Microfluidic-Based Cationic Cholesterol Lipid siRNA Delivery Nanosystem: Highly Efficient In Vitro Gene Silencing and the Intracellular Behavior

Safe and efficient delivery of small interfering RNA (siRNA) is essential to gene therapy towards intervention of genetic diseases. Herein, we developed a novel cationic cholesterol lipid derivative (CEL) in which cholesterol hydrophobic skeleton was connected to L-lysine cationic headgroup via a hexanediol linker as the non-viral siRNA delivery carrier. Well-organized CEL/siRNA nanocomplexes (100–200 nm) were prepared by microfluidic-assisted assembly of CEL and siRNA at various N/P ratios. The CEL and CEL/siRNA nanocomplexes have lower cytotoxicity compared with bPEI25k. Delightfully, we disclosed that, in Hela–Luc and H1299–Luc cell lines, the micro-fluidic-based CEL/siRNA nanocomplexes exhibited high siRNA transfection efficiency under both serum-free condition (74–98%) and low-serum circumstances (80–87%), higher than that of lipofectamine 2000. These nanocomplexes also showed high cellular uptake through the caveolae/lipid-raft mediated endocytosis pathway, which may greatly contribute to transfection efficiency. Moreover, the time-dependent (0–12 h) dynamic intracellular imaging demonstrated the efficient delivery to cytoplasm after lysosomal co-localization. The results indicated that the microfluidic-based CEL/siRNA nanosystems possessed good stability, low cytotoxicity, high siRNA delivery efficiency, rapid cellular uptake and caveolae/lipid raft-dependent internalization. Additionally, this study provides a simple approach for preparing and applying a “helper lipid-free” cationic lipid siRNA delivery system as potential nanotherapeutics towards gene silencing treatment of (tumor) diseases.     

Efficient synthesis of tyrosol from L-tyrosine via heterologous Ehrlich pathway in Escherichia coli

For the efficient conversion of L-tyrosine(L-Tyr) to tyrosol, which is an aromatic compound widely used in the pharmaceutical and chemical industries, a novel four-enzyme cascade pathway based on the Ehrlich pathway of Saccharomyces cerevisiae was designed and reconstructed in Escherichia coli. Then,the expression levels of the relevant enzymes were coordinated using a modular approach and gene duplication after the identification of the pyruvate decarboxylase from Candida tropicalis(Ct PDC) as ...     

吸附交联法固定化蔗糖异构酶及其在异麦芽酮糖制备中的应用

以壳聚糖为载体、戊二醛为交联剂,采用吸附交联法对重组短小芽孢杆菌来源的蔗糖异构酶进行固定化。以表观酶活力回收率为指标,对壳聚糖浓度、戊二醛加量、游离酶加量、固定化时间等条件进行了优化;并考察了温度、pH、固定化酶加量、反应时间以及底物浓度等因素对固定化蔗糖异构酶转化生产异麦芽酮糖的影响。结果表明,最佳固定化条件为：壳聚糖质量浓度3 g/dL、戊二醛加量（体积分数）0.75%、酶加量50 U/g、固定化时间16 h,此时固定化酶活力回收率达到70.3%;最佳转化条件为：温度30 ℃、初始pH 4.5、酶用量15 U/g,转化10 h,蔗糖质量浓度600 g/L,异麦芽酮糖最大产物得率达到87.8%。在最佳的转化条件下连续转化16次,产物得率仍保持在87.52%,显示该固定化酶具有良好的操作稳定性及较高的异麦芽酮糖合成能力。     

重组酿酒酵母全细胞催化合成莱鲍迪苷A

用经密码子优化后合成的甜叶菊UDP糖基转移酶UGT76G1编码基因构建了表达该酶的重组酿酒酵母YPH499(pYES2-UGT)。建立了通过柠檬酸钠调节酿酒酵母细胞内由葡萄糖到尿苷二磷酸葡萄糖(UDPG)的代谢通量,用于催化甜菊苷合成莱鲍迪苷A的全细胞催化方法。优化后的催化体系为:甜菊苷1 g/L,葡萄糖20g/L,普郎尼克F68 10 g/L,MgCl2 6 g/L,柠檬酸钠15 g/L,pH 7.2,细胞密度200 g湿细胞/L反应液,在37℃下经72 h后转化生成莱鲍迪苷A 267.89 mg/L。     

旋涂法制备PVDF/PTFE复合膜及其性能研究

旋涂技术具备薄膜厚度精确可控、高性价比、节能、低污染等优点,在薄膜制备方法中脱颖而出。本文使用旋涂法制备了聚偏氟乙烯(PVDF)/聚四氟乙烯(PTFE)复合膜,并对其制备工艺进行了优化,同时对旋涂成膜机理进行了研究。通过单因素试验,分析了PVDF溶液浓度、旋涂仪的旋转速度和成膜温度对复合膜的拉伸强度和水通量的影响,利用电子扫描显微镜(SEM)对复合膜胶粘层的微孔孔径变化进行了分析,结果表明,PVDF/PTFE复合膜制备的最佳工艺:PVDF溶液浓度为6 g/L,旋涂仪的旋转速度为2500 r/min以及成膜温度为90℃。该条件下制备的复合膜的膜厚度为200μm,平均孔径为20μm,拉伸强度可达到26.34MPa,水通量可达到605.2 L/(m2·h)。此外,分析旋涂PVDF/PTFE复合膜成型的机理,其原因是由于PVDF溶胶分子与PTFE基膜之间存在范德华力等分子间力以及溶胶分子渗透到基膜微孔之间所形成的钩钉键等粘合作用。     

四种速生杨木制浆及漂白性能的研究

本文对四种5年生速生阔叶材的制浆及漂白性能进行了比较研究，结果表明：三倍体毛白杨的纤维较长，分布均一，具有较大的长宽比和小的壁腔比，纤维素含量较高，木素含量较低，是一种较好的造纸原料。四种原料分别进行了碱性亚纳一蒽醌法蒸煮并采用DED三段漂白，对其纸浆性能进行检测分析，结果表明：三倍体毛白杨易蒸煮、易漂白，其成浆得率和白度较高，物理强度较好。     

猪皮胶原的酶法提取

以工业化的蛋白酶为原料 ,采用 3种不同的工艺 ,对提取猪皮胶原的生物酶法进行了研究。研究发现 :猪皮的酶水解产物分子呈不连续分布 ,主要集中在某些分子量处 ,与碱法工艺明胶分子量的分布明显不同 ;且酶处理条件不同 ,水解得率与产物分子量的分布也不同。     

浅析坦比哀多教堂的传承和影响

该文从坦比哀多教堂对古典建筑的传承,对阿尔伯蒂理念的发扬,帕拉第奥对坦比哀多的推崇以及坦比哀多对后世建筑的影响这四个方面,来分析这一文艺复兴集中式宗教建筑的开山之作的传承与影响,以及在西方建筑历史的重要意义.     

潮汕传统建筑装饰题材的深层语义

归纳整理了潮汕传统建筑装饰题材的发展沿革、取材类型、取材特点,并结合儒学思想、民俗美学、社会美学等方面的理论,初步分析了当地传统建筑装饰题材所承载的民俗文化和美学取向.     

婴幼儿食品源阪崎克罗诺杆菌的3种分型方法

阪崎克罗诺杆菌污染婴幼儿食品可能导致新生儿和婴幼儿脑膜炎、败血症和坏死性小肠结肠炎,对其健康造成严重威胁。采用脉冲场凝胶电泳(PFGE)、多位点序列分型(MLST)和肠杆菌间保守重复序列PCR(ERIC-PCR)对分离于婴幼儿配方乳粉、米粉和辅食中的阪崎克罗诺杆菌进行分子分型,计算3种分型方法的辛普森多样性指数(DI)值,评价3种方法的分型效率,以便精确和快速开展阪崎克罗诺杆菌流行病学研究。PFGE、MLST和ERIC-PCR可分别将37株阪崎克罗诺杆菌分为35个PFGE基因型、28个ERIC-PCR型和23个序列(ST)型,分型结果的DI值分别为99.55%,96.40%和98.05%。3种方法对阪崎克罗诺杆菌均具有较高的分型能力,其中PFEG分型能力最高,分型效率最佳。