低温对2种紫草科植物△6 脂肪酸脱氢酶基因表达的影响

以紫草科植物新疆软紫草(Arnebia euchroma)和假狼紫草(Nonea capsica(willd.)G.Don)为材料,翻译延伸因子EF1a为内参基因,利用半定量逆转录聚合酶链式反应(semi-RT-PCR)技术,对2种紫草植物中γ-亚麻酸合成关键酶基因△6-脂肪酸脱氢酶基因的表达进行研究。结果表明,△6-脂肪酸脱氢酶基因在紫草植物的叶、茎、根中均表达,在低温诱导不同时间后紫草植物叶片中△6-脂肪酸脱氢酶基因的表达有所增强。气相色谱分析显示,其产物γ-亚麻酸在低温下积累,推测紫草科植物叶片中△6-脂肪酸脱氢酶基因表达量增加可能与适应低温胁迫的逆境有关。     

猪ERK6基因编码区的克隆及组织表达谱分析

利用RT-PCR方法扩增了猪ERK6基因编码区的序列，将扩增产物与pMD18-T载体连接，构建了重组质粒；重组质粒经PCR、酶切鉴定后进行测序；采用Northern杂交和半定量RT-PCR方法在猪不同部位骨骼肌中分析了ERK6基因转录本的个数、大小及组织表达谱。结果显示，所克隆的猪ERK6基因片段长1113bp，含有1个1104bp的开放阅读框（ORF），该ORF编码367个氨基酸；该基因与已报道的人ERK6基因核苷酸序列相似性为90％；猪ERK6基因在肌肉组织中只有一个转录本，大小约1．5kb。此基因在骨骼肌中的表达量最高，心、子宫次之，卵巢最低；而在脂肪、胃、肝、脾、肺、肾、十二指肠和胰腺中未见表达。结果表明，猪ERK6基因与已报道的小鼠和人ERK6基因一样，主要在骨骼中特异表达。     

Succession and phylogenetic profile of eukaryotic communities in rice straw incorporated into a rice field: Estimation by PCR-DGGE and sequence analyses

Abstract

Succession and the phylogenetic profile of eukaryotic communities associated with rice straw decomposition in a rice field were studied using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis followed by 18S rDNA sequencing. Nylon mesh bags containing leaf sheaths or blades were buried in the plow layer of a rice field under flooded conditions after transplanting (Experiment 1) and under drained conditions during the off-crop season (Experiment 2). In addition, rice straw samples in Experiment 2 were taken out before plowing in spring and re-placed in the rice field under flooded conditions at transplanting. Statistical analyses based on DGGE patterns showed that eukaryotic communities were divided into two groups, namely group A before the placement in soil, after the mid-season drainage in Experiment 1 and under the drained conditions in Experiment 2 and group B before the mid-season drainage in Experiment 1 and under the flooded conditions in Experiment 2. Based on the sequence analysis of DGGE bands, which characterized the eukaryotic communities, succession of the communities was revealed, that is, most of the bands in group A were closely related to fungi, whereas the bands in group B were closely related to protozoa. These results indicated that eukaryotic communities associated with rice straw decomposition in the rice field are mainly affected by soil conditions, such as oxic or reduced conditions, irrespective of rice straw parts (leaf sheaths and blades).     

饲粮调控对猪胴体脂肪酸的组成改善及氧化稳定性的增强效应

从猪胴体脂肪酸组成特点及其改变的可能性入手 ,系统综述了饲粮中添加不同种类与水平的油脂、全脂油籽与饼粕对猪胴体脂肪酸组成的改善 ,以及维生素E、硒、维生素C、类胡萝卜素与肌肽对不饱和脂肪酸含量较高的胴体脂肪氧化稳定性的增强效应 ,并指出了今后的研究和发展方向。     

鸡软骨细胞体外分离培养鉴定以及过表达miR-15a对软骨细胞的影响

为分析miR-15a在肉鸡不同组织中的表达情况,并探究过表达miR-15a对体外培养鸡软骨细胞的影响。本研究首先通过倒置显微镜观察、PCR、凝胶电泳和甲苯胺蓝染色对体外分离培养的软骨细胞进行鉴定。通过实时荧光定量PCR检测miR-15a在肢体内外翻畸形(valgus-varus deformity, VVD)组和健康组肉鸡中(各3只)各组织的表达量。之后通过CCK8和EDU方法分析软骨细胞过表达miR-15a后细胞增殖情况。软骨细胞转染miR-15a mimics后,通过qPCR检测软骨细胞的标志基因Collagen-2、Aggrecan、Collagen-10,成熟分化基因Runx2、Sox9、VEGF、MMP9,炎性因子IL-1β、IL-6、IL-8、IL-10、TNF-α、TGF-β3以及凋亡基因Fas、FasL、Bcl-2的表达量。并构建FKBP5 3'UTR的野生型载体和突变型载体,通过双荧光素酶检测报告检测miR-15a与FKBP5的靶向关系。结果表明,本研究所用的胰蛋白酶、Ⅱ型胶原酶和透明质酸酶联合消化法成功分离得到状态良好的软骨细胞。荧光定量结果显示,miR-15a在各组织中均有表达,与健康组相比,miR-15a在VVD组的肝(P < 0.01)、脾(P < 0.05)、胸腺(P < 0.01)中的表达量显著升高,在心和胸肌组织中的表达量显著降低(P < 0.01)。CCK8与EDU分析结果显示,与NC组相比,过表达miR-15a组软骨细胞增殖速度显著降低(P < 0.01),增殖细胞数量显著减少(P < 0.01)。qPCR结果显示,与mimics NC组相比,miR-15a mimics组的软骨细胞标志基因Aggrecan、成熟分化基因Sox9、Runx2表达量显著降低(P < 0.05),Fas基因表达量极显著上升(P < 0.01),FasL基因和抗凋亡基因Bcl-2极显著下降(P < 0.01)。成功构建了FKBP5 3'UTR野生型和突变型载体,双荧光素酶检测报告结果显示预测的靶基因FKBP5与miR-15a没有靶向关系。本研究成功分离并鉴定了鸡软骨细胞,过表达miR-15a抑制鸡软骨细胞增殖、成熟和分化并促进细胞凋亡。     

Assessment of Rhizospheric Microorganisms of Transgenic Populus tomentosa with Cowpea Trypsin Inhibitor (CpTI) Gene

To have a preliminary insight into biosafety of genetically transformed hybrid triploid poplars (Populus tomentosa × P. bolleana) × P. tomentosa with the cowpea trypsin inhibitor (CpTI) gene, two layers of rhizospheric soil (from 0 to 20 cm deep and from 20 to 40 cm deep, respectively) were collected for microorganism culture, counting assay and PCR analysis to assess the poten-tial impact of transgenic poplars on non-target microorganism population and transgene dispersal. When the same soil layer of suspension stock solution was diluted at both 1:1 000 and 1:10 000 rates, there were no significant differences in bacterium colony numbers between the inoculation plates of both transgenic and non-transgenic poplars. The uniform results were revealed for both soil layer suspension solutions of identical poplars at both dilution rates except for non-transgenic poplars at 1:10 000 dilution rates from the same type of soil. No significant variation in morphology of both Gram-positive and Gram-negative bacteria was observed under the microscope. The potential transgene dispersal from root exudates or fallen leaves to non-target microbes was repudiated by PCR analysis, in which no CpTI gene specific DNA band was amplified for 15 sites of transgenic rhizospheric soil samples. It can be concluded that transgenic poplar with the CpTI gene has no severe impact on rhizospheric microorganisms and is tentatively safe to surrounding soil micro-ecosystem.     

The method for designing the profile of forest roads supported by genetic algorithm

This study examines a method for designing an optimum profile of a forest road in a short matter of time. In this method, a genetic algorithm (GA) searches for points where the gradient changes and dynamic programing (DP) designs the longitudinal slope. This method can be applied to the ground heights of an actual forest road. The evaluation function of the DP assesses the total expenditure. The inverse of this function is the fitness function. Crossover and mutation are operators of an evolution simulation of a GA. They are each repeated 20 times and 420 profiles are designed and assessed, all within about 10 min. A comparison of the profile designed by the GA with that of an actual forest road shows only one section where the formation level differed substantially. Overall, both profiles seem to show a high degree of correspondence. From this result, we believe that the profile designed by the GA is best, or the one of the best. A conventional method takes about 65 h to calculate all changing points of gradients. However, by using the GA, the calculation time was reduced to about 1/389 that of the coventional method, showing that the method which uses the GA through the profile design is very useful.     

Effects of polyline simplification of dynamic GPS data under forest canopy on area and perimeter estimations

The objectives of this study were to determine Global Positioning System (GPS) positional errors while moving under the forest canopy and to clarify the effects of polyline simplification on area and perimeter estimations. We used the Pathfinder Pro XR and GPSMAP 76S, which are categorized as “high-end mapping” and “general navigation” GPS receivers, respectively. The field tests were conducted in both natural and plantation forests. The results showed that the Pathfinder Pro XR, which has better multipath rejection technology, worked well, especially in the plantation forest under unfavorable conditions of higher stand density. We used analysis of variance to clarify the effects of the receiver type, positioning mode, stand type, and polyline simplification method on area and perimeter estimations. The receiver type and positioning mode were found to be significant factors that affected area estimation. The Pathfinder Pro XR estimated the area more accurately than the GPSMAP 76S, and differential GPS estimated the area more accurately than autonomous GPS. With respect to the perimeter, the receiver type, positioning mode, and polyline simplification method were found to be significant factors. The results showed that perimeter estimation was improved by using the velocity filter, and further improved by using the velocity filter and Douglas-Peucker algorithm, especially when the Pathfinder Pro XR was used. The GPSMAP 76S estimated the perimeter accurately without any filtering because its default speed filter worked well, even though the GPSMAP 76S is a general navigation GPS receiver.     

The effects of within-species and between-species variation in wood density on the photodegradation depth profiles of sugi (Cryptomeria japonica) and hinoki (Chamaecyparis obtusa)

Low density wood is more rapidly eroded than denser wood when exposed to the weather, possibly because it is more susceptible to photodegradation. Fourier transform infrared microscopy was used to examine: (1) the depth of photodegradation in earlywood and latewood of sugi (Japanese cedar) and earlywood of hinoki (Japanese cypress) exposed for up to 1500 h to artificial sunlight emitted by a xenon lamp (375 W/m² within the 300 to 700 nm spectral range); and (2) the relationship between the density of wood tissues and depth of photodegradation. The depth of photodegradation varied between species (sugi and hinoki) as well as within a growth ring (sugi earlywood and latewood), and there was an inversely proportional relationship between depth of photodegradation and wood density. These findings may explain why low density earlywood is more rapidly eroded than latewood during weathering, and more generally, why there is an inverse relationship between the density of wood species and their rate of erosion during artificial and natural weathering. Part of this work was presented at the 54th Annual Meeting of the Japan Wood Research Society, Sapporo, August 2004