铁皮石斛花总RNA提取方法的比较研究

为筛选铁皮石斛(Dendrobiumofficinale)花总RNA提取方法,对8种提取方法进行了比较研究,包括改良CTAB-LiCl法(M1)、改良CTAB-异丙醇法(M2)、改良SDS-LiCl法(M3)、改良SDS-异丙醇法(M4)、多糖多酚植物RNA提取试剂盒法(M5)、柱式植物RNAout 2.0试剂盒法(M6)、RNAprep Pure多糖多酚植物总RNA提取试剂盒法(M7)和Biospin多糖多酚植物总RNA提取试剂盒法(M8)。结果表明,以M4和M5提取的总RNA带型清晰,完整性好,A260 nm/A280 nm为1.8～2.0,A260 nm/A230 nm大于2.0,RNA产率分别为(159.45±1.45)和(170.84±3.53)μg/g。利用M4、M5提取霍山石斛、金钗石斛、鼓槌石斛和美花石斛花的总RNA,样品的完整性、浓度和纯度均符合质量要求。以M4、M5提取的铁皮石斛总RNA为模板,扩增Actin基因片段,扩增产物大小与预期一致且条带单一。这说明M4、M5方法操作简便,结果重复性好,能够较好地提取石斛属植物花的总RNA。     

灰绿藜RNA提取方法及在cDNA文库构建中的应用

目的:为分离灰绿藜耐盐相关新基因,构建叶片cDNA文库,探索从盐生植物中提取高质量RNA的方法.方法:以灰绿藜叶片为材料,应用RNA Plant Reagent法(Tiangen)、Plant RNA purification Reagent法(Invitrogen)、UNIQ柱式总RNA试剂盒提取法(Sangon)、SDS-LiCl法、TRIZOL试剂法(Invitrogen)等进行总RNA的提取,并构建其全长cDNA文库和抑制消减文库.结果:RNAPlant-Reagent法、Plant RNA purification Reagent法和UNIQ柱式总RNA试剂盒提取法获得的RNA完整性均不好;SDS-LiCl法提取的RNA完整性好(28S条带亮度是18S的2倍、条带锐利清晰)、纯度高(A280/A260为1.89±0.03,A260/A230为2.23±0.02),适合用于直接反转录及构建全长cDNA文库,并获得成功;TRIZOL试剂法简单快捷,产率高(2724±82μg/g),完整性较好,可经mRNA纯化后用于抑制消减文库的构建并获得成功.结论:建立了用于构建高质量cDNA文库的灰绿藜RNA的制备方法.     

大青杨叶片总RNA的快速提取方法

目的:寻求快速提取大青杨叶片总RNA的方法。方法:分别用改良CTAB法、改良SDS法、改良TRIzol法及某公司总RNA提取试剂盒提取大青杨叶片总RNA,并用紫外光谱分析、凝胶电泳方法对提取的总RNA进行鉴定。结果:用改良CTAB法和改良TRIzol法能有效地去除蛋白及多糖,提取到的总RNA纯度高,D260nm/D280nm分别为2.05和1.78,RNA的完整程度优于试剂盒法。结论:改良CTAB法为大青杨叶片总RNA的最佳提取方法。     

从富含多糖和多酚的柿果中提取具转录活性RNA的方法

通过吸光比值分析 ,反转录实验比较TRIzoL法、Tris 硼酸法、异硫氰酸胍法、市售经典总RNA抽提试剂盒、FlashUNIQ 10柱离心总RNA抽提试剂盒 ,以及改进的CTAB法提取成熟期柿果中的总RNA的可行性 ,结果表明 :改进的CTAB法效果较好 ,柿果中RNA的吸光比值A2 60 A2 3 0 在 1.5～ 2 .0之间 ,A2 60 A2 80 在 1.7～ 1.9之间。采用设定的锚定引物Oligo(dT) 1 2 A分别对提取物进行反转录 ,并结合随机引物进行PCR扩增 ,琼脂糖凝胶电泳 ,结果仅有改进的CTAB法提取的成熟期柿果中的总RNA具有转录活性     

滨海沙地人工林树种水分利用效率和叶片养分浓度比较及其关系分析

以福建省福州市滨海后沿沙地人工营造的湿地松、木麻黄、尾巨桉、肯氏相思和纹荚相思人工林为研究对象,运用稳定碳同位素法测定植物水分利用效率,探索叶片氮、磷养分状况与水分利用效率之间的差异及相互关系。结果表明:(1)湿地松和肯氏相思的叶片δ13 C均与尾巨桉差异显著(P0.05),但湿地松、木麻黄、肯氏相思、纹荚相思之间的叶片δ13 C均无显著差异;肯氏相思、纹荚相思的叶片δ15 N均显著高于湿地松、木麻黄和尾巨桉。(2)不同树种水分利用效率大小依次为:湿地松肯氏相思木麻黄纹荚相思尾巨桉,且湿地松叶片的WUE显著高于尾巨桉36.87%,而与其他树种的水分利用效率均无显著差异。(3)各树种叶片氮浓度大小依次为:肯氏相思纹荚相思尾巨桉木麻黄湿地松,但仅肯氏相思和纹荚相思与湿地松之间差异达到显著水平;各树种叶片磷浓度大小依次为:尾巨桉肯氏相思纹荚相思湿地松木麻黄,其中尾巨桉与湿地松、木麻黄、纹荚相思之间差异显著;木麻黄和纹荚相思人工林叶片的N/P显著高于尾巨桉人工林,其余树种之间的叶片N/P均无显著差异。(4)叶片δ13 C与叶片N浓度呈显著二次曲线相关关系,叶片δ15 N与叶片N浓度呈极显著线性正相关关系(P0.01)。研究发现,滨海沙地不同树种人工林间水分利用效率、叶片氮浓度、磷浓度、N/P均存在显著差异,水分利用效率与叶片氮浓度、磷浓度均呈显著负相关关系,氮、磷含量是影响滨海沙地沿海防护林树种水分利用效率的重要因子。     

红树植物叶片总RNA提取方法比较与优化

分别采用三种植物RNA提取试剂盒、改良的CTAB-LiCl法和CTAB-异丙醇法五种方法提取四种红树植物叶片总RNA,并用紫外光谱、琼脂糖凝胶电泳、cDNA合成和real-time PCR等手段对提取效果进行鉴定.反复试验表明:五种方法对白骨壤Aricennia marina叶片总RNA提取均有良好的效果.Tiangen RNA提取试剂能够成功提取秋茄Kandelia candel、桐花树Aegiceras corniculatum、白骨壤的总RNA,提取的木榄Bruguiera gymnorrhiza总RNA浓度低、杂质较多,且稳定性较差,在对该方法改良后则能够成功提取木榄的总RNA.Invitrogen试剂盒对木榄和秋茄叶片均有良好的提取效果,但是对桐花树提取的RNA易受到蛋白的污染.CTAB-LiCl法和CTAB-异丙醇法提取的RNA总产量偏低,提取时间较长,且实时定量结果表明,两种方法反转录效率较试剂盒方法也偏低.     

广西高峰林场相思树的真菌病害调查

相思树是从澳大利亚引进的速生丰产林种,在广西以高峰林场为主要种植基地。经初步调查,不同相思树上发生的真菌性病害不同,马占相思树、厚荚相思树、直干相思树、纹英相思树、杂交相思发生炭疽病,其病原茵为胶孢炭疽菌(Colletotrichum gloeosporioides),在马占相思、直干相思、纹荚相思和杂交相思的叶片上只产生病原菌的无性阶段,而在厚荚相思叶片上可同时出现无性阶段和有性阶段;马占相思树和杂交相思树上发生的白粉病菌为白粉菌Oidium sp．,其中杂交相思白粉病发生普遍而严重,发病期多在3-5月;黑木相思树发生叶尖枯病,病原为异色拟盘多毛孢(Pestalotiopsis versicolor),一年四季均可发病;马占相思树干基部心腐病发生严重,造成木材的大量损失,病原尚未得到确定。本文对病害的症状特点、病原菌形态特征进行了描述。     

五种不同组织结构特点昆虫的总RNA提取效率比较

【目的】比较RNAeasy抽提试剂盒法和Trizol提取法对不同种昆虫总RNA提取的效率,为不同种昆虫总RNA提取方法选择提供参考。【方法】采用RNAeasy抽提试剂盒法和Trizol提取法分别对九香虫Aspongopus chinensis Dallas、白背飞虱Sogatella furcifera Horváth、中华蜜蜂Apis cerana cerana、黄蜻Pantala flavescens和蒿金叶甲Chrysolina aurichalcea进行总RNA提取,通过紫外分光光度计测定总RNA吸光度(OD)值,利用琼脂糖凝胶电泳检测等方式对RNA质量进行评估。【结果】2种提取方法提取不同昆虫总RNA中,Trizol法提取白背飞虱(同翅目)、蒿金叶甲(鞘翅目)和九香虫(半翅目)的总RNA浓度较高,但纯度较低。电泳结果显示总RNA有降解,完整性较差,RNAeasy抽提试剂盒法提取黄蜻(蜻蜓目)和中华蜜蜂(膜翅目)的浓度较低,纯度较高,电泳结果显示完整性较好。【结论】Trizol法更适合提取小型或几丁质含量高的昆虫,RNA抽提试剂盒法更适合几丁质含量较少、腹软且有大量体液的昆虫总RNA提取。     

快速提取棉花蕾期高质量RNA的改良方法

棉花组织中因富含棉酚、多糖和单宁等次生代谢物,RNA难分离,易降解,导致现有RNA快速提取试剂盒提取的棉花RNA产率低、质量差,无法应用于实验操作。针对棉花组织次生代谢物多的特点,以酸酚法为基础,结合RNA吸附柱,通过对幼苗期和蕾期棉花叶片RNA提取,总结出一种快速提取棉花组织RNA的方法。与热硼酸法、酸酚法及试剂盒法相比,所述方法具有步骤少、产率高、质量好等特点,操作简单、整个实验在1h内完成,可开发出提取多糖多酚类生物组织RNA试剂盒。     

Acinetobacter sp. Y1的氨氮去除性能及其关键酶活性

【目的】研究Acinetobacter sp.Y1的氨氮(NH_4~+-N)去除性能及其关键酶的提取与酶活性。【方法】以柠檬酸钠为碳源,硫酸铵为氮源,研究菌株Y1的NH_4~+-N去除性能;采用正交实验优化超声波破碎法提取粗酶的条件,SDS-PAGE分析比较渗透压休克法和超声波破碎法获得的粗酶;检测关键酶——羟胺氧化还原酶(HAO)、亚硝酸盐还原酶(NIR)、硝酸盐还原酶(NAR)的酶活性。【结果】24 h内菌株Y1的菌密度(OD600)可达1.280,对NH_4~+-N、总氮(TN)和COD的降解率分别达到98%、94%和92%,硝化过程中羟胺、亚硝酸盐氮、硝酸盐氮不积累,反硝化产生N2;超声波破碎法提取粗酶的最佳工作条件为:破碎功率50 W,工作与间歇时间分别为4 s和7 s,OD600为1.250,总工作时间20 min,关键酶HAO、NIR和NAR的比活力分别为0.011、0.002和0.018 U/mg;渗透压休克法得到的HAO比活力是0.067 U/mg。【结论】Acinetobacter sp.Y1能同时高效去除NH_4~+-N、TN和COD。优化超声波破碎法提取粗酶的条件,检测到HAO、NIR和NAR的酶活性,且渗透压休克法比超声波破碎法更适合用来提取HAO。     

Three-dimensional xylem networks and phyllode properties of co-occurring Acacia

Reduced leaf size is often correlated to increased aridity, where smaller leaves demand less water via xylem conduits. However, it is unknown if differences in three-dimensional (3D) xylem connectivity reflect leaf-level adaptations. We used X-ray microtomography (micro-CT) to quantify 3D xylem connectivity in ～5 mm diameter branch sections of co-occurring semi-arid Acacia species of varied phyllode size. We compared 3D connectivity to minimum branch water potential and two-dimensional (2D) vessel attributes derived from sections produced by micro-CT. 2D attributes included vessel area, density, vessel size to number ratio (S) and vessel lumen fraction (F). Trees with terete phyllodes had less negative water potentials than broad phyllode variants. 3D xylem connectivity was conserved across all trees regardless of phyllode type or minimum water potential. We also found that xylem connectivity was sensitive to vessel lumen fraction (F) and not the size to number ratio (S) even though F was consistent among species and phyllode variants. Our results demonstrate that differences in phyllode anatomy, and not xylem connectivity, likely explain diversity of drought tolerance among closely related Acacia species. Further analysis using our approach across a broader range of species will improve understanding of adaptations in the xylem networks of arid zone species.     

华南地区桉树/相思混交种植的林木生长效应

将桉树U6与厚荚相思按不同比例和不同模式进行混交种植,测定二者的生长参数.结果表明：2～3年生混交林中,随着桉树比例的增大,相思免受风害的效果越来越显著,桉树/相思混交比例为3:1时,风害率下降了26.14%;但不同处理间桉树或相思保存率没有显著差异;桉树/相思混交种植对相思生长有一定抑制作用,混交林中相思胸径约为其纯林的90%,混交种植虽对桉树高生长影响不明显,但对胸径生长有显著促进作用,并随相思比例的增加而越发明显.6年生的桉树/相思1:1混交林分,桉树胸径>15 cm所占比例达到了32.1%,而桉树纯林仅为5.83%;桉树/相思2∶1株间混交的种植模式使总生物量达到最大,为198.8 m³·hm^-2,是桉树纯林的118.8%,相思纯林的169.9%;桉树与相思混交种植对大径材桉树的培育是一种理想的种植模式.     

马占相思的日光合作用和日碳固定总量研究 (英文)

 马占相思（Acacia mangium）是一种具有较高经济价值和生态效益的热带速生树种。为了研究其光合作用对一天内光强、温度等环境因子变化的响应,揭示其迅速生长的生理生态机制,在叶片水平上测量了马占相思在24 h内的净光合速率(Pn)、蒸腾速率、气孔导度(Gs)、胞间CO2浓度(Ci)、每日固定碳的总量、夜间消耗碳的总量以及日间光合系统Ⅱ光化学效率(Fv/Fm)的变化。结果显示：其净光合速率在正午降低了14.5%;其较高的蒸腾速率和气孔导度有助于保持叶片日间较高的碳固定速率;光合系统Ⅱ对于高光强和高温有较强的忍受度;每日固定碳的总量为502 mmol CO2·m-2,夜间碳的损失为54 mmol CO2·m-2。研究结果在一定程度上揭示了马占相思在热带高光强和高温条件下迅速生长的生理机制。     

Hydraulic conductance of Acacia phyllodes (foliage) is driven by primary nerve (vein) conductance and density

We determined effects of venation traits on hydraulic conductance of phyllodes (foliage), using an array of Acacia s.str. species with diverse phyllode morphologies as the source of variation. Measurements were made on phyllodes from 44 species, grown in common gardens but originating from different positions along a precipitation gradient. K(phyllode) varied 18-fold and was positively correlated with primary nerve hydraulic conductance, and with primary nerve (vein) density but not with minor nerve density, in contrast with previous studies of true leaves in other dicotyledons. Phyllodes with higher primary nerve density also had greater mass per area (PMA) and larger bundle sheath extensions (BSEs) from their minor nerves. We suggest that higher primary nerve conductivity and density may decrease the distance travelled in the high-resistance extra-xylem pathways of the phyllode. Further, larger BSEs may increase the area available for dispersion of water from the xylem to the extra-xylem tissue. High PMA phyllodes were more common in acacias from areas receiving lower annual precipitation. Maximizing efficient water movement through phyllodes may be more important where rainfall is meagre and infrequent, explaining relationships between nerve patterns and the climates of origin in Australian phyllodinous Acacia.     

乳腺叶状囊肉瘤超声征象与病理对照分析

目的：探讨乳腺叶状囊肉瘤超声影像学特征及与病理组织学的相关性。方法：回顾性分析经手术病理证实的15例乳腺叶状囊肉瘤的超声影像学表现,并与病理诊断结果进行对照,分析肿块大小、形态、内部回声、后方回声增强、有无钙化,内部血流、与周围组织有无粘连等影像学表现与病理组织学的相关性。结果：15例乳腺叶状囊肉瘤其病理组织分化呈现多样性,其中恶性分级呈低度恶性的有7例,中度恶性的有5例,高度恶性的有3例。15例乳腺叶状囊肉瘤经超声诊断为乳腺叶状囊肉瘤9例,乳腺癌4例,纤维腺瘤2例,诊断的符合率为60%。乳腺叶状囊肉瘤有多样种病理类型,超声声像图上以不均匀中低回声表现为主,主要表现为肿块大,形态分叶状;边缘清楚,多见囊性变,少见或罕见钙化,血流信号丰富。结论：乳腺叶状囊肉瘤的超声表现具有比较典型的影像特征,与其病理类型相关,超声检查对乳腺叶状囊腺瘤具有较高的诊断及临床应用价值。     

几种提取白木香茎干总RNA方法的比较

目的:通过比较多种RNA提取方法,确定白木香茎干RNA提取的有效手段,为白木香伤害形成沉香药材的分子研究奠定基础。方法:取2年生白木香茎,分别用CTAB法、TRIzol法、异硫氰酸胍-SDS法、Qiagen试剂盒和Nor gen试剂盒法提取总RNA,通过琼脂糖凝胶电泳和紫外分光光度法测定其产量与质量,并通过cDNA合成及小样本克隆测序检查RNA的可用性。结果:异硫氰酸胍-SDS法及Norgen试剂盒法提出的RNA条带清晰,完整性好;D260nm/D280nm值可达2.0左右,纯度较高,其产量足够应用于cDNA合成;其他方法不能提出RNA。结论:异硫氰酸胍-SDS法及Norgen试剂盒法可用于白木香茎干RNA提取。     

Trizol RNA提取法在雪貂组织的流感病毒H3N2定量PCR检测中优于RNeasy mini kit（Qiagen）柱子法

目的比较RNeasy mini kit（Qiagen）柱子法和Trizol法提取RNA在雪貂肺和肠组织H3N2 SYBRGreen PCR定量中的去干扰作用,从而找到适合该定量体系的RNA提取方法。方法比较Trizol法,RNeasy minikit柱子法,改良RNeasy mini kit柱子法1和改良RNeasy mini kit柱子法2提取肺肠组织RNA的质量,RNA逆转录成cDNA后,利用SYBR Green PCR方法检测样品中H3N2的载量,比较产物的特异性,综合评价RNA提取方法。结果 4种方法提取的肺和肠组织的RNA质量不相同,紫外分光光度仪测得RNeasy mini kit柱子法提取的RNA的A260/280低于1.8,其余3种方法的A 260/280在1.8～2.1之间,A 260/230只有Trizol法能达到2.0左右,其余3种方法均远低于2.0。电泳可见Trizol法提取的RNA有3条带：28 s、18 s和5 s,而RNeasy mini kit柱子法的RNA除了有28 s、18 s外,还有基因组DNA,改良RNeasy mini kit柱子法1和2提取的RNA只有28 s和18 s带。RNA逆转录后进行荧光定量PCR,从溶解曲线看,不论是鼻甲刮取物还是肺肠组织的样品模板,4种方法获得的样品与标准品均为单一峰溶解曲线峰,并且波峰位置重叠。而鼻甲刮取物定量的产物跑琼脂糖凝胶电泳均为单一的特异性条带,而肺肠组织的产物电泳发现：Trizol法获得的模板定量产物与标准品一致,为单一的特异性条带,而其它3种方法获得的模板则均有非特异性条带。结论鼻甲刮取物的病毒定量选用RNeasy mini kit柱子法提取定量结果与Trizol法一样可靠,说明对于简单样品该体系及引物非常适用,结果可信。对于组织样品肺和肠,Trizol法获得的样品定量结果比其它3种方法可靠。     

Epidemiology and control of virus diseases of vegetables

In order to assess the effect of an introduced gall wasp on the weed Acacia longifolia, it was necessary to study the vegetative growth and reproductive potential of uninfested trees. The growth of branches on uninfested trees was studied over one year. The results yielded a growth phenology which is notably different from the growth phenology of A. longifolia described by Milton (1980). Milton (1980) observed no flushing, but a gradual increase and decline in growth during spring and late summer, respectively. The present study showed distinct flushing, and a marked lack of growth during spring. Examination of the effect of reproduction on the vegetative growth of A. longifolia explains the disparity. A. longifolia reproduces during spring. Comparison of phyllode addition during spring with pod production for three successive years at one site, and for two years at another, shows that phyllode addition is low during years of high pod production, and vice versa. Between trees, phyllode addition is also inversely related to pod production within a reproductive season. Because the present growth phenology was done in a year of high pod production, growth during spring was negligible, in contrast with Milton's (1980) phenology which was done in a year of low pod production and high phyllode addition. Depending on the year in which the plant was studied, its reproductive potential and vegetative growth could have been under- or overestimated by as much as 62% and 90%, respectively. This study shows that the phenology of weeds with an annual cycle needs to be examined for at least two years in order not to under- or overestimate their reproductive or growth potential. This is especially important when assessing the efficacy of biological control agents.