龙葵抗阿特拉津生物型叶绿体基因文库的建立

用对阿特拉津(Atrazine)除草剂抗性的龙葵生物型B_(12)株系作材料,制备叶绿体DNA。B_(12)株ctDNA(叶绿体DNA)经BamHI酶解,在0.7％琼脂糖凝胶电泳上呈现24条带,其中最大的片段为18.6kb,最小的片段为1kb。用pBR322作为载体,构建B_(12)株ctDNA BamHI片段文库。通过与探针的分子杂交,从中筛选出含有编码叶绿体32kd蛋白质的阿特拉津抗性基因的克隆pSB135和含有ATP合酶α亚单位基因的克隆pSB132。     

番木瓜果糖-2,6-二磷酸酶CpF2KP基因克隆和蛋白表达纯化

番木瓜是岭南四大名果之一,在我国东南部地区广泛种植,因其具有食用和药用双重价值,因此深受人们的青睐。果糖-6-磷酸,2-激酶/果糖-2,6-二磷酸酯酶(fructose-6-phosphate,2-kinase/fructose-2,6-bisphosphatase,F2KP)是一个独特的双功能酶,具有激酶功能域和酯酶功能域,能催化生物体内糖代谢的重要调节物果糖-2,6-二磷酸(Fru-2,6-P_(2))的合成和降解。为了研究番木瓜中编码该酶的基因CpF2KP的功能,得到目的蛋白尤为重要。本研究从番木瓜基因组中提取到CpF2KP基因的编码序列(coding sequence,CDS)序列,该基因CDS全长2274 bp。将该基因CDS全长扩增之后选用pGEX-4T-1载体进行原核表达。对载体pGEX-4T-1用EcoRⅠ和BamHⅠ进行双酶切,利用基因重组的方式将扩增序列构建到原核表达载体上。经过诱导条件探索,SDS-PAGE结果显示GST-CpF2KP重组蛋白的大小约为110 kDa,诱导CpF2KP蛋白表达的最适条件为:异丙基β-D-硫代半乳糖苷(isopropyl beta-D-thiogalactopyranoside,IPTG)浓度为0.5 mmol/L,温度28℃。对诱导后的CpF2KP蛋白进行纯化,得到了纯化的单一目的蛋白。此外,检测了该基因的组织表达特性发现该基因在种子中表达量最高,在果肉中表达量最低。该研究为进一步深入揭示番木瓜CpF2KP蛋白的功能及研究该基因参与的生物学过程提供了重要基础。     

西双版纳国家级自然保护区蚂蚁-树互作网络空间变异

网络分析(network analysis)可以同时分析群落中的物种多样性和种间关系, 为了解生态群落的稳定性机制提供了新的分析思路和方法。本研究从西双版纳国家级自然保护区的纳板河、勐仑和勐腊(补蚌)三个地点采集了树栖性蚂蚁及树木的种类和数量数据, 对蚂蚁-树组成的二分网络进行了分析, 探讨了3个采样点物种的多样性、网络指标以及群落指标之间的关系。我们采用零模型的方法比较了3个样点的标准化网络参数差异。结果表明: 蚂蚁和树木的物种数以及树的异质性指数(Shannon-Wiener多样性指数、Simpson多样性指数)都呈现出勐仑 > 纳板河 > 补蚌的趋势。树木-蚂蚁的灭绝曲线系数大小关系同样为勐仑 > 纳板河 > 补蚌, 灭绝曲线与树的物种数及异质性指数大小趋势一致, 而与蚂蚁的异质性指数并不吻合。根据Z值的绝对值来看, 网络参数(加权嵌套性、平均连接数、特化水平、模块性、连接度)与群落参数(灭绝曲线系数、生态位重叠)的大小趋势相同, 表现出勐仑 > 纳板河 > 补蚌的趋势。综上所述, 蚂蚁-树互作网络的稳定性(灭绝曲线系数)主要由树的数量和异质性指数决定。网络的加权嵌套性和网络中节点的平均连接数也能促进群落的稳定性。而在一个特化的(数值越大表示专性互作越多)和模块化(具有较多密切互作的节点单元)的网络中, 当低营养级物种灭绝时高营养级物种数量将迅速减少。     

荒漠结皮层藓类植物死亡对表层土壤水分蒸发和入渗的影响

水分是荒漠植物生长最主要的限制因子,藓类结皮作为荒漠土壤表层重要覆被物,对土壤水分蒸发入渗具有重要影响。研究表明,在全球气候变化背景下,不确定的降水格局变化导致结皮层藓类植物出现集群死亡现象,但这一过程对荒漠地表土壤水分蒸发与入渗过程的影响及其机理尚不清楚。以古尔班通古特沙漠齿肋赤藓结皮为研究对象,利用便携式渗透计和蒸发仪,研究了结皮层藓类植物死亡对土壤水分蒸发与入渗的影响。结果表明,与裸沙相比,藓类结皮的存在显著抑制了水分入渗,而藓类植物死亡的结皮层抑制作用最大,其初渗速率、稳渗速率和累积入渗量分别是活藓类结皮的39.89%、85.91%及64.48%,仅为裸沙的5.96%、13.13%及20.42%。在水分蒸发初期,裸沙的水分蒸发速率明显高于活藓类结皮和藓类植物死亡的结皮层,但藓类植物死亡的结皮层维持相对稳定的蒸发速率的时间长于裸沙和活藓类结皮,这也导致最终累计蒸发量以藓类植物死亡的结皮层最高、裸沙最低。可见,荒漠生物土壤结皮中藓类植物死亡会明显减少土壤水分入渗、增大水分蒸发,进一步影响荒漠表层土壤水分格局,从而影响生物土壤结皮与维管植物的水分利用关系。     

Comparison of the Efficiency of Banna Miniature Inbred Pig Somatic Cell Nuclear Transfer among Different Donor Cells

Somatic cell nuclear transfer (SCNT) is an important method of breeding quality varieties, expanding groups, and preserving endangered species. However, the viability of SCNT embryos is poor, and the cloned rate of animal production is low in pig. This study aims to investigate the gene function and establish a disease model of Banna miniature inbred pig. SCNT with donor cells derived from fetal, newborn, and adult fibroblasts was performed, and the cloning efficiencies among the donor cells were compared. The results showed that the cleavage and blastocyst formation rates did not significantly differ between the reconstructed embryos derived from the fetal (74.3% and 27.4%) and newborn (76.4% and 21.8%) fibroblasts of the Banna miniature inbred pig (P>0.05). However, both fetal and newborn fibroblast groups showed significantly higher rates than the adult fibroblast group (61.9% and 13.0%; P<0.05). The pregnancy rates of the recipients in the fetal and newborn fibroblast groups (60% and 80%, respectively) were higher than those in the adult fibroblast group. Eight, three, and one cloned piglet were obtained from reconstructed embryos of the fetal, newborn, and adult fibroblasts, respectively. Microsatellite analyses results indicated that the genotypes of all cloning piglets were identical to their donor cells and that the genetic homozygosity of the Banna miniature inbred pig was higher than those of the recipients. Therefore, the offspring was successfully cloned using the fetal, newborn, and adult fibroblasts of Banna miniature inbred pig as donor cells.     

Determinants of 14-3-3σ Protein Dimerization and Function in Drug and Radiation Resistance

Many proteins exist and function as homodimers. Understanding the detailed mechanism driving the homodimerization is important and will impact future studies targeting the “undruggable” oncogenic protein dimers. In this study, we used 14-3-3σ as a model homodimeric protein and performed a systematic investigation of the potential roles of amino acid residues in the interface for homodimerization. Unlike other members of the conserved 14-3-3 protein family, 14-3-3σ prefers to form a homodimer with two subareas in the dimeric interface that has 180° symmetry. We found that both subareas of the dimeric interface are required to maintain full dimerization activity. Although the interfacial hydrophobic core residues Leu¹² and Tyr⁸⁴ play important roles in 14-3-3σ dimerization, the non-core residue Phe²⁵ appears to be more important in controlling 14-3-3σ dimerization activity. Interestingly, a similar non-core residue (Val⁸¹) is less important than Phe²⁵ in contributing to 14-3-3σ dimerization. Furthermore, dissociating dimeric 14-3-3σ into monomers by mutating the Leu¹², Phe²⁵, or Tyr⁸⁴ dimerization residue individually diminished the function of 14-3-3σ in resisting drug-induced apoptosis and in arresting cells at G₂/M phase in response to DNA-damaging treatment. Thus, dimerization appears to be required for the function of 14-3-3σ.     

Niche Partition of Bacteriovorax Operational Taxonomic Units Along Salinity and Temporal Gradients in the Chesapeake Bay Reveals Distinct Estuarine Strains

The predatory Bacteriovorax are Gram-negative bacteria ubiquitous in saltwater systems that prey upon other Gram-negative bacteria in a similar manner to the related genus Bdellovibrio. Among the phylogenetically defined clusters of Bacteriovorax, cluster V has only been isolated from estuaries suggesting that it may be a distinct estuarine phylotype. To assess this hypothesis, the spatial and temporal distribution of cluster V and other Bacteriovorax phylogenetic assemblages along the salinity gradient of Chesapeake Bay were determined. Cluster V was expected to be found in significantly greater numbers in low to moderate salinity waters compared to high salinity areas. The analyses of water and sediment samples from sites in the bay revealed cluster V to be present at the lower salinity and not high salinity sites, consistent with it being an estuarine phylotype. Cluster IV had a similar distribution pattern and may also be specifically adapted to estuaries. While the distribution of clusters V and IV were similar for salinity, they were distinct on temperature gradients, being found in cooler and in warmer temperatures, respectively. The differentiation of phylotype populations along the salinity and temporal gradients in Chesapeake Bay revealed distinct niches inhabited by different phylotypes of Bacteriovorax and unique estuarine phylotypes.