Molecular cloning and expression analyses of a new gene encoding 3-hydroxy-3-methylglutaryl-CoA synthase from Taxus × media

A new full-length cDNA encoding 3-hydroxy-3-methylglutaryl-CoA synthase (designated as TmHMGS, GenBank Accession No. AY644708), which catalyses the condensation of acetyl CoA and acetoacetyl CoA to form 3-hydroxy-3-methylglutaryl-CoA as an early step in the taxol biosynthetic pathway, was isolated from young leaves of Taxus × media by rapid amplification of cDNA ends (RACE) for the first time. The full-length cDNA of TmHMGS contained a 1431 bp open reading frame (ORF) encoding a deduced protein of 476 amino acid residues. The deduced protein had an isoelectric point of 5.23 and a calculated molecular mass of about 53 kDa. Amino acid sequence comparison analysis showed that TmHMGS had high similarity with a number of HMGSs ranging from Schizosaccharomyces pombe to humans, with much higher identity with other HMGSs from plants than those from yeast and humans. Phylogenic analysis showed that TmHMGS had closest relationship with HMGS from Pinus sylvestris. Tissue expression pattern analysis showed that TmHMGS expressed in needles and stems at similar level, but no expression could be detected in roots. Expression of TmHMGS was all induced by under different elicitors such as silver nitrate, ammonium ceric sulphate and methyl jasmonate, revealed that TmHMGS was an elicitor-responsive gene.     

芝麻愈伤组织诱导与植株再生体系的建立

以芝麻栽培种(Sesamum indicum, 2n=26)、野生种(S. radiatum, 2n=64; S. schinzianum, 2n=64)及其远源杂交后代(S. schinzianum × S. indicum)为材料, 研究了不同基因型、外植体类型、激素种类及其浓度对芝麻愈伤组织诱导及植株再生的影响, 建立了芝麻愈伤组织诱导及高频植株再生的技术体系。结果表明, 6-BA/NAA激素组合有利于绿色紧密型愈伤组织的形成及分化; 最佳愈伤组织诱导及分化培养基为MS+ 0.1 mg·L^–1NAA + 2.0 mg·L^–16-BA+ 30 g·L^–1蔗糖。在该培养条件下, 野生种下胚轴愈伤组织的诱导率最高为97.50%, 分化率为94.02%; 栽培种下胚轴愈伤组织的诱导率最高为40.60%, 分化率为8.16%; 远缘杂交后代幼胚外植体愈伤组织的诱导率最高为46.67%, 分化率为89.29%。该研究结果为芝麻转基因技术体系的建立及新种质创制奠定了基础。     

ARK2 stabilizes the plus-end of microtubules and promotes microtubule bundling in Arabidopsis

Microtubule dynamics and organization are important for plant cell morphogenesis and development. The microtubule-based motor protein kinesins are mainly responsible for the transport of some organelles and vesicles, although several have also been shown to regulate microtubule organization. The ARMADILLO REPEAT KINESIN (ARK) family is a plant-specific motor protein subfamily that consists of three members (ARK1, ARK2, and ARK3) in Arabidopsis thaliana. ARK2 has been shown to participate in root epidermal cell morphogenesis. However, whether and how ARK2 associates with microtubules needs further elucidation. Here, we demonstrated that ARK2 co-localizes with microtubules and facilitates microtubule bundling in vitro and in vivo. Pharmacological assays and microtubule dynamics analyses indicated that ARK2 stabilizes cortical microtubules. Live-cell imaging revealed that ARK2 moves along cortical microtubules in a processive mode and localizes both at the plus-end and the sidewall of microtubules. ARK2 therefore tracks and stabilizes the growing plus-ends of microtubules, which facilitates the formation of parallel microtubule bundles.     

3，4苯并芘对内皮祖细胞生物学功能的影响

目的研究3,4苯并芘（BaP）对人脐血来源的内皮祖细胞（EPC）生物学功能的影响。方法密度梯度离心法分离获取人脐血单个核细胞,采用贴壁培养法培养MNC中的EPC,通过Dil标记的乙酰化低密度脂蛋白（Dil-ac-LDL）摄取实验和FITC标记的植物凝集素（FITC-UEA-I lectin）结合实验鉴定细胞。消化收集第3代细胞,分别采用细胞计数试剂盒（CCK-8）、黏附能力测定实验、Transwell小室法及Matrigel体外成血管试验观察BaP对EPC增殖能力、粘附能力、迁移能力及成血管能力的影响,并检测各组细胞培养上清液SOD含量。采用单因素方差分析及LSD-t检验进行统计学分析。结果采用贴壁培养法能成功培养出EPC;与正常对照组相比,BaP呈浓度依赖性降低EPC的增殖能力{正常对照组OD（1.02±0.04）显著高于BaP各组[BaP①组OD（0.66±0.04）,BaP②组OD（0.55±0.04）,BaP③组OD（0.35±0.05）,均P〈0.01],BaP染毒组间增殖能力差异亦有统计学意义（两两比较,均P〈0.01）};与正常对照组比较,BaP呈浓度依赖性降低EPC的黏附能力{正常对照组[（117.50±17.16）个/200倍镜]显著高于BaP各组[BaP①组（80.00±14.46）个/200倍镜,BaP②组（66.00±9.06）个/200倍镜,BaP③组（49.80±10.72）个/200倍镜,均P〈0.01],BaP染毒组间黏附能力差异亦有统计学意义（两两比较,均P〈0.05）};与正常对照组相比,EPC的迁移能力亦呈BaP浓度依赖性降低{正常对照组[（46.10±4.51）个/400倍镜]显著高于BaP各组[BaP①组（35.50±4.95）个/400倍镜,BaP②组（26.80±4.08）个/400倍镜,BaP③组（19.50±2.84）个/400倍镜,均P〈0.01],BaP染毒组间迁移能力差异亦有统计学意义（两两比较,均P〈0.01）};与正常对照组比较,EPC的成血管能力亦呈BaP浓度依赖性降低{正常对照组[（33.20±3.70）个/100倍镜]显著高于BaP各组[BaP①组（22.00±3.39）个/100倍镜,BaP②组（16.20±2.59）个/100倍镜,BaP③组（10.80±2.39）个/100倍镜,均P〈0.01],BaP染毒组间成血管能力差异亦有统计学意义（两两比较,均P〈0.05）}。同时细胞培养上清液中SOD的活力也呈BaP浓度依赖性地降低{正常对照组[（22.6±2.19）U/ml]高于BaP各组[BaP①组（15.94±1.68）U/ml,BaP②组（12.5±1.58）U/ml,BaP③组（6.9±1.55）U/ml,均P〈0.01],BaP染毒组间SOD活力差异亦有统计学意义（两两比较,均P〈0.01）}。结论 BaP体外诱导显著影响EPC的多种生物学功能,其机制可能与氧化损伤有关。     

不明原因发热的临床综合诊断

不明原因发热是临床常见疑难疾病,其定义随着疾病种类的不断变化及诊断流程的进步而逐步完善。不明原因发热(feverof unknown origin,FUO)病因包括感染性疾病、风湿免疫性疾病及肿瘤性疾病三大类,三大病因各自具有不同临床特征。详细病史采集、全面体格检查及常规实验室检查对诊断FUO至关重要,必要时应及时进行相关影像学及病理学检查,疾病的最终诊断应综合分析病史及实验室检查。     

Later Passages of Neural Progenitor Cells from Neonatal Brain Are More Permissive for Human Cytomegalovirus Infection

Congenital human cytomegalovirus (HCMV) infection is the most frequent infectious cause of birth defects, primarily neurological disorders. Neural progenitor/stem cells (NPCs) are the major cell type in the subventricular zone and are susceptible to HCMV infection. In culture, the differentiation status of NPCs may change with passage, which in turn may alter susceptibility to virus infection. Previously, only early-passage (i.e., prior to passage 9) NPCs were studied and shown to be permissive to HCMV infection. In this study, NPC cultures derived at different gestational ages were evaluated after short (passages 3 to 6) and extended (passages 11 to 20) in vitro passages for biological and virological parameters (i.e., cell morphology, expression of NPC markers and HCMV receptors, viral entry efficiency, viral gene expression, virus-induced cytopathic effect, and release of infectious progeny). These parameters were not significantly influenced by the gestational age of the source tissues. However, extended-passage cultures showed evidence of initiation of differentiation, increased viral entry, and more efficient production of infectious progeny. These results confirm that NPCs are fully permissive for HCMV infection and that extended-passage NPCs initiate differentiation and are more permissive for HCMV infection. Later-passage NPCs being differentiated and more permissive for HCMV infection suggest that HCMV infection in fetal brain may cause more neural cell loss and give rise to severe neurological disabilities with advancing brain development.     

The Investigation of Protein Diffusion via H-Cell Microfluidics

In this study, we developed a microfluidics method, using a so-called H-cell microfluidics device, for the determination of protein diffusion coefficients at different concentrations, pHs, ionic strengths, and solvent viscosities. Protein transfer takes place in the H-cell channels between two laminarly flowing streams with each containing a different initial protein concentration. The protein diffusion coefficients are calculated based on the measured protein mass transfer, the channel dimensions, and the contact time between the two streams. The diffusion rates of lysozyme, cytochrome c, myoglobin, ovalbumin, bovine serum albumin, and etanercept were investigated. The accuracy of the presented methodology was demonstrated by comparing the measured diffusion coefficients with literature values measured under similar solvent conditions using other techniques. At low pH and ionic strength, the measured lysozyme diffusion coefficient increased with the protein concentration gradient, suggesting stronger and more frequent intermolecular interactions. At comparable concentration gradients, the measured lysozyme diffusion coefficient decreased drastically as a function of increasing ionic strength (from zero onwards) and increasing medium viscosity. Additionally, a particle tracing numerical simulation was performed to achieve a better understanding of the macromolecular displacement in the H-cell microchannels. It was found that particle transfer between the two channels tends to speed up at low ionic strength and high concentration gradient. This confirms the corresponding experimental observation of protein diffusion measured via the H-cell microfluidics.     

家燕营巢特征及其对城市环境的适应性

城市面积在全球范围内迅速扩张,一些鸟类种群通过改变营巢特征,在与自然生境截然不同的城市中筑巢繁殖。但目前城市环境对于鸟类营巢影响的研究较缺乏。为了解鸟类营巢对城市环境的适应,于2016、2019年在黑龙江哈尔滨的城市与乡村环境,分别测量家燕（Hirundo rustica）巢（如,大小及形状）及巢址特征等（如,距地面和屋顶距离）参数,以探究：（1）家燕巢特征在乡村及城市生境是否存在差异？（2）家燕巢特征在年际间是否存在变化？并为城市家燕种群的保护提供理论依据及合理建议。研究采用Kruskal-Wallis秩和检验以及Wilcoxon秩和检验比较分析所测量的巢特征参数在城乡之间、年际间的差异,并对组间参数进行线性判别分析（LDA,Linear Discriminant Analysis）。结果发现,城乡间具有显著差异：（1）与乡村相比,城市巢距离屋顶更远,距地相对更近（P<0.05）;（2）城市巢更浅（P<0.05）;（3）从2016到2019年,城市和乡村巢都变得更深,半径更大（P<0.05）。根据这些发现,推测城市楼房建筑的楼道为家燕繁殖提供了相对更为封闭、安全的环境,旧巢及较为丰富的支撑物为家燕提供了适宜的巢址,有可能节省亲鸟在营巢上的繁殖投入;但同时应当警惕门窗关闭、资源受限、人为干扰等不利因素可能造成的生态陷阱。