合和杂合的抗病植株,其符合率分别达到100.0%和94.5%。     

烟草青枯病抗性与分子标记的关联分析

为了找到与烟草青枯病抗性相关的分子标记、加快抗病品种的选育,以78份烟草种质为自然群体材料,选用20对简单重复序列(Simple Sequence Repeat,SSR)和37对微卫星锚定片段长度多态性(Microsatellite-anchored Fragment Length Polymorphism,MFLP)引物组合对烟草种质材料进行基因分型,并利用等位变异数据进行主成分分析和群体结构分析,同时采用TASSEL3.0软件对烟草青枯病抗性与等位变异进行连锁不平衡关联分析。结果显示:SSR和MFLP标记在78份烟草种质中共检测到252个等位变异,主成分分析和群体结构分析均将烟草种质分为5个组群;关联分析发现6个MFLP标记位点与烟草青枯病抗性显著相关,对青枯病抗性的解释率在8.33%~19.49%之间。这些分子标记可为评价具有青枯病抗性潜力的烟草种质材料提供依据。      

基于SSR分子标记的部分烤烟种质资源遗传多样性研究

为准确评价部分烟草种质资源的遗传多样性,利用42对已发表多态性较好的烟草SSR标记引物对38份烤烟种质资源进行分析,筛选出22对引物在这些种质间存在多态性,平均等位位点数为3.68个,Nei's多样性指数平均为0.451,Shannon信息指数平均达0.789。UPGMA聚类分析表明,红花大金元有明显特异性。遗传结构分析显示,参试材料可分为2个类群,类群1和类群2分别包含16份和22份种质材料,且类群1变异范围大于类群2,类群2材料多表现为自然株高在210 cm以下,叶数20~25,茎围10~11 cm,田间赤星病较严重,青枯病发病相对较轻。研究结果为提升SSR分子标记筛选效率和部分烤烟种质创新应用提供理论依据。     

烟草PVY隐性抗病基因的分子标记及其适用性

马铃薯Y 病毒(Potato virus Y,PVY)是危害烟草的重要病害, 种植抗病品种是经济有效的防治措施。分子标记辅助选择可提高抗病育种效率。来源于X-射线诱变的Virgin A Mutante(VAM) 的隐性抗PVY 基因位点(va)被广泛应用于烟草抗病育种。为了提高va 位点的育种利用效率, 根据烟草隐性抗PVY 基因(感病基因)eIF4E-1 基因序列, 设计特异扩增的引物CF2GR11,开发eIF4E-1 基因的分子标记, 并检测了该标记与抗性的遗传距离和在常见烟草资源中的适用性。CF2GR11 在云烟87、红花大金元和K326 等感PVY 品种可扩增出500 bp 产物, 在NC102、NC55 和K326PVY 等抗病品种无扩增条带。以抗、感PVY 亲本构建的101 个F₂ 单株为定位群体, 遗传连锁分析表明, CF2GR11 标记与烟草PVY 感病基因的遗传距离为0.99 cM。对46 份PVY 抗性明确的栽培烟草资源的检测表明, 供试资源的标记检测结果与抗性的吻合度为100%, 表明CF2GR11 标记适用性高。该目的基因标记可用于抗PVY 育种的辅助选择和抗PVY 资源的鉴定。      

Characterization of Callosobruchus chinensis (L.) resistance in Vigna umbellata (Thunb.) Ohwi & Ohashi

Resistance to azuki bean weevil, Callosobruchus chinensis, was studied in a series of field and laboratory experiments in two accessions of rice bean (Vigna umbellata (Thunb.) Ohwi & Ohashi), one accession of black gram (V. mungo (L.) Hepper), and one accession of mungbean, (V. radiata (L.) Wilczek). Weevil damage to immature pods of the rice bean accessions, ‘Menaga’ and ‘Miyazaki’, was significantly less than to the susceptible mungbean, VC1973A. In mature pods, the pest damage to the pod wall of Menaga was significantly higher than to VC1973A, whereas the damage to Miyazaki was similar to VC1973A. Seeds within the pods of both rice bean accessions were resistant no matter when the pods were harvested. When the insects were exposed directly on dry seeds, both rice bean accessions and a black gram accession VM2164 were resistant to them. In artificial seeds made by mixing flour of the individual resistant Vigna accessions with VC1973A and subsequently exposed to bruchid oviposition, the higher the quantity of resistant Vigna flour the lower the number of bruchids that emerged from such seeds. No bruchids emerged from artificial seeds containing crude starch fraction from the three resistant Vigna accessions when such seeds were exposed to bruchid infestation, whereas many insects emerged from the seeds containing starch of VC1973A or flour of VC1973A alone. In artificial seeds made by mixing crude protein fractions of the three resistant Vigna accessions with flour of VC1973A, as the concentration of protein increased the number of C. chinensis adults that emerged decreased. Fractionation of crude proteins into acetone-precipitable proteins and peptide and amino acid portions resulted in the loss of antibiosis effect. Artificial seeds made from purified starch-polysaccharides fraction, however, exhibited antibiosis effects if prepared from the rice bean seed of Menaga and Miyazaki but not if made from the black gram seed, VM2164.     

Fat and fatty acids of white lupin (Lupinus albus L.) in comparison to sesame (Sesamum indicum L.)

The study was undertaken to compare fat and fatty acid profiles in white lupin (Lupinus albus ssp. albus) and sesame (Sesamum indicum L.), representing two different families, Fabaceae and Pedaliaceae. Fat levels were 10.74% and 55.44% in seeds of white lupin and sesame, respectively. The results indicated that oleic, linolenic and arachidic acids in seed fat were higher in white lupin than in sesame cultivars. Oleic acid was the predominant fatty acid in white lupin, whereas linoleic acid was predominant in sesame. Fat content (%) was statistically significantly correlated with linoleic, linolenic and arachidic acids at the genotypic level. The fatty acid composition of white lupin is useful for human consumption. Although oil content of white lupin was lower than that of sesame, white sweet lupin could be improved.     

Investigation of organic extractives from unifloral chestnut (Castanea sativa L.) and eucalyptus (Eucalyptus globulus Labill.) honeys and flowers to identification of botanical marker compounds

Ultrasound-assisted extraction was employed to investigate the composition of extractable compounds from unifloral chestnut and eucalyptus honeys. 1-phenylethanol and 2′-aminoacetophenone are the most powerful botanical markers of chestnut honey. Additionally, cis-cinnamyl alcohol, and p-hydroxyacetophenone were found exclusively in chestnut honey. Organic extractives from whole chestnut flowers were predominated by 1-phenylethanol, nonanal, benzyl alcohol, and nonanoic acid. Of the 16 compounds found in the flower extract, 13 were present in honey as well. Some of them are transferred directly to honey, while in other cases modification reactions take place. Eucalyptus honey is characterized by 2-hydroxy-5-methyl-3-hexanone and 3-hydroxy-5-methyl-3-hexanone, as well as exo-2-hydroxycineole and an unknown norisoprenoid. Aside from, acetoin, nonanal, methyl nonanoate, and dehydrovomifoliol were present in higher concentrations. In eucalyptus flower extracts, norisoprenoids were the most abundant compounds, all of which were also present in the honey samples. 3-Oxo-α-ionone comprised half of the total amount, followed by eucalyptol.     

普通烟草长叶柄突变性状的遗传分析

叶柄是烟草叶片的重要组成部分,烟草长叶柄性状遗传分析可用于研究烟草叶柄和叶片的发育。利用甲基磺酸乙酯（EMS）诱变普通烟草烤烟品种中烟100和红花大金元各获得一个（极）长叶柄突变体（M48和M50）,将两个突变体分别与各自的野生型中烟100和红花大金元及普通烟草烤烟品种革新三号杂交获得F₁,F₁自交获得F₂群体,F₁与其相应的野生型亲本回交获得BC₁F₁群体。对各类型材料进行表型调查和统计分析结果显示:F₁均表现为（极）长叶柄突变表型,在F₂群体和BC₁F₁群体中,经x2检验,（极）长叶柄突变表型与野生型表型的理论分离比均符合3:1和1:1,表明（极）长叶柄突变性状由染色体上一对显性单基因控制。上述结果为进一步定位和克隆（极）长叶柄突变基因,揭示其在烟草叶柄和叶片发育中的作用奠定基础。      

Extrusion of a hard-to-cook bean (Phaseolus vulgaris L.) and quality protein maize (Zea mays L.) flour blend

Heated extrusion was tested as an alternative process for incorporating “hard-to-cook” beans into food products. A 3² factorial design was used to evaluate extrusion conditions for a 40/60 (w/w) blend of “hard-to-cook” beans and quality protein maize. Tested extrusion variables were temperature (155, 170 and 185 °C) and moisture content (15.5, 17.5 and 19.5 g/100 g). Screw speed was fixed at 130 rpm. The extrudates obtained at 155 and 170 °C with 15.5% moisture had the best physical characteristics and were chosen for comparative analysis of nutritional changes between the unprocessed “hard-to-cook” bean/quality protein maize flour blend and the resulting extrudates. In vitro protein digestibility was higher in the extrudates (80%) than in the flour blend (76%). In vitro starch digestibility was higher at 155 °C (89%) and 170 °C (92%) than in the flour blend (12%). Processing conditions decreased dietary fibre content by 38% at 155 °C and 44% at 170 °C.     

Flavonoids of Zizyphus jujuba L. and Zizyphus spina-christi (L.) Willd (Rhamnaceae) fruits

The ripe edible fruits of jujube, Zizyphus jujuba Miller (syn. Z. sativa Gaertner, Z. vulgaris Lam.) and Christ’s thorn jujube Zizyphus spina-christi (L.) Willd (Rhamnaceae family) were phytochemically investigated, comparing their quali-quantitative flavonoids profile. Twelve compounds from both methanol extracts have been recognized as quercetin, kaempferol, and phloretin derivatives by means of HPLC/ESI-MS analyses. Six major compounds have been purified by Sephadex LH-20 column chromatography followed by HPLC and were characterized using NMR spectroscopy. One C-glycoside, 3′,5′-di-C-β-d-glucosylphloretin, was detected in Z. spina-christi. The quantitative analysis of all compounds was also reported showing a higher content of flavonoids in Z. jujuba.     

Repellence and fumigant toxicity of propionic acid against adults of Sitophilus granarius (L.) and S. oryzae (L.)

The biological activity of propionic acid towards adults of the primary stored-grain pests, Sitophilus granarius and S. oryzae was investigated by electroantennographic, behavioural and fumigant toxicity assays. Electroantennograms revealed the sensitivity of both sexes of the two species to propionic acid. In two-choice pitfall bioassays, the compound showed dose-dependent repellent effects even in the presence of wheat odours, which attracted the beetles. In the fumigation assay, propionic acid was effective in killing weevil adults. The LC₅₀ values, calculated for both species at 23 and 30 °C, ranged from 5 to 10 μg/L air and are comparable to those of other known fumigants. Propionic acid could have potential for applications in IPM programs for stored-grain beetles because of its safety, high volatility, repellency and fumigant activity.     

Insecticidal activities of constituents identified in the essential oil from leaves of Chamaecyparis obtusa against Callosobruchus chinensis (L.) and Sitophilus oryzae (L.)

Analysis by gas chromatography-mass spectrometry led to the identification of 20 volatiles from the steam distilled oil of the leaves from Chamaecyparis obtusa Siebold et Zuccarni. Seven constituents of the oil were tested for contact and fumigant activity against adults of Callosobruchus chinensis (L.) and Sitophilus oryzae (L.). Responses varied with compound and dose rather than increasing exposure time beyond 1 or 2 days. In the impregnated-paper test with the constituents [bornyl acetate, (+)-limonene, myrcene, α-phellandrene, α-pinene, sabinene and terpinolene], at 0.1 mg/cm², bornyl acetate, α-phellandrene and terpinolene caused 97%, 97% and 87% mortality, respectively, against C. chinensis adults 1 day after treatment. At 0.05 mg/cm², moderate activity was achieved with bornyl acetate (56% mortality), α-phellandrene (75%) and terpinolene (55%). Against S. oryzae adults, at 0.26 mg/cm², terpinolene caused 93% mortality 2 days after treatment, whereas 80% mortality 4 days after treatment was observed with bornyl acetate and α-phellandrene. The toxicity of these compounds was significantly decreased at 0.18 mg/cm². In a fumigation test with S. oryzae adults, bornyl acetate and terpinolene were much more effective in sealed containers than in open ones, indicating that the insecticidal activity of these compounds was largely attributable to fumigant action. These naturally occurring materials derived from leaves of Chamaecyparis could be useful for managing populations of C. chinensis and S. oryzae.     

Weight loss of wheat infested with gamma-radiated Sitophilus oryzae (L.) and Rhyzopertha dominica (F.)

Sitophilus oryzae (L.) and Rhyzopertha dominica (F.) adults were treated with 2·5, 5, 10, 25 or 50 krad of gamma radiation and confined on samples of uninfested wheat. Except for those treated at 2·5 krad the amounts of wheat consumed by the adults and their progeny were much less than that eaten by the controls. Five weeks after treatment with 25 krad, total feeding damage by R. dominica and S. oryzae was 11 and 3 per cent, respectively, of the control and most of this feeding occurred during the first week after treatment. The amount of damage caused by live but sterile insects in bulk grain treated with gamma radiation would probably not be a serious problem because of the great reduction in feeding brought about by the radiation.