鸡Ii结合Rab5a和Rab7b分子的分析

本研究旨在探明鸡恒定链（invariant chain，Ii）与内吞体转运蛋白Rab5a和Rab7b结合的结构域和在细胞内共定位的特征。首先，用PCR和基因突变技术将Ii胞浆区与跨膜区[Ii_{（Cyt-Tra）}]、Ii CLIP （class Ⅱ-associated invariant chain peptide）-三聚体区[Ii_{（CLIP-TRIM）}]和Ii突变体[Ii_{（M81-87aa）}、Ii_{（M91-99aa）}和Ii_{（M81-99aa）}]分别插入pET-32a和pEGFP-C1构建相应的原核和真核重组质粒。其次，将构建的含有绿色荧光蛋白的重组质粒与实验室保存的含有红色荧光Rab5a和Rab7b的重组质粒共转染至人胚胎肾细胞系293 T，观察它们的共定位。将构建的原核重组质粒进行表达和纯化，最后用拉下法和免疫印迹检测Ii与Rab5a和Rab7b的结合域。结果表明，成功构建Ii结构域及Ii突变体的重组质粒。Ii_{（Cyt-Tra）}及Ii突变体均能与Rab5a和Rab7b在细胞内共定位，而Ii_{（CLIP-TRIM）}与空载体却不能。Ii的胞浆区和跨膜区是与Rab5a和Rab7b结合的功能结构域，而不是CLIP与三聚体区。综上所述，鸡Ii与Rab5a和Rab7b共定位和结合的区域是其胞浆区和跨膜区，而不是内质网腔区。这些结果提示Rab分子参与了Ii在胞内细胞器的转运机制，为进一步研究Ii及其载体在细胞内的转运机制和功能提供了新的途径。     

凤阳山不同林分土壤腐殖质特征及影响因素

以浙江省凤阳山海拔1 300~1 400 m处不同林分类型(阔叶混交林、针阔混交林、杉木林、竹林)为对象,测定不同土层土壤基本理化性质、酶活性及腐殖质质量分数,分析土壤腐殖质特征及影响因素,为凤阳山自然保护区的土壤肥力和可持续经营发展提供理论依据。结果表明:土壤腐殖质质量分数针阔混交林最高,阔叶混交林、竹林、杉木林次之;胡敏酸质量分数针阔混交林最高,杉木林最低;富里酸质量分数针阔混交林最高,竹林最低。除磷酸酶活性随土层加深无统一规律外,4种林分脲酶活性、过氧化氢酶活性及蔗糖酶活性土壤随土层加深皆呈现降低趋势。土壤pH值、土壤密度与土壤腐殖质质量分数、胡敏酸质量分数和富里酸质量分数均呈现显著的负相关;除磷酸酶活性相关性不显著外,土壤脲酶活性、蔗糖酶活性、过氧化氢酶活性、总孔隙度、毛管孔隙、非毛管孔隙与土壤腐殖质质量分数、胡敏酸质量分数和富里酸质量分数均呈现显著的正相关。凤阳山不同林分类型对土壤腐殖质特征的影响较土层深度更显著,土壤理化性质及酶活性与土壤腐殖质质量分数有着密切关系。     

Expression of Anti-Lipopolysaccharide Factor Isoform 3 in Chlamydomonas reinhardtii Showing High Antimicrobial Activity

Antimicrobial peptides are a class of proteins with antibacterial functions. In this study, the anti-lipopolysaccharide factor isoform 3 gene (ALFPm3), encoding an antimicrobial peptide from Penaeus monodon with a super activity was expressed in Chlamydomonas reinhardtii, which would develop a microalga strain that can be used for the antimicrobial peptide production. To construct the expression cluster, namely pH2A-Pm3, the codon optimized ALFPm3 gene was fused with the ble reporter by 2A peptide and inserted into pH124 vector. The glass-bead method was performed to transform pH2A-Pm3 into C. reinhardtii CC-849. In addition to 8 μg/mL zeocin resistance selection, the C. reinhardtii transformants were further confirmed by genomic PCR and RT-PCR. Western blot analysis showed that the C. reinhardtii-derived ALFPm3 (cALFPm3) was successfully expressed in C. reinhardtii transformants and accounted for 0.35% of the total soluble protein (TSP). Furthermore, the results of antibacterial assay revealed that the cALFPm3 could significantly inhibit the growth of a variety of bacteria, including both Gram-negative bacteria and Gram-positive bacteria at a concentration of 0.77 μM. Especially, the inhibition could last longer than 24 h, which performed better than ampicillin. Hence, this study successfully developed a transgenic C. reinhardtii strain, which can produce the active ALFPm3 driven from P. monodon, providing a potential strategy to use C. reinhardtii as the cell factory to produce antimicrobial peptides.     

施氮水平对7S亚基缺失大豆根系形态和结瘤固氮的影响

为有效推广功能型大豆7S亚基缺失品种,以7S亚基缺失大豆品系东富2号为研究对象,设置4种施氮水平(纯N),N0(0 mg·kg~(-1))、N1(25 mg·kg~(-1))、N2(50 mg·kg~(-1))、N3(75 mg·kg~(-1)),采用桶栽法研究大豆根系形态和结瘤固氮对不同施氮水平的响应。结果表明:N1(25 mg·kg~(-1))水平下根系干重加大,根冠比增大,根瘤固氮潜力高,单株产量较高。N2(50 mg·kg~(-1))水平下根长、根表面积、根体积在生育后期增长较快,根系干重较大,根冠比低,固氮酶活性最高,单株籽粒产量最高。N3(75 mg·kg~(-1))水平下植株干重较大,无效生长较多,根瘤数少,固氮潜力和根冠比低,单株产量不高。综合籽粒产量和根系特性指标,功能型大豆7S亚基缺失品系东富2号的适宜施肥量为25~50 mg·kg~(-1)。     

Neuroprotective Metabolites from Vietnamese Marine Derived Fungi of Aspergillus and Penicillium Genera

Low molecular weight secondary metabolites of marine fungi Aspergillus flocculosus, Aspergillus terreus and Penicillium sp. from Van Phong and Nha Trang Bays (Vietnam) were studied and a number of polyketides, bis-indole quinones and terpenoids were isolated. The structures of the isolated compounds were determined by 1D and 2D NMR and HR-ESI-MS techniques. Stereochemistry of some compounds was established based on ECD data. A chemical structure of asterriquinone F (6) was thoroughly described for the first time. Anthraquinone (13) was firstly obtained from a natural source. Neuroprotective influences of the isolated compounds against 6-OHDA, paraquat and rotenone toxicity were investigated. 4-Hydroxyscytalone (1), 4-hydroxy-6-dehydroxyscytalone (2) and demethylcitreoviranol (3) have shown significant increasing of paraquat- and rotenone-treated Neuro-2a cell viability and anti-ROS activity.     

青霉菌发酵液对大豆幼苗生长及生理特性的影响

采用室内生理检测及生物化学方法,研究了青霉菌发酵液对大豆幼苗生长及生理特性的影响。结果表明:用一定浓度的青霉菌发酵液对大豆浸种和茎叶处理,大豆幼苗的生长呈明显上升趋势,处理后大豆的株高、根长、株鲜重和根鲜重均明显得到了提高,且生长状态好于对照,不同浓度处理间具有明显差异,在发酵液浓度为5 g·L-1时,效果最佳。尤其是对根鲜重的影响,促进率最高可达56.21%,同时增强了幼苗根系活力,最大可以提高57.82%,浸种和茎叶处理后,分别提高了叶绿素含量41.25%和15.40%;超氧化物歧化酶活性12.20%和11.70%;过氧化物酶31.33%和29.95%;过氧化氢酶活性42.8%和20.0%,这为发酵液在大豆田推广应用、提高大豆产量提供了重要理论依据。     

miR-140-3p inhibits viability,invasion and migration of non-small-cell lung cancer A549 cells by targeting PD-L1

AIM: To explore the target relationship between microRNA-140-3p (miR-140-3p) and programmed cell death ligand 1 (PD-L1) and their effect on the viability, migration and invasion of non-small-cell lung cancer A549 cells.METHODS: RT-qPCR was used to detect the miR-140-3p expression in HLF-1, A549 and H1299 cells, and then the A549 cells with the most significant difference were selected as the subsequent research object. TargetScan software and dual-luciferase reporter assay were performed to predict and confirm the target relationship between miR-140-3p and PD-L1. RT-qPCR and Western blot were used to determine the effects of miR-140-3p mimic and inhibitor on PD-L1 expression level. MTT assay was used to detect the viability of A549 cells. Transwell assay was performed to detect the migration and invasion abilities of the A549 cells.RESULTS: miR-140-3p was significantly down-regulated in the A549 cells and H1299 cells (P<0.05). Transfection with miR-140-3p mimic decreased the expression of PD-L1 and inhibited the viability, migration and invasion of the A549 cells. Transfection with pcDNA3.0-PD-L1 reversed the inhibitory effect of miR-140-3p on the viability, migration and invasion of the A549 cells.CONCLUSION: miR-140-3p inhibits the viability, migration and invasion of A549 cells by targeting PD-L1.