高羊茅光周期调控基因FaCONSTANS亚细胞定位与差异表达分析

构建高羊茅光周期调控基因FaCONSTANS与绿色荧光蛋白基因GFP融合的植物表达载体P-FaCONSTA NS-hGFP,基因枪转化法转入洋葱表皮细胞,荧光显微镜检测融合基因的瞬时表达.并运用实时荧光定量PCR研究光周期调控基因FaCONSTANS在长日照与短日照条件下昼夜24 h的表达差异.结果表明:FaCONST...     

高羊茅FaChit1启动子的克隆及其与gus基因融合表达载体的构建

本研究以获得的FaChit1基因cDNA序列设计引物,采用染色体步移技术从高羊茅基因组中分离了一段FaChit1基因启动子区域。序列分析结果显示,该启动子长度为935bp,含有保守性元件TATA和CAAT-Box,以及与植物逆境胁迫应答相关的多个顺式作用元件,如W-box、ABRE、MYB及MYC转录因子结合位点等。另外构建了含不同长度FaChit1启动子区域(-935bp、-651bp、-233bp)与gus基因融合植物表达载体,分别命名为pFaChit1P-Ⅰ、pFaChit1P-Ⅱ和pFaChit1P-Ⅲ,为进一步进行该启动子的功能分析奠定基础。     

植物开花光周期反应的分子调控机制

植物开花时间受到日照长短季节性变化的调节,拟南芥和水稻中与光周期反应相关基因的分离,使人们得以认识植物开花光周期反应的分子调控机制。植物感知日照长短的变化主要由CONSTANS(CO)基因的表达所控制。CO能够将光信号与生物钟信号整合,调节开花基因FLOWERING LOCUS T(FT)的表达,并最终控制植物的开花时间。本文对这一研究的最新进展进行了综述。     

光伏提水技术在西藏的推广前景

为了西藏能源短缺的农牧区发展光伏提水技术提供科学依据,本研究介绍了太阳能光伏提水技术的发展现状,分析了西藏地区的太阳能资源储量,以及光伏提水技术在西藏地区的推广应用优势及潜力,并对其进行了经济评价。     

四川白鹅PRL基因的克隆与生物信息学分析

为了探明鹅就巢性产生的分子机理及不同鹅种之间的遗传分化,采用RT-PCR方法从四川白鹅垂体组织克隆到了PRL基因,并对其核苷酸序列和推导的氨基酸序列等进行了分析.四川白鹅PRL基因编码区含有690个核苷酸(GenBank NO.GQ202542),编码229个氨基酸,与其他禽类PRL具有高度保守性;其中与东北白鹅和籽鹅...     

CCT基因调控热带甜玉米光周期敏感性

热带甜玉米种质具有抗病性好、株型紧凑、产量高等优点,然而光周期敏感性严重制约了热带甜玉米种质在中国大部分地区的利用。关于甜玉米光周期敏感性的调控机制尚不清楚。本研究调查了50份热带和温带甜玉米自交系材料的光周期敏感性,同时分析了甜玉米自交系中主要光周期控制关键CCT基因的基因型和表达。结果显示热带甜玉米自交系抽穗、吐丝和散粉期的光周期敏感性指数都显著大于温带甜玉米自交系;启动子和UTR区域的突变影响了热带甜玉米自交系中ZmCOL3和ZmCCT9的表达,暗示了ZmCOL3和ZmCCT9可能是调控热带甜玉米自交系光周期敏感性的关键因子。本研究为热带甜玉米种质的利用和遗传改良提供基础。     

枸杞LbCO基因的克隆与生物信息学分析

CONSTANS(CO)基因是高等植物叶片中光周期途径的关键成分,生物钟及光信号控制CO基因的表达。本研究以宁夏枸杞(Lycium barbarum L.)品种之一‘宁杞7号’叶片为实验材料,通过转录组测序结果筛选,利用RT-PCR技术扩增得到‘宁杞7号’的CO基因cDNA全长序列,命名为LbCO。利用生物信息学手段对所获得的序列进行结果及功能预测,结果显示:LbCO基因的cDNA序列全长1224 bp,编码407个氨基酸,分子质量为44.83 k D,理论等电点pI=5.58,不稳定指数为39.66,是一种稳定的非分泌蛋白。该蛋白亚细胞定位于细胞核,二级结构中无规则卷曲占比最高(54.48%),其次为α-螺旋(28.99%)。系统进化分析表明枸杞LbCO蛋白与其它物种的CO蛋白具有较高的同源性,其中与辣椒属CO蛋白亲缘关系最近。本研究为后续进一步探讨该基因编码的蛋白在调控枸杞花期研究方面提供理论参考。     

高羊茅种子产量因子与产量的通径分析

采用多因素正交试验设计,通过大样本相关、通径和逐步回归分析后表明,高羊茅5个种子产量因子对种子产量的直接贡献大小排序为:生植枝数/m2＞小花数/小穗＞种子粒数/小穗＞单粒种子重＞小穗数/生植枝;提高小花数/小穗和种子粒数/小穗可最有效提高高羊茅的种子产量,其次是提高生植枝数/m2.     

高羊茅对重金属镉的抗性分析

为了解不同高羊茅品种对重金属镉的抗性差异,探讨其修复镉污染土壤的作用,通过形态和生理指标测定,对不同浓度镉胁迫前后的高羊茅品种形态和生理指标进行比较分析。结果表明：低浓度氯化镉胁迫下,高羊茅品种‘猎狗’的茎叶鲜重、最大根长、根鲜重、叶绿素含量等指标呈增长趋势,到达一定阈值后则受到抑制;相对电导率、MDA含量、脯氨酸含量则随镉浓度的增大而持续升高;根系活力则表现为随镉浓度的升高而逐渐下降。进一步利用50 mg/L的氯化镉胁迫处理9个高羊茅品种,对茎叶鲜重、根鲜重、最大根长等形态指标的变化情况进行分析评估。试验结果显示,多数品种的抗性与根部性状的表现更为密切;综合抗性由强到弱的排列顺序为：‘交战Ⅱ’>‘毕加索’>‘猎狗’>‘杰作’>‘新秀’>‘碧翠A’>‘可奇思’>‘爱瑞’>‘达芬奇’。对于镉污染程度较高的土壤,可选择综合抗性较高的品种‘交战II’进行治理修复;对于镉污染程度较低的土壤,可选用抗性一般但地上部分生物量较高的品种‘新秀’。     

高羊茅高效组织培养再生体系的建立

植物分子育种与基因功能鉴定是建立在高效的组织培养再生体系基础上的。本研究以高羊茅(Festuca arundinacea Schreb)为材料,探讨了不同外植体状态、基本培养基类型、外源激素组合与固化剂种类与浓度对愈伤组织诱导的影响,并进行了愈伤组织分化长芽与生根研究。结果表明:愈伤组织诱导过程中,离体成熟胚比完整种子作外植体愈伤组织出现早、质量优、诱导率高;基本培养基的选择上以N6诱导率最高,MS最低;激素组合研究中高羊茅三个品种(SAFari, FirewarⅡ, Barleduc)在无6-BA培养基中,生长激素选择上都是2,4-D优于IAA,2~2.5 mg/L 2,4-D在三个品种中诱导率都在50%以上,而在添加0.5 mg/L 6-BA时诱导率都偏低;固化剂选择上则以Phytagel与Agarose二都最优,Phytagel固化剂最佳诱导浓度为2~4 g/L。分化长芽最佳培养基为MS+0.1 mg/L 2,4-D+0.1 mg/L 6-BA+0.5 mg/L NAA+20 mg/L AgNO3+2%蔗糖+0.6%琼脂,生根壮苗最佳培养基为1/2MS+0.5 mg/L NAA+3%蔗糖+0.3%Phytagel。本研究为高羊茅的高效转基因体系建立与CRISPR基因功能研究提供准备。     

CO基因的调控表达与植物光周期反应

光周期是影响植物花发育的重要环境因素,CONSTANS（CO）是拟南芥光周期遗传调控途径中的关键因子,目前已在许多植物中发现了具有N 端类锌指结构B-box和C端CCT 保守结构域的CONSTANS-like (COL)基因,但它们的功能却不尽相同。本文概述了CO基因的结构、家族成员及其在光周期途径中的作用机制,以期为植物光周期遗传机理研究提供参考。     

农杆菌介导的高羊茅遗传转化体系的优化

以高羊茅品种猎狗5号的胚性愈伤组织作为受体材料,通过gus基因瞬时表达检测,探讨预培养培养基、预培养时间、菌液浓度、感染时间、乙酰丁香酮浓度、共培养条件等不同因素对农杆菌介导转化的影响,结果表明,高羊茅品种猎狗5号遗传转化优化条件为胚性愈伤组织先在预培养培养基(MS 6-BA0．5mg／L NAA0．5mg／L)预培养5d;然后用农杆菌菌液(OD600=0．5),感染时间10-20min,在23-25℃的温度下,共培养基(MS 蔗糖30g／L 葡萄糖10g／L CA0．5g／L Glu0．5g／L 2,4-D1．5mg／L AS 150μmol／L agar 8g／L,pH5．2～5．6)上共培养3d。在共培养基中添加AS 150μmol／L有助于提高gus基因瞬时表达率。     

Cloning and Sequence Analysis of Lipoxygenase Gene cDNA from Cucumber Fruit （Cucumis sativus L.）

Lipoxygenases are nonheme-iron-containing dioxygenases that catalyze the hydroperoxidation of unsatrated fatty acids containing a cis, cis-1,4-pentadiene structure producing hydroperoxy acids with conjugated dienes. LOX activity has been found in a wide range of plants. Typical substrates for LOX in plants are linoleic acid and linilenic acid fatty acids. The function of various LOXs in plants is unknown, but their participation in all stages of plant growth and development has been suggested （Hildebrand, 1989; Siedow, 1991）. Some of the physiological processes in whicn lipoxygenses have been implicated include wounding （Saravitz and Siedow, 1996）, pathogen attack （Melan et al., 1993）, seed germination （Kato et al., 1992）, fruit ripening （Ferrie et al., 1994）, plant senescence （Paliyath and Droillard, 1992）. The study on the role of lipoxygenase in ripening and senescence fruit focused on tomato and strawberry. Cloning LOX gene of cucumber fruit will make us further understand the molecularaction mode of this enzyme during fxuit ripening and senescence. In this paper we isolated the partial nucleotide sequences of cucumber fiuit lipoxygenase gene and discuss the characterization of it.     

草莓果实多酚氧化酶基因的克隆及序列分析

多酚氧化酶（polyphenol oxidase,PPO）是一类植物体内广泛存在的由核基因编码的能与铜结合的金属蛋白酶。它是诱导许多果蔬等农产品产生酶促褐变的主要原因,同时在植物的生长发育、抗病虫害及果实品质形成等方面也起到了一定的作用。本文用RACE的方法,从草莓幼果cDNA中成功扩增出PPO基因的3′端部分序列。序列分析表明该基因片段编码424个氨基酸。与其他物种PPO核苷酸序列比较相似性为72％-80％,氨基酸相似性为82％～87％。草莓多酚氧化酶基因的成功克隆,为草莓抗褐变的研究及果实品质的改良打下了坚实的理论基础。     

Genetic Diversity in Tall Fescue Ecotypes from Alabama I. Maturity, Morphological Traits, and Disease Reaction

There is a large hectarage of tall fescue in Alabama. Recognition of the deleterious effects of the endophytic fungus Acremonium coenophialum has resulted in substantial acceptance of new endophyte-free cultivars. Destruction of old pastures and the concomitant loss of potentially valuable adapted germplasm could be a valid cause for concern. The objectives of this study were to evaluate genetic variation for maturity and plant morphological traits, both among and within tall fescue cv. ‘Kentucky 31’ populations from 18 to 38 year-old pastures in Alabama. Ten populations of approximately 50 plants each were collected from old Alabama tall fescue pastures. Two pastures were sampled in each of five counties, representing the five major land use areas of the state. In 1990, spaced plants of all populations were evaluated at heading time for maturity, morphological, and disease traits at two locations in central Alabama. Endophyte infection level of the populations ranged from 2 to 100 %. Maturity was highly correlated with tiller length and flag leaf width and was used as a covariate for these traits in the analysis of variance. Leaf rust and net blotch ratings were negatively correlated. Significant variation among populations was observed for maturity and flag leaf dimensions, most variation being due to differences among source counties. Significant variation was observed within all populations for maturity and morphological traits, but only in seven populations for leaf rust. No within-population variation was detected for net blotch. The average precipitation for the month of May explained 81 % of the variation among populations for maturity. Age of the pasture sampled had a significant effect on tiller diameter (r²= 0.55), and latitude of the pasture sampled was the best individual predictor of leaf width (r²= 0.42). This study reveals that considerable genetic diversity, largely influenced by climatic conditions, exists among ‘Kentucky 31’ tall fescue ecotypes from Alabama.     

Continuous Plant Regeneration from Established Embryogenic Cell Suspension Cultures of Italian Ryegrass and Tall Fescue

The suitability of different protocols was compared for entire plant regeneration by somatic embryogenesis, of the forage plants Lolium multiflorum Lam. (Italian ryegrass) and Festuca arundinacea Schreb. (tall fescue). In the first protocol, miniature embryos were used as starting material, while mature seeds were retained in the other two. Whichever the considered protocol, undifferentiated calli were produced on Murashige and Skoog MS medium supplemented with 2,4-D. The calli were subcultured in the dark on solid MS agar medium, containing 5 mg/1 2,4-D (protocol 2) or on solid MS medium followed by transfer to a rotated liquid MS medium with 2 mg/1 2,4-D (protocol 1). In these conditions, induction of somatic embryogenesis occurred, and whole plants were regenerated during a limited lapse of time, upon transfer in the light, to MS medium supplemented with BAP but devoid of 2,4-D. The simultaneous elimination of 2,4-D and transfer to light appeared essential for full regeneration of the plants. Using this characteristic, an additional step was added to a new protocol (protocol 3) in which microcalli, cultured on liquid MS medium containing 5 mg/1 2,4-D, were transferred to the same medium with 2 mg/1 2,4-D, in the dark. In these conditions, the suspensions kept their embryogenic potential for months. In all cases, plantlets were successfully transferred into the soil. An evaluation of the somaclonal variation potential of the plants issued from each protocol is now underway.     

三河马生长激素基因的克隆与序列分析

为进一步探明三河马遗传分化,参考牛(M57764)、羊(AF002110)、猪(M17704)、人(J03071)和鼠(X12630)等哺乳动物GH基因序列,设计一对特异引物,用PCR方法从三河马基因组中扩增出一条长约2kb的DNA片段。PCR产物经pGEM-Teasy载体转化感受态DH5α株大肠杆菌,获得重组克隆子。DNA测序表明:三河马生长激素基因DNA序列长1923bp,含完整的5个外显子和4个内含子,与猪、狗、牛、羊的GH的cDNA序列同源性分别为94.47%,92.63%,90.06%和90.37%。其cDNA所编码氨基酸与猪、狗、牛、羊同源性分别为97.21%,96.74%,88.84%和88.37%,表明该基因在进化过程中是保守的。