马尾松捕光叶绿素a/b结合蛋白基因cab-Pm1的克隆与原核表达

光合作用是光合生物将光能转化为化学能,并同时将无机物转化成有机物贮存在生物体的过程.现已证明光合生物对光能的吸收、传递和转化都是在光合膜上具有一定的分子排列和空间构象的色素蛋白复合体上进行的(郁飞等,2001).在高等植物体内,捕光叶绿素a/b结合蛋白基因(cab)属于光合系统基因(Raghvendra,1998),其编码的蛋白质与色素形成捕光色素蛋白复合体(LHC),是一类把能量迅速传至反应中心引起光化学反应的色素蛋白复合体,大部分的叶绿素分子都结合在光系统I(PSⅠ)和光系统Ⅱ(PSⅡ)的天然色素蛋白复合物上.     

银杏叶绿素a/b结合蛋白基因(GbLhcb4)及其启动子克隆

为了探究银杏中叶绿素a/b结合蛋白基因(Gb Lhcb4)的结构及其相关的转录调控元件,基于Roche454高通量测序获得的Gb Lhcb4基因片段,采用RACE技术分离得到该基因的c DNA序列,并通过染色体步移技术获得该基因启动子序列,结合生物信息学技术分析基因序列的特征、系统进化地位及启动子区域的转录调控元件。序列分析表明,Gb Lhcb4基因全长1 200 bp,含有一个834 bp开放阅读框,编码277个氨基酸,相对分子量为30.65 k D,等电点为5.17。同源分析表明,该基因编码蛋白与北美云杉Ps Lhcb4蛋白(Gen Bank登录号:ABK21281.1)相似性最高,为74.7%。启动子顺式作用元件预测表明,启动子区域包含多个光响应元件及逆境响应元件,同时还存在激素响应、组织特异性表达、细胞周期等相关调控元件。此研究分离得到银杏Gb Lhcb4基因及启动子序列,并进行了相关的生物信息学分析,为探究银杏Gb Lhcb4基因应答环境信号的转录调控机制提供了分子基础。     

玫瑰β-1,3-葡聚糖合成酶基因(RrCalS)克隆与生物信息学分析

为了探明胼胝质对玫瑰授粉亲和性的影响,本研究以‘唐红’玫瑰花柱为试材,采用RT-PCR和RACE方法获得了玫瑰β-1,3-葡聚糖合成酶基因的cDNA全长,命名为RrCalS。该基因全长5742 bp,开放阅读框5295 bp,编码1764个氨基酸。推导编码蛋白的分子量为204.7kD, PI值为9.00,在164-265位具有pfam14288结构域,在869-1642位具有pfam02364保守结构域,属于葡聚糖合成酶超家族;该蛋白属于疏水性、非分泌型蛋白,具有16个跨膜结构域,含有32个Ser磷酸化位点、21个Thr磷酸化位点、12个Tyr磷酸化位点;该蛋白的α-螺旋占49.49%,无规则卷曲占22.68%,β-转角占7.94%;该蛋白与Fragaria vesca等8种植物的Cals氨基酸序列同源性达73%以上,且它们的系统进化关系与传统分类结果一致。本研究为进一步深入研究玫瑰授粉不亲和机理,提高玫瑰育种理论和技术水平奠定了基础。     

杜仲HDR基因全长cDNA克隆与序列分析

以杜仲叶片cDNA为模板,采用反转录RCR及RACE技术分离出HDR基因的cDNA克隆,命名为EuHDR。EuHDR基因cDNA全长1 653 bp,5’端非编码区长82 bp,3’端非编码区长188 bp,编码460个氨基酸,与喜树HDR基因序列相似性最高,达82%;推导EuHDR氨基酸序列中包含转运肽序列(A1-A33)及植物HDR蛋白多个保守的功能位点(A117,A208,A262,A345);EuHDR蛋白二级结构α-螺旋占35.65%,β-折叠占19.78%,螺环结构占44.57%;EuHDR蛋白三级结构为单体形式,呈不规则的三叶草形状;系统进化分析表明EuHDR蛋白与葡萄HDR蛋白的亲缘关系最为接近。     

白蜡虫β1-微管蛋白基因cDNA克隆及序列分析

采用RACE和RT-PCR对白蜡虫雌成虫β1-微管蛋白基因进行了克隆,获得的cDNA全长序列为1 492 bp,完整开放阅读框为1 341 bp,编码447个氨基酸残基,包含β微管蛋白的保守区域和GTP结合位点,理论分子量约为50.20 Kda,等电点为4.75。同源性分析表明:所获得的β1-微管蛋白与其它昆虫的β微管蛋白基因在氨基酸水平上是高度同源的,该基因cDNA序列已经递交到Genbank,获得的登录号为JF731244。     

松材线虫果胶酶基因Bxpel1的克隆及其生物信息学分析

以松材线虫Bursaphelenchus xylophilus转录本为模板,通过RT-PCR技术克隆果胶酶Bxpel1基因。结果表明,松材线虫果胶酶Bxpel1基因序列全长为795bp,GC含量为50.44%,编码252个氨基酸。通过Blast比对克隆的Bxpel1基因与已知序列(Gen-Bank ID:AB232908.1)的同源性达到98%。系统进化分析表明,Bxpel1基因编码产物的氨基酸序列与拟松材线虫Bursaphelenchus mucronatus,燕麦真滑刃线虫Aphelenchus avenae的Bxpel1蛋白序列具有高度同源性。Bxpel1基因编码的产物的0～25序列有可能是跨膜结构域,而其他序列均位于膜外,其二级结构主要是以无规则卷曲和β-折叠为主,信号肽的剪切位点位于第17至第18位氨基酸之间,主要在细胞外发挥生物学作用。Bxpel1基因的克隆及其生物信息学分析对进一步研究Bxpel1基因在松材线虫致病过程中的功能具有重要意义。     

杨树热激转录因子HsfAld的克隆、表达及单核苷酸多态性分析

利用RT-PCR方法,首次由小叶杨受热激胁迫后构建的cDNA文库中扩增获得一热激转录因子HsfAldcDNA克隆,测序结果表明克隆的PsHsfAld cDNA片段总长为2 036 bp,基因内部含有完整的开放阅读框架,大小为1 449 bp,可编码长度为482个氨基酸残基的蛋白质,所推导的蛋白质氨基酸序列在HSF DBD结构功能域与拟南芥AtHsfAld和水稻OsHsfAl蛋白的相似性分别为86.3%和87.4%.组织特异性Realtime-PCR结果显示,Ps HsfAld主要在杨树叶片和根部组织中高丰度表达.非生物胁迫、激素及糖诱导表达表明,PsHsfAld不仅受高温、干旱与盐诱导表达,还受到激素中GA3与ABA,及葡萄糖与蔗糖信号的上调表达.组合利用MEGA4.0和DnaSP4.50.7软件对小叶杨36株基因型个体的PsHsfAld序列进行比对和分析,共检测到207个单核苷酸多态性(SNP)位点,SNP频率为1/16 bp,多样性指数π为0.007 72.在编码区域,共检测到69个SNP位点,其中37个为同义突变,31个为错义突变,1个为无义突变;非同义突变与同义突变的比率0.132＜1,推测在小叶杨物种演化过程中,纯化选择是该基因内同义SNP位点主要的进化驱动力.     

茶树2个MYB转录因子基因的克隆及表达分析

MYB类转录因子是一类包含一段保守的DNA结合结构域的基因家族,广泛地参与植物发育和植物次生代谢的调节。根据前期芯片杂交和文库筛选得到的2个MYB转录因子的部分序列,采用RT-PCR和RACE技术分离得到它们的全长基因:CsMYB1和CsMYB2,在GenBank的登录号分别为HQ660373和HQ660374。序列分析表明:CsMYB1基因全长1132bp,开放阅读框长879bp,编码292个氨基酸,推测的蛋白分子量约为32.9ku,理论等电点为8.13;CsMYB2基因全长1020bp,其中开放阅读框长675bp,编码224个氨基酸,推测的蛋白分子量约为25.4ku,理论等电点为9.05。2个基因编码的蛋白均具有明显的R2R3MYB结构域,且在R3结构域的下游都含有1个相对保守的C1(LIXXGIDPXTHR)基序。同源性分析表明:茶树CsMYB1和CsMYB2编码的氨基酸序列与其他植物的MYB类转录因子具有较高的相似性,其中CsMYB1编码的氨基酸序列与陆地棉MYB1的相似性为57%,CsMYB2编码的氨基酸序列与葡萄MYBC2的相似性为75%。利用荧光定量PCR技术检测2个转录因子基因在遮荫处理条件下的表达规律,及其在茶树不同组织中的表达特性,结果表明:CsMYB1和CsMYB2在不同组织中均有表达,但表达量具有明显区别,其中CsMYB2在叶片中的相对表达量是根中的100多倍;而遮荫处理能明显降低叶片中的花青素含量,并提高CsMYB1的表达,但对转录因子CsMYB2的影响不大。     

荔波连蕊茶肌动蛋白基因全长cDNA的克隆及其表达分析

根据植物肌动蛋白(Actin)基因编码区的保守序列设计引物,以山茶属植物荔波连蕊茶幼嫩茎段为材料,提取总RNA,进行RT-PCR。采用RACE技术扩增获得1 631 bp的Actin基因全长cDNA序列,命名为ClActin1(GenBank登录号KF366912)。序列分析表明,ClActin1开放阅读框(ORF)为1 134 bp,编码377个氨基酸,5’非编码区90 bp,3’非编码区407 bp。预测的ClActin1蛋白分子量为41.69 kD,理论等电点为5.31,具有Actin超基因家族的保守结构域和肌动蛋白家族特有的特征信号序列。ClActin1与GenBank中收录的其它植物肌动蛋白核苷酸序列的相似性在82%以上,氨基酸序列的相似性在97%以上。与其它植物肌动蛋白比较并构建系统进化树,结果显示山茶肌动蛋白与茶树和杨树的肌动蛋白的亲缘关系最为密切。实时荧光定量PCR结果显示,该基因在荔波边蕊茶不同组织器官及不同发育时期的表达量稳定,表明ClActin1基因可作为内参基因。     

油茶乙酰辅酶A羧化酶BC亚基全长cDNA克隆及序列分析

以油茶‘湘林1号’品种的近成熟种子为材料,采用简并PCR、RACE、交错PCR等技术,获得了油茶乙酰辅酶A羧化酶BC亚基基因的全长cDNA克隆。该基因cDNA序列全长1 901 bp,含有一个1 599 bp的ORF,编码533个氨基酸残基。BC蛋白的等电点pI为6.88,分子量为58 509.3 u,含两个跨膜结构域,是一个不稳定的非分泌蛋白。模体搜索结果表明在BC上具有N-糖基化位点、蛋白激酶C磷酸化位点、ATP结合位点等多个功能结构域。同源建模的模型中发现12个α-螺旋,整个BC蛋白为一个内凹的结构。该基因已登录到GenBank,并被命名为co-bc。     

Simultaneous determination of Cu(II), Zn(II), Fe(II), Mn(II), Co(II) and Ni(II) with a single sweep polarography

A new method of simultaneous determining Cu(II), Zn(II), Fe(II), Mn(II), Co(II) and Ni(II) with a single sweep polarography was developed. The experiment condition was studied and the results show that the optimum base solution is that the residue got from evaporating 25 mL mixed solution, 2.5 mL 1.0 mol/L ethylenediamine, 3.0 mL 1.0 mol/L ammonia-ammonium chloride, 5.0 mL 10 % sodium sulfite and 0.5 mL 0.5 % gelatin were mixed and diluted to 25 mL with water. Under the condition of this base solution, the polarography wave shapes of the six cations are ideal and without overlap of the peaks. In this base solution, Cu(II), Zn(II), Fe(II), Mn(II), Co(II) and Ni(II) in synthetic samples were determined with relative errors 0.4–9.4% and relative standard deviations 0.8%-12.2%. Under this experiment condition, one fold of Pb(II), Cr(VI) and Mo(VI) do not interfere with the determination for Cu(II). This method has been used to the simultaneous determination of the amount of the six cations in the samples of soil and Beiqi Tea. The relative standard deviations were 0. l%-2.1% and recoveries were 93.5%-99.0 %.     

大气候室测定家具中甲醛及其他VOC的释放量

采用30m(3)大气候室测定家具中释放的甲醛和其他有机挥发物,并用气相色谱仪和液相色谱仪等分析.结果表明:家具中释放的甲醛和总有机挥发物(TVOC),超过相关标准限量;TVOC中二甲苯、乙酸丁酯和正十一烷相对较多;甲醛为总醛类挥发物的88%.基材甲醛释放量为E2级时,通过贴面并涂饰处理,仍不能阻止甲醛的释放.     

Estimating vertical vegetation density through a SPOT5 imagery at multiple radiometric correction levels

There is growing concern about remote sensing of vertical vegetation density in rapidly expanding peri-urban interfaces.A widely used parameter for such density,i.e.,leaf area index (LAI),was measured in situ in Nanjing,China and then correlated with two vegetation indices (VI) derived from multiple radiometric correction levels of a SPOT5 imagery.The VIs were a normalized difference vegetation index (NDVI) and a ratio vegetation index (RVI),while the four radiometric correction levels were i) post atmospheric correction reflectance (PAC),ii) top of atmosphere reflectance (TOA),iii) satellite radiance (SR) and iv) digital number (DN).A total of 157 LAI-VI relationship models were established.The results showed that LAI is positively correlated with VI (r varies from 0.303 to 0.927,p < 0.001).The R 2 values of "pure" vegetation were generally higher than those of mixed vegetation.The average R 2 values of about 40 models based on DN data (0.688) were higher than that of the routinely used PAC (0.648).Independent variables of the optimal models for different vegetation quadrats included two vegetation indices at three radiometric correction levels,indicating the potential of vegetation indices at multiple radiometric correction levels in LAI inversion.The study demonstrates that taking heterogeneities of vegetation structures and uncertainties of radiometric corrections into account may help full mining of valuable information from remote sensing images,thus improving accuracies of LAI estimation.     

Photosynthesis,growth and yield of soybean and maize in a tree-based agroforestry intercropping system on the Loess Plateau

Agroforestry is the most effective way to restore the disturbed lands on the Loess Plateau and to develop the poor local economy. In order to maximize the potential benefits of tree-based intercropping systems, photosynthesis, growth and yield of soybean and corn were studied by measuring photosynthetic active radiation (PAR), plant water deficit and soil moisture in a 4-year-old plantation of walnut (Juglans regia L.) and plum (Prunus salicina) grown at a spacing of 5 m × 3 m on the Loess Plateau. The effects of tree competition significantly reduced PAR, net assimilation (NA), growth and yield of individual soybean or corn plants growing nearer (1 m near tree row) to tree rows. NA was highly correlated with growth and yield of the both crops. These correlations were higher for corn than soybeans, with corn, rather than soybeans being more adversely impacted by tree shading. Plum, rather than walnut had the greatest competitive effect on PAR and NA. Daily plant water deficits were non-significantly and poorly correlated with NA and growth and yield of the both crops. However, soil moisture (20 cm depth) was significantly correlated with biomass and yield of both crops. Possible remediation strategies are discussed to reduce tree competitive interactions on agricultural crops.     

Topography related spatial distribution of dominant tree species in a tropical seasonal rain forest in China

The degree to which variation in species distribution is predictable from topographic variation is of considerable current interest. In this paper, canonical correspondence analysis (CCA), linear regression and principal coordinates of neighbour matrices (PCNM) models were used to explain the variation in the distributions of the 13 dominant species in a 20-ha tropical rain-forest plot in China. The results showed that: (1) Tree distribution maps show that some species are mainly found in the gullies of the plot, whereas others occur on the slopes. Which indicates topographic variables are important factors for the distribution pattern of species. (2) Both linear regression and CCA results show that convexity and elevation are the most important variables effecting distribution of trees. For saplings, elevation, convexity and aspect explain 15.3%, 9.0% and 10.1% of the total variation of species abundance. For poles, elevation and convexity explain 19.3% and 11.4% respectively. However, only 5.3% of the total variation is explained for adults. (3) The PCNM results showed that topography alone explained 20%, 24% and 5% of the total variation of species abundance for saplings, poles and adults, respectively. Overall evidence for topographic control of the tropical tree distribution is strong, but the explanatory power of topographic variables was a small part of the total of variation.     

Cloning and Sequence Analysis of a Full-length cDNA Encoding Light-harvesting Complexes of Photosystem II in Phyllostachys edulis

The light-harvesting chlorophyll a/b-protein complex plays an important role in photosynthesis of plants. A full-length cDNA of light-harvesting chlorophyll a/b （cab） gene was cloned from the first strand of Moso （Phyllostachys edulis） cDNA through RT-PCR and RACE methods, named as cabPhEIO （cab gene 10 from Ph. edulis）. The length of cab- PhEIO （GenBank accession number： EU118754） is 1 151 bp, which contains an open reading frame encoding 283 amino acids from 81st to 932nd position. The bioinformatics analysis indicated that the protein encoded by cab-PhElO had a chlorophll a/b binding domain （83rd -247th position）, two protein kinase C-phosphorylation sites, three Nmyristoylation sites and a yia A/B double helix domain.The amino acid sequence of cab-PhElO showed high similarity with the cab genes of Oryza sativa, Zea mays, Hordeum vulgare, and Vitis vinifera, more than 80%, respectively, which indicated that cab-PhElO gene belongs to lhcb5 gene family.     

毛白杨转录因子PtCBF5的表达模式分析

利用RT-PCR手段克隆得到1个毛白杨CBF基因cDNA,命名为PtCBF5(GenBank注册:DQ354395)。PtCBF5全长837bp,该序列包括起始密码子ATG和42bp的5′末端非编码区,终止密码子TGA和98bp的3′末端非编码区,开放阅读框编码231个氨基酸。RT-PCR半定量研究表明:PtCBF5在叶片组织中的表达量高,茎、根中的表达量低。组培苗在低温、干旱、NaCl、ABA条件下处理诱导24h,在低温和干旱的条件下PtCBF5表达量在诱导30min后开始增加,分别在2h和6h达到峰值,24h后又恢复到初始水平;而高盐和ABA存在的条件下未有显著变化。PtCBF5在植物体适应寒冷和干旱的过程中可能有着重要作用,尤其在逆境初期。     

应用苏特灵Bt防治黄褐天幕毛虫的试验

该文应用苏特灵Bt生物杀虫剂防治黄褐天幕毛虫Malacosoma neustria，筛选适宜的用药浓度，防治最佳虫龄和时间，为大面积防治提供可靠依据。试验结果表明：600－800倍液，1－2龄幼虫群集网幕中施药，杀虫率95.5%以上。     

沙棘果实生长期间维生素含量变化研究

本文对乌鞘岭地区天然分布的中国沙棘和西藏沙棘果实生长与成熟期间维生素C,PP,E等含量的动态变化进行了系统的调查和分析测定。结果表明:V_C主要存在于果肉中,且随果实发育而增加,果实变色期(花后107—110天)达到最高值,随后急速下降。但完熟后果肉中V_C减少时种子内V_C却有所增加。中国沙棘V_C水平远较西藏沙棘为高(5倍以上)。果实发育前期果肉中V_(PP)含量很高,以后逐渐下降,完熟期中国沙棘果实内V_(PP)再度升高,其变化与呼吸速率的变化表现出相同的消长规律。V_E主要存在于种子里,总趋势是随成熟而增加。种子变色期V_E上升至第一高峰值,随后下降,果实完熟后达到第二高峰值。西藏沙棘V_E含量高于中国沙棘。本文对不同维生素可能起的生理作用也进行了讨论。