A comparison of internal standards for plant cytophotometry

Plant cell nuclei were compared with chicken erythrocyte nuclei for use as internal standards for microspectrophotometry. The amount of DNA per nucleus and the coefficient of variation for measurement of individual nuclei were determined for cells from dormant embryos of Pinus taeda and Pinus coulteri, from onion root tips and from chicken erythrocytes. The chicken erythrocytes had the least variability and thus were best suited for use as a standard. Onion root tips were least suitable, with a coefficient of variation 2 1/2 times that of erythrocytes. Although onion root tips have been used as an internal standard in other studies, their mitotic activity, in contrast with the nonreplication of DNA of mature erythrocytes, is reflected in a broad distribution of nuclei with values in the 2C-4C range. Coulter pine mature embryos were at the 3C level, whether dry or hydrated, while loblolly pine embryos were in the 2C state. This confirms previous reports. The coefficient of variability for the pine embryo cells was 1 1/2 times that of erythrocytes for nonhydrated seeds and twice the erythrocyte value for hydrated seeds. The larger 2C values for pine (26 pg for P. taeda and 17 pg for P. coulteri) are closer to values expected for many plant species than the 3 pg level of the chicken erythrocytes. Dormant P. taeda embryo cells (2C) are suggested as an alternative where the experimental material has large DNA values and/or chicken erythrocytes are difficult to procure. Large sample size is recommended for the plant materials if they are to be used as internal standards in Feulgen cytophotometry.     

马尾松与粘盖乳牛肝菌菌根共生体形成过程

对菌根共生机制的研究是对其进行应用的前提,到目前为止,绝大多数外生菌根（ectomycorrhiza,ECM）的建立过程尚不明晰,在一定程度上限制了这些ECM真菌在林业中的应用。本研究以我国南方地区主栽树种之一——马尾松Pinus massoniana和其林下优势ECM真菌——粘盖乳牛肝菌Suillus bovinus为材料,在无菌条件下研究两者菌根共生体形成过程的形态学特征。结果表明马尾松与粘盖乳牛肝菌的共生过程分为2个阶段：（1）预共生阶段,即物理接触之前,粘盖乳牛肝菌可通过释放挥发物和分泌物促进马尾松根系伸长和分枝;（2）共生阶段,又可分为3个时期。接种后第4天,粘盖乳牛肝菌菌丝体开始与马尾松根系接触并形成附着胞进入接触期;第7天菌丝开始侵入根系内部,侵入期开始;第28天菌套和哈氏网形成,即菌套和哈氏网形成期,该时期菌根化根尖开始膨大,随后继续发育至二叉分枝状菌根形成。在发育顺序方面,哈氏网与菌套同步发育,但哈氏网成形早于菌套。以上结果可对后续ECM共生机制的深入研究及马尾松高效菌根化育苗技术的开发提供参考。     

东北红松纯林菌根外生菌根真菌群落与环境因子的相关性

外生菌根是木本植物根系与真菌形成的共生结构,外生菌根真菌在红松等外生菌根树种的定植与森林生态系统的保持方面起到至关重要的作用。明确菌根系统内外生菌根真菌群落组成是揭示菌根共生机制的前提条件。本研究利用Illumina Hiseq测序平台对生长季内红松纯林内根围土壤及菌根样品ITS2区进行高通量测序,分析其外生菌根真菌群落结构随季节的变化规律,同时通过统计学的方法分析了红松根系微生态中外生菌根真菌群落结构组成变化与其他生物因素、非生物因素的相关性。结果如下：（1）从6月份到10月份,5个月的菌根样品测序共得到741个真菌OTUs,利用FUNGuild数据库分析,其中85个OTUs归类为外生菌根真菌,优势属（相对丰度>5）为蜡壳菌属Sebacina、乳牛肝菌属Suillus、Meliniomyces、红菇属Russula、棉革菌属Tomentella、须腹菌属Rhizopogon和缘腺革菌属Amphinema。6月份菌根中外生菌根真菌的多样性最大,显著高于其他月份。（2）红松林外生菌根真菌群落组成受到土壤pH、有效磷含量、有效钾含量和土壤有效氮含量的影响,它们与外生菌根真菌优势属相对丰度呈现正相关或负相关。（3）根围土壤内真菌是影响红松根系外生菌根真菌相对丰度的另一重要因素,其中,包括普可尼亚属Pochonia、产丝齿菌属Hyphodontia、镰刀菌属Fusarium、Collembolispora、枝穗霉属Clonostachys、Apodus、鹅膏属Amanita在内的土壤真菌与根内外生菌根真菌的相对丰度呈线性关系。同时,超过85%的根内外生菌根真菌与同一取样地的土壤共有,可以认为侵染和扩散是红松根内外生菌根真菌群落形成的主要方式,同时兼有植物根系的选择,因为根内并不包括所有土壤中存在的外生菌根真菌,其机制需要进一步人工模拟试验验证。     

Evidence of natural reciprocal hybridisation between Pinus uliginosa and P. sylvestris in the sympatric population of the species

Peat-bog pine Pinus uliginosa Neumann has become extinct or rare in many parts of Europe. It is supposed to hybridize with P. sylvestris but the influence of hybridization to genetic erosion of the P. uliginosa gene pool is unknown. In the presented study, the crossability between P. uliginosa and P. sylvestris was analyzed in a sympatric population at Węgliniec reserve in Poland. The aim of the study was to prove natural hybridisation and to estimate the influence of this phenomenon on possible natural gene pool erosion of peat-bog pine. A sequence polymorphism in the trnF–trnL cpDNA region of P. uliginosa and P. sylvestris was used to develop the species diagnostic PCR-RFLP marker. The marker of paternally transmitted cpDNA was applied in haplotype analyses of the progeny from open-pollinated P. uliginosa seeds (collected in 2000–2002) and from P. sylvestris ones (in 2002). An inconsistency in species diagnostic cpDNA haplotypes of seedlings and parental trees was observable both for P. uliginosa and P. sylvestris (about 1% and 2% of hybrids seeds, respectively). The results prove the occurrence of reciprocal hybridisation between the species. The influence of hybridisation on the natural gene pool protection and measures to reintroduce peat-bog pine is discussed.     

Nuclear DNA content of Pinus sylvestris (L.) as determined by laser flow cytometry

Nuclear DNA content was determined in nuclei isolated from needles, stems and roots of in vitro grown seedlings and from megagametophytes and embryo of mature seeds in three accessions of Pinus sylvestris L. One accession was from Inari, northern Finland at timber line, and two accessions were from the Alpine region in Italy. Nuclei were mechanically isolated by a chopping method, stained with propidium iodide, and DNA content was determined using an EPICS PROFILE laser flow cytometer. Nuclei isolated from leaves of barley (Hordeum vulgare L. cv. Sultan; 2C=11.12 pg) were used as an internal standard for measurement of pine nuclei. Mean 1C nuclear DNA content of P. sylvestris was 27.88 pg as determined from megagametophyte tissue. Mean 2C value was 52.25 pg as determined from stem and root tissue, and 55.58 pg as determined from embryo tissue. The ratio of 2C to 1C value was 1.87 and 1.99, respectively. Extracts of nuclei from needles contained propidium iodide-absorbing debris which may have interfered with measurements and resulted in lower 2C values than those obtained from stem and root.     

Molecular and paleo-climatic data uncover the impact of an ancient bottleneck on the demographic history and contemporary genetic structure of endangered Pinus uliginosa

With the current rate of biodiversity loss, conservation management practices require a comprehensive understanding of eco-evolutionary relationships, history, and genetic structure of species. Assessments of genetic diversity are crucial, especially in rare, endemic, or threatened forest tree species with small and isolated populations, such as peat bog pine (Pinus uliginosa N.). Here, we used a novel approach, combining genetic diversity assessment, ecological niche modeling, and population demography inference to explore the complex history of a few remnant populations of this endangered pine. To asses the relative influence of isolation and fragmentation on genetic diversity in the taxonomic context, the patterns of genetic variation found in P. uliginosa were contrasted with those observed in its close relatives with much bigger distribution ranges and larger populations (Pinus sylvestris, Pinus mugo, and Pinus uncinata). We found a similar level of genetic diversity across the species at nuclear loci but contrasting patterns of variability distribution at chloroplast markers. We detected the signatures of an ancient genetic bottleneck dated at around 26 400 years ago, indicating a drastic reduction in the population size of P. uligionosa during the Last Glacial Maximum. In addition, we found substantial differentiation between current populations as a result of enhanced genetic drift during long-lasting isolation. The research suggests potential conservation management strategies for peat bog pine and emphasizes the importance of using complementary approaches for their successful development.     

长白松的木材解剖及其分类问题

关于长白松的分类问题一直有争议^[6-11],根据作者对吉林省安图县二道白河产的长白松的木材解剖研究,发现其木材的射线管胞内壁具深锯齿,颇与欧洲赤松(Pinus sylvestris Linn.)相近^{[14] [15]},但以其木射线薄壁组织细胞水平壁无纹孔和晚材最后数列管胞弦壁上存在着排列成轴向单行小形具缘纹孔来看,这又显属赤松(Pinus densiflora Sieb.et Zucc.)的特征。因此,作者认为长白松的木材结构是处于欧洲赤松与赤松之间而更偏于赤松^[10],故确认其当归属于赤松的种群范围。     

供磷水平对梨砧木生长和15N-尿素吸收利用特性的影响

以一年生盆栽豆梨、川梨、木梨实生苗为试材,采用¹⁵N同位素示踪技术,研究了不同供磷水平(P₂O₅分别为0、50、100、150、200 kg·hm^-2,分别用P₀、P₁、P₂、P₃、P₄表示)对3种梨砧木生长和¹⁵N-尿素吸收、利用特性的影响.结果表明: 随着供磷水平的提高,砧木的株高、地径、干质量、根系总表面积、总根长、根尖数、根系活力、根系呼吸速率、Ndff值和氮素利用率均先升高后降低,但不同砧木升降幅度不同,且各指标出现峰值的磷水平不同.在同一磷素水平下,木梨的株高、地径、干质量均最高,川梨次之,豆梨最低,根系形态指标和根系呼吸速率呈相同规律,而Ndff值和氮素利用率表现不同.在不同磷素水平下,木梨在P₃处理各指标均达到最高水平,而川梨和豆梨分别在P₂和P₁处理各指标均达到最高水平;砧木各器官的Ndff值在不同磷水平下均以茎最高;木梨的最高氮素利用率(种植季的肥料N回收率)为9.6%、川梨为8.9%、豆梨为8.3%.以上结果表明,不同梨砧木生长和氮素吸收利用特性在不同磷水平下响应不同,生产中应根据砧木需磷特性合理施磷.     

供磷水平对梨砧木生长和15N-尿素吸收利用特性的影响

以一年生盆栽豆梨、川梨、木梨实生苗为试材,采用¹⁵N同位素示踪技术,研究了不同供磷水平(P₂O₅分别为0、50、100、150、200 kg·hm^-2,分别用P₀、P₁、P₂、P₃、P₄表示)对3种梨砧木生长和¹⁵N-尿素吸收、利用特性的影响.结果表明: 随着供磷水平的提高,砧木的株高、地径、干质量、根系总表面积、总根长、根尖数、根系活力、根系呼吸速率、Ndff值和氮素利用率均先升高后降低,但不同砧木升降幅度不同,且各指标出现峰值的磷水平不同.在同一磷素水平下,木梨的株高、地径、干质量均最高,川梨次之,豆梨最低,根系形态指标和根系呼吸速率呈相同规律,而Ndff值和氮素利用率表现不同.在不同磷素水平下,木梨在P₃处理各指标均达到最高水平,而川梨和豆梨分别在P₂和P₁处理各指标均达到最高水平;砧木各器官的Ndff值在不同磷水平下均以茎最高;木梨的最高氮素利用率(种植季的肥料N回收率)为9.6%、川梨为8.9%、豆梨为8.3%.以上结果表明,不同梨砧木生长和氮素吸收利用特性在不同磷水平下响应不同,生产中应根据砧木需磷特性合理施磷.     

中国东北产“樟子松”的核型研究

作者研究了中国东北所产的"樟子松"的核型。结果表明,大兴安岭山地生长的樟子松与欧洲赤松的核型类似,而沙地生长的樟子松的核型则有明显的变异。根据核型比较研究,作者认为,把山地樟子松视为欧洲赤松的原变种,沙地樟子松应视为另一变种。"樟子松"分布于黑龙江大兴安岭海拔400-900米山地及大兴安岭以西、西南一带沙地。因为它生长迅速,成材期短,能耐贫瘠干旱,为石质山地和草原沙地主要造林树种之一,在我国东北、内蒙、华北地区营林事业中占有重要的地位。多年来,在植物分类学上对于"樟子松"的归属问题还存在着一些分歧:V.L.Komarov(1901)、北川政夫(1939、1979)、竹内亮(1958)等人认为,中国东北产的"樟子松"与分布于欧亚大陆的欧洲赤松(Pinus sylvestris)同属一种;D.I.Litvinov(1905)及陈嵘(1937)、刘慎谔(1955)、吴仲伦(1956),郑万钧等(1961、1978)的研究认为东北 的樟子松均为欧洲赤松的变种(Pinus sylvestris var.mongolica LitV.);近来又有的作者认为中国东北地区的"樟子松"根据产地不同可分为山地樟子松(分布于大兴安岭山地)与沙地樟子松(分布于大兴安岭以西的呼伦尔草原之沙地上)两种类型,并认为前者为欧洲赤松原变种,而后者则为欧洲赤松之另一变种。     

云南松苗木生长与生物量的相关性及通径分析

为了解云南松苗木生长对生物量的影响程度和相对重要性,以采自云南省昆明市宜良县禄丰村林场种子培育的2年生云南松苗木为材料,对苗高、地径、主根长、侧根长、针叶长和侧根数6个生长性状及各组分生物量进行测定,运用相关性分析和逐步回归分析剔除了对生物量影响不大的性状,在此基础上采用通径分析方法研究了云南松苗木生长性状指标与生物量指标间的关系。结果表明：云南松苗木生长性状和生物量之间均呈现极显著的正相关关系。各生长性状对生物量均有直接或间接的正向效应,直接影响云南松苗木生物量的优势因素为地径和苗高,针叶长、侧根长和主根长对生物量的影响也起正向效应。各生长性状对生物量的直接效应均较大,而通过其他性状的间接效应均较小,其中侧根长通过其他性状对生物量产生的间接影响最大。依据相关性分析和通径分析,建立了云南松各组分生物量与生长性状之间的数学模型,可用于云南松苗期生物量的估测。     

样本量对云南松幼苗生物量模型构建及预估精度的影响

为探究不同样本量对生物量模型构建及建模精度的影响，以实际调查的20个家系，共615株云南松幼苗为例，通过编写计算机程序建立进行简单随机抽样，构建不同样本量云南松幼苗各器官及单株生物量异速生长方程。利用决定系数（R²）、估计值标准误（SEE）、均方根误差（RMSE）、总相对误差（RS）及平均误差绝对值（MAB），对模型拟合优度与精度进行比较分析。结果表明：幂函数方程可较好地用于估测云南松幼苗生物量；随着样本量的增加模型精度评估指数MAB呈幂函数形式逐渐减小；当样本数量小于200时MAB较为敏感，模型精度较差，样本量大于200时，其精度随样本量逐渐增加，但变化幅度逐步减小并趋于稳定。因此，根据MAB的变化趋势，样本量达到200时可以构建精度较高且稳定模型。     

Nonspecific light loss and intrinsic DNA variation problems associated with feulgen DNA cytophotometry.

Nonspecific light loss by the cell-wall-plus-cytoplasm (CWC) can cause a 50% increase in Feulgen absorption units in peanut root-tip nuclei as determined by scanning at 450 nm, whereas this phenomenon is not evident with chicken erythrocytes. A two wavelength scanning method of subtracting nonspecific 450 nm absorption from 550 nm Feulgen absorption values eliminated the nonspecific light loss in CWC, However, the two wavelength scanning method is time consuming and somewhat impractical with a regular scanning microdensitometer such as Vickers M85. Elimination of the problem of nonspecific light loss is suggested by careful determination of background setting with the spot position close to the nucleus in CWC. The accuracy of the CWC background setting method was further tested by comparison with subtraction method. The use of plant nucleis as an internal standard in plant DNA measurements was also evaluated. Significant variation among the replicate slides due to the variation in pine nuclear DNA amounts was observed and plant nuclei generally are not reliable internal standards. Mature chicken erythrocytes are recommended as an internal standard because the cell type and metabolic state is known.     

典型黑土区主要树种根系构型特征及其对固土能力的影响

为量化典型黑土区主要树种根系构型特征,探究其对固土能力的影响,以该区分布较广的榆叶梅、小叶锦鸡儿、白桦、糖槭、红皮云杉、樟子松单株个体为研究对象,采用全根挖掘和WinRHIZO Pro LA2004分析系统相结合对其根系空间分布、几何形态、分形等特征进行测定,同时采用原位整株根系拉拔的方法量化根系垂直拉拔力。结果表明: 榆叶梅以倾斜根为主,小叶锦鸡儿、白桦、糖槭和红皮云杉以水平根为主,樟子松根系在水平和垂直分布上较为均衡;除白桦总根表面积和红皮云杉总根长外,灌木树种总根长、总根表面积显著大于乔木,落叶阔叶乔木总根长、总根表面积显著大于针叶常绿乔木,白桦总根体积显著大于小叶锦鸡儿、糖槭、红皮云杉和樟子松;榆叶梅、小叶锦鸡儿和白桦根系分形维数和分形丰度显著大于红皮云杉和樟子松;榆叶梅、小叶锦鸡儿和糖槭整株根系平均最大垂直拉拔力显著大于白桦、樟子松和红皮云杉。主要受根系总根长、总根表面积和倾斜根数量的影响,榆叶梅、小叶锦鸡儿和糖槭根系表现出较强的固土能力,可作为典型黑土区水土保持植被构建中优先选择的树种。     

High DNA content of Sprekelia formosissima Herbert (Amaryllidaceae) and Ophioglossum petiolatum Hook. (Ophioglossaceae)

Ophioglossum petiolatum and Sprekelia formosissima root tips were chemically determined to have 170± 12 pg and 180±12 pg DNA/cell respectively, or 2.8 and 3.0 times the 60±4 pg DNA/ cell of Tradescantia sp. clone 02 root tips. These values were compared with those predicted from nuclear volume measurements. General qualifications of the nuclear volume-DNA content relationship are discussed. Microspectrophotometrically determined relative DNA values for Feulgen stained half-telophase root tip nuclei of O. petiolatum and S. formosissima were 2.8 and 2.6 the value for Tradescantia. The value for Sprekelia is among the highest in the angiosperms, and Ophioglossum probably has the highest nuclear DNA content of ferns. O. petiolatum has 131±3 pg DNA per dormant spore.     

氮添加和水分胁迫对红松、水曲柳幼苗生物量分配的影响

氮沉降和暖干化是我国东北地区面临的主要生态问题,它将对森林生态系统产生怎样的影响一直是生态学研究的热点.本研究以东北温带阔叶红松林中2个关键树种——红松和水曲柳为研究对象,探讨水分胁迫和氮(N)添加对其幼苗短期(55周)生长的影响.结果表明: 红松与水曲柳幼苗生长对N添加和水分胁迫的响应有明显差异.红松对水分胁迫更加敏感,在处理早期(10周)水分胁迫降低了红松幼苗叶生物量,提高了根生物量;N添加只在水分胁迫发生时显著减少了红松根和植株总生物量.水曲柳对N添加的反应更加敏感,氮添加迅速增加了水曲柳茎、根和总生物量;而只有持续的水分胁迫才对水曲柳的茎、根和总生物量有显著影响.红松和水曲柳在持续水分胁迫和N添加处理下,叶、根生物量占比和地上、地下生物量之比都趋于维持一个稳定值,说明两个树种都有很强的自我调节能力.以上结果说明,在未来干旱条件下,红松采取的是“积极”的调整策略,而水曲柳则是“被动”的适应策略,因而相比较而言,红松存活和适应能力可能更强;而在N沉降增加的环境下,水曲柳受益会更大.这些结果可为预测未来东北温带森林群落演替动态提供参考.     

冠层光谱组成对红松和蒙古栎幼苗生长和光合荧光特性的影响

本研究选择长白山阔叶红松林优势树种红松和蒙古栎幼苗为对象,研究其植株形态、生长和光合荧光特性对5种光谱处理的响应。结果表明: 红松与蒙古栎的形态结构与生长主要受蓝光与紫外B区(UV-B)辐射调控。滤除蓝光后两种幼苗的植株叶面积比和相对生长速率均显著降低,而滤除UV-B辐射显著增加了红松的叶面积比和相对生长速率,分别上升了41.8%和47.7%,降低了蒙古栎的株高、总叶面积和生物量积累。滤除UV-B辐射显著降低了2种幼苗的光合荧光调节能力,红松的下降幅度较低,其非调节性耗散的量子产量(Φ_NO)升高31.6%,反映光合荧光调节能力的Φ_NPQ/Φ_NO值降低37.5%。2个树种幼苗具有明显不同的光谱适应策略,蒙古栎幼苗偏向于利用光谱变化调整自身形态增加光捕获能力,而红松更注重调整光合荧光特征以提高碳同化效率。     

褐环乳牛肝菌对樟子松和油松根际土壤真菌多样性的影响

为了解褐环乳牛肝菌Suillus luteus对樟子松Pinus sylvestris var. mongolica和油松Pinus tabulaeformis根际土壤真菌群落结构的影响,采用Illumina MiSeq高通量测序,对两种松树接种S. luteus与对照组的根际土壤真菌群落结构进行研究。结果显示,4个样品共获得原始序列347 681条,归为5个真菌门。综合各土壤样品真菌Alpha多样性指数及OTUs-Venn图,发现接种S. luteus后樟子松和油松根际土壤真菌相对丰度与对照存在一定差异。群落结构分析表明,接种S. luteus提高了樟子松和油松根际土壤担子菌和壶菌的相对丰度,抑制了子囊菌门真菌。优势属由原来的Geopora转变为Suillus。通过PCA分析与NMDS分析发现接种S. luteus后樟子松和油松根际土壤真菌群落差异性增加,且接种S. luteus后樟子松根际土壤真菌群落结构相似性显著高于油松根际土壤真菌群落结构相似性。接种S. luteus的樟子松和油松土壤真菌丰富度降低、多样性增加,接种S. luteus改变了樟子松和油松根际土壤真菌群落结构和优势真菌种群。试验结果为樟子松和油松促生真菌菌种筛选和育苗工作提供了依据。     

亚热带红壤侵蚀区马尾松不同套种模式生态系统碳平衡

以福建省长汀县红壤侵蚀区马尾松低效林套种杨梅、无患子、油茶及黄栀子的改造模式林分为研究对象,对林分各组分生物量年净生长量、含碳率及土壤异养呼吸进行定位观测,分析套种模式对低效马尾松林分生态系统碳储量格局及碳平衡的影响。结果表明: 杨梅、无患子、油茶、黄栀子和马尾松不同器官含碳率的变化范围分别为41.1%～50.1%、42.2%～50.6%、45.1%～48.9%、44.7%～49.6%和46.1%～51.9%。不同树种同一器官之间的含碳率存在显著差异。马尾松套种杨梅及马尾松套种无患子模式植被层碳储量及年净增碳储量最高,分别为67.62～68.42 t·hm^-2和9.21～9.45 t·hm^-2·a^-1,马尾松套种油茶和马尾松套种黄栀子模式较小,分别为31.96～36.24 t·hm^-2和4.09～4.16 t·hm^-2·a^-1,马尾松纯林对照最小,分别为17.01 t·hm^-2和2.00 t·hm^-2·a^-1。土壤异养呼吸年通量从高到低依次为马尾松套种杨梅模式(7.41 t·hm^-2·a^-1)>马尾松套种油茶模式(5.89 t·hm^-2·a^-1)>马尾松套种无患子模式(5.86 t·hm^-2·a^-1)>马尾松套种黄栀子模式(4.95 t·hm^-2·a^-1)>马尾松纯林对照(2.45 t·hm^-2·a^-1)。马尾松套种杨梅和马尾松套种无患子模式的年净生态系统碳平衡分别为2.04和3.27 t C·hm^-2·a^-1,表现为“碳汇”,马尾松套种油茶和马尾松套种黄栀子模式及马尾松纯林对照的年净生态系统碳平衡分别为-1.80、-0.80和-0.45 t C·hm^-2·a^-1,表现为“碳源”。总体上,短期内马尾松低效林套种杨梅或无患子能够提升林分的固碳增汇效益。