GFP基因在新疆小拟南芥和拟南芥中的表达分析

以质粒pMCB30为模板,扩增GFP基因,连接到载体pCMBIA2300-35S-OCS上,构建过量表达载体p35S:GFP,将其转入农杆菌GV3101.通过农杆菌介导法将p35S:GFP载体分别转入新疆特色植物小拟南芥和拟南芥中.T0代经含有卡那霉素的1/2MS培养基筛选,获得了T1代转基因小拟南芥2株,T1代转基因拟南芥9株.通过激光共聚焦显微镜观察,在转基因小拟南芥和拟南芥的根尖细胞中均可检测到GFP绿色荧光蛋白;对转基因植株进行PCR扩增,均可检测到GFP基因,表明GFP基因已成功转入小拟南芥和拟南芥中.该研究建立了小拟南芥的遗传转化体系,为进一步利用GFP基因和进一步研究小拟南芥的功能基因奠定基础.     

Intrinsic Effects of Gold Nanoparticles on Oxygen–Glucose Deprivation/Reperfusion Injury in Rat Cortical Neurons

This study aimed to investigate the potential effects of gold nanoparticles (Au-NPs) on rat cortical neurons exposed to oxygen–glucose deprivation/reperfusion (OGD/R) and to elucidate the corresponding mechanisms. Primary rat cortical neurons were exposed to OGD/R, which is commonly used in vitro to mimic ischemic injury, and then treated with 5- or 20-nm Au-NPs. We then evaluated cell viability, apoptosis, oxidative stress, and mitochondrial respiration in these neurons. We found that 20-nm Au-NPs increased cell viability, alleviated neuronal apoptosis and oxidative stress, and improved mitochondrial respiration after OGD/R injury, while opposite effects were observed for 5-nm Au-NPs. In terms of the underlying mechanisms, we found that Au-NPs could regulate Akt signaling. Taken together, these results show that 20-nm Au-NPs can protect primary cortical neurons against OGD/R injury, possibly by decreasing apoptosis and oxidative stress, while activating Akt signaling and mitochondrial pathways. Our results suggest that Au-NPs may be potential therapeutic agents for ischemic stroke.

     

An Escherichia coli MutY mutant without the six-helix barrel domain is a dimer in solution and assembles cooperatively into multisubunit complexes with DNA

Escherichia coli MutY is an adenine and weak guanine DNA glycosylase involved in reducing the mutagenic effects of 7,8-dihydro-8-oxoguanine (GO). MutY contains three structural domains: an iron-sulfur module, a six-helix barrel module with the helix-hairpin-helix motif, and a C-terminal domain. Here, we demonstrate that the mutant MutY(Delta26-134), which lacks the six-helix barrel domain, cannot complement the mutator phenotype of a mutY mutant in vivo. However, the mutant can still bind DNA and has weak catalytic activity at high enzyme concentrations. The mutant is a dimer in solution and assembled into two and multiple (up to five) complexes with 20- and 44-bp DNA fragments, respectively, in a concentration-dependent manner. Higher order complexes with DNA substrates containing A/GO mismatches were formed at lower protein concentrations than with the A/G mismatch and homoduplex DNA. Measurement of equilibrium binding using fluorescence anisotropy showed that the mutant protein retains some specificity for A/GO-containing DNA substrates and that the binding event is highly cooperative. This is consistent with the MutY structure determined, which indicates that GO specificity is contributed by both the six-helix barrel and C-terminal domains. The nonspecific binding of MutY(Delta26-134) to DNA suggests a model in which the specific binding of mismatched DNA by MutY involves sequential interactions, in which one MutY molecule scans the DNA and enhances binding of another MutY molecule to the A/GO mismatch.     

Genome sequencing and comparison of two nonhuman primate animal models, the cynomolgus and Chinese rhesus macaques

The nonhuman primates most commonly used in medical research are from the genus Macaca. To better understand the genetic differences between these animal models, we present high-quality draft genome sequences from two macaque species, the cynomolgus/crab-eating macaque and the Chinese rhesus macaque. Comparison with the previously sequenced Indian rhesus macaque reveals that all three macaques maintain abundant genetic heterogeneity, including millions of single-nucleotide substitutions and many insertions, deletions and gross chromosomal rearrangements. By assessing genetic regions with reduced variability, we identify genes in each macaque species that may have experienced positive selection. Genetic divergence patterns suggest that the cynomolgus macaque genome has been shaped by introgression after hybridization with the Chinese rhesus macaque. Macaque genes display a high degree of sequence similarity with human disease gene orthologs and drug targets. However, we identify several putatively dysfunctional genetic differences between the three macaque species, which may explain functional differences between them previously observed in clinical studies.     

Effects of domestic wastewater slow infiltration on the growth of poplar 'Zhonglin 2001' plantation

A slow infiltration experiment with different hydraulic loads (0, 3, 6, 9, 12, and 15 cm per week) of domestic wastewater was conducted in a 'Zhonglin 2001' poplar plantation to study the effects of the wastewater slow infiltration on the growth of the plantation. Comparing with the control (0 cm), the other five treatments increased the soil organic matter, total N, total P, total K, and Na+ contents in the plantation averagely by 1.940 g x kg(-1), 0.115 g x kg(-1), 0.029 g x kg(-1), 1.454 g x kg(-1) and 0.030 g x kg(-1), respectively. At lower hydraulic loads (3-12 cm per week), the poplar biomass growth and the N, P and Na+ contents in different poplar organs averagely increased by 17.583 t x hm(-2) x a(-1), 3.086 g x kg(-1), 0.645 g x kg(-1), and 0.121 g x kg(-1), with the maximum (36.252 t x hm(-2) x a(-1), 13.162 g x kg(-1), 5.137 g x kg(-1), and 0.361 g x kg(-1), respectively) at hydraulic loads 6-12 cm per week. The further increase of the hydraulic load decreased the poplar biomass growth and the N, P and Na+ contents in different poplar organs. The K content in different poplar organs decreased with increasing hydraulic load. Treating with domestic wastewater increased the leaf length, decreased the leaf asymmetry, and delayed leaf-falling. At high hydraulic load (15 cm per week), the higher soil Na+ and water contents would threat the poplar growth. The proper domestic wastewater hydraulic loads for the growth of poplar 'Zhonglin 2001' plantation would be 3-12 cm per week.     

Comparative Genomic Analysis of N2-Fixing and Non-N2-Fixing Paenibacillus spp.: Organization,Evolution and Expression of the Nitrogen Fixation Genes

We provide here a comparative genome analysis of 31 strains within the genus Paenibacillus including 11 new genomic sequences of N₂-fixing strains. The heterogeneity of the 31 genomes (15 N₂-fixing and 16 non-N₂-fixing Paenibacillus strains) was reflected in the large size of the shell genome, which makes up approximately 65.2% of the genes in pan genome. Large numbers of transposable elements might be related to the heterogeneity. We discovered that a minimal and compact nif cluster comprising nine genes nifB, nifH, nifD, nifK, nifE, nifN, nifX, hesA and nifV encoding Mo-nitrogenase is conserved in the 15 N₂-fixing strains. The nif cluster is under control of a σ⁷⁰-depedent promoter and possesses a GlnR/TnrA-binding site in the promoter. Suf system encoding [Fe–S] cluster is highly conserved in N₂-fixing and non-N₂-fixing strains. Furthermore, we demonstrate that the nif cluster enabled Escherichia coli JM109 to fix nitrogen. Phylogeny of the concatenated NifHDK sequences indicates that Paenibacillus and Frankia are sister groups. Phylogeny of the concatenated 275 single-copy core genes suggests that the ancestral Paenibacillus did not fix nitrogen. The N₂-fixing Paenibacillus strains were generated by acquiring the nif cluster via horizontal gene transfer (HGT) from a source related to Frankia. During the history of evolution, the nif cluster was lost, producing some non-N₂-fixing strains, and vnf encoding V-nitrogenase or anf encoding Fe-nitrogenase was acquired, causing further diversification of some strains. In addition, some N₂-fixing strains have additional nif and nif-like genes which may result from gene duplications. The evolution of nitrogen fixation in Paenibacillus involves a mix of gain, loss, HGT and duplication of nif/anf/vnf genes. This study not only reveals the organization and distribution of nitrogen fixation genes in Paenibacillus, but also provides insight into the complex evolutionary history of nitrogen fixation.     

林木分布格局多样性测度方法: 以阔叶红松林为例

林木分布格局是森林结构的重要组成部分, 直接影响森林生态系统的健康与稳定, 维持森林结构多样性被认为是保护生物多样性的最佳途径。本研究探讨了林木分布格局多样性的测度方法, 以期为揭示森林结构多样性提供理论依据。格局多样性研究的关键在于选择合适的生物多样性测度方法和具有分布属性的格局指数。本研究通过统计角尺度分布频率和Voronoi多边形边数分布频率, 运用Simpson指数分别计算角尺度多样性和Voronoi多边形边数分布多样性, 作为表达林木分布格局多样性指数的方法, 并以我国东北吉林蛟河的3个100 m × 100 m的阔叶红松(Pinus koreansis)林长期定位监测标准地为例, 分析了林木分布格局的多样性。结果表明: 无论是角尺度分布还是Voronoi多边形的边数分布都接近正态分布, 角尺度分布中随机分布林木的频数最多, 占55%以上; Voronoi多边形的类型多达10个以上, 50%以上的林木有5-6株最近相邻木。利用Simpson指数衡量林木格局多样性, 角尺度分布与Voronoi多边形的边数分布都显示出聚集分布的林分比随机分布林分的格局多样性高。研究还发现, 两种格局判定方法得出的Simpson指数值有所不同, 角尺度分布的多样性数值明显低于Voronoi多边形的边数分布的多样性数值, 主要原因是二者的等级数量不同。可见, 林木分布格局多样性研究应选择具有分布属性的格局指数, 但由于各指数反映的角度不同, 所以在分析比较不同林分格局多样性时应采用相同的分析方法。     

拟南芥根毛衰老死亡过程的PCD检测

根毛是植物体吸收养分的重要器官, 自然条件下根毛的寿命很短, 仅能存活2–3周, 随即脱落死亡。以模式植物拟南芥(Arabidopsis thaliana)根毛为材料, 对根毛死亡的细胞学特征进行了报道。结果发现, 根毛衰老死亡后细胞内的原生质体发生了收缩, 并在胞质中观察到凝集物的出现; 通过原位末端标记(TUNEL)检测, 发现幼根上的根毛细胞核DNA发生了片段化。上述结果表明, 拟南芥根毛的衰老死亡很可能是植物体自主调控的程序性细胞死亡(PCD)。另外, 当根毛衰老死亡后,细胞核大多会迁移到靠近根毛基部的位置, 且正常的长管状根毛发生旋转扭曲。