Dvl1及其突变体腺病毒载体的构建与在MSCs中表达鉴定

Dvl1及其结构缺失突变体ΔDIX（dsh and axin）和ΔDEP（dsh,Egl-10 and pleckstrin）腺病毒载体的构建并验证其在MSCs（mesenchymal stem cells）中的表达情况。将目的基因定向克隆至pAdTrack-CMV穿梭载体上,并经PmeⅠ线性化后在BJ5183细菌中与pAdEasy-1骨架质粒同源重组,获得重组腺病毒载体,在QBI-293A细胞中包装及扩增,实时荧光定量PCR和Western bolt验证Dvl1在MSCs中的表达情况。经PCR、PacⅠ单酶切鉴定及测序分析,成功构建Ad-Dvl1、Ad-ΔDIX和Ad-ΔDEP腺病毒载体,目的基因序列与GenBank报道一致,并选出150 MOI为最适感染MSCs的感染复数,成功构建了Ad-Dvl1、Ad-ΔDIX和Ad-ΔDEP腺病毒载体,并获得高滴度的病毒子,qPCR和Western blot证实Dvl1在MSCs中高表达并增加了β-catenin在细胞中的累积,为进一步研究Dvl1在MSCs迁移过程的作用奠定了基础。     

一种大规模体数据压缩体绘制策略

针对基于GPU的大规模体数据直接体绘制过程中遇到的显存不足的问题，提出了一种大规模体数据的压缩绘制策略.该策略结合小波变换和分类矢量量化进行数据压缩，采用基于GPU的光线投射算法进行绘制，在绘制时，只解压变换当前绘制所需要的极少数数据，并结合多分辨率绘制，实现实时交互.基于CUDA的实验表明：该压缩绘制策略有效解决了显存不足问题，并且在保证交互绘制帧率的前提下获得了较好的成像质量.     

自动折叠喷洒臂的设计及运动学特性

为了解决当前卷盘式喷灌机所配套的喷灌车自动化水平较低的问题,设计了一种基于电动缸驱动的自动折叠喷洒臂,并在喷洒臂之间连接处设计了一种当喷洒臂完全展开时能够实现自密封的端面密封铰链.基于喷洒臂完全折叠时收拢于车架两侧,完全伸展时铰链左右铰合密封等特征建立各杆件参数约束方程,确定了喷洒臂的各杆件尺寸.通过矢量方程法对喷洒臂进行运动学分析,以电动杆推杆的转角、位移、速度、加速度为输入量,各杆件的角位移、角速度、角加速度为输出量,通过Simulink搭建喷洒臂的运动仿真框图并进行分析.结果表明:完全折叠时,喷管1与水平方向呈90°夹角;伸展过程中,各喷管运动平稳,且始终保持在水平直线方向;完全展开时,3个喷管转动90°,且各喷管铰接处完全闭合不发生水泄漏现象,说明设计具有一定的合理性.     

Preparation of Adjuvant Diluent for Live Vaccine against Classical Swine Fever and Evaluation of its Immune Effects

The aim of this study was to develop a novel oil-in-water (O/W) emulsion as adjuvant diluents (AD) for live vaccine against classical swine fever (CSF) that could effectively enhance the immune effect of vaccine.The AD was prepared by high-pressure homogenization technique.Formulations and preparation parameters were optimized with response surface design.Its stability, particle size, polydispersity (PDI) and Zeta potential were characterized.The humoral immune response and cellular immune response of the AD were evaluated with BALB/c mice by intramuscular injection.The particle size of the AD prepared by optimized formulation and parameters was 100.4 nm, PDI was 0.147, and Zeta potential was —28.7 mV.The experiment results showed that the AD had good stability.The AD was inoculated combined with live vaccine against CSF into BALB/c mice by intramuscular injection.The results showed that the live vaccine against CSF specific immune responses could be evoked in mice by co-inoculation with AD and vaccine.The cellular immune response levels in co-inoculated groups were significantly higher than control group (P<0.05), with obvious phenomena of higher levels of IFN-γ, IL-6 and IL-4 in serum.The result revealed that cellular immune capability significantly improved with the AD.The results strongly revealed that cellular immune capability significantly improved by introducing AD for effective immune-adjuvant for live vaccine against CSF.     

Immunogenicity of multiple-epitope antigen gene of HCV carried by novel biodegradable polymers

In order to develop a promising vaccine candidate utilizing a combined approach to induce both antibody production and T-cell activity, the DNA fragment containing MA of HCV with five conserved epitopes was synthesized. Two types of HCV vaccine candidates (the DNA type and DNA/polymers) were constructed using MA. PLA-PEG-PLA and PLGA-PEG-PLGA were synthesized and used as micelles with encapsulated plasmid pcDNA3.1(+)-MA. The preparation of copolymers, the cloning and analysis of recombinant plasmid DNA, in vitro expression, and immunogenicity in transgenic mice were evaluated in detail. The results indicated that even single immunization and oral immunization with DNA/polymers achieved satisfying immune responses in vivo tests. As biodegradable and nontoxic triblock copolymers, the novel copolymers demonstrated a great advantage, as they made long-term and single-immunizing vaccines possible; in addition, the copolymers showed a better adjuvant effect and scarcely any side effects.     

拟南芥DUF647家族成员RUS4植物表达载体的构建及亚细胞定位

ROOT UV-B SENSITIVE4 (RUS4)是拟南芥DUF647蛋白家族的一个功能未知的成员。RUS1和RUS2曾经报道和根UV-B-感应途径相关联,并在拟南芥早期幼苗形态发生和发育中发挥重要作用。为了探究RUS4的分子功能,本研究对RUS4蛋白进行了亚细胞定位分析。首先,我们通过Gateway TOPO载体系统,构建了融合蛋白表达载体pMDC83-RUS4;然后,通过农杆菌介导的花苞转化法,获得了转化pMDC83-RUS4的转基因拟南芥株系;对转基因植物叶肉原生质体的荧光显微镜观察显示,RUS4-GFP信号与叶绿体的自发荧光共定位,说明RUS4蛋白定位叶绿体中。该结果为进一步研究RUS4的分子功能奠定了良好的基础。     

基于PCA-SVR-ARMA的狮头鹅养殖禽舍气温组合预测模型

为提高狮头鹅养殖禽舍气温预测精度,提出了基于主成分分析(Principal Component Analysis,PCA)、支持向量回归机(Support Vector Regression,SVR)融合自回归滑动平均(Autoregressive Moving Average,ARMA)模型的狮头鹅养殖禽舍气温组合预测模型。在建模过程中,运用主成分分析法筛选狮头鹅养殖禽舍气温的关键影响因子,消除变量之间冗余信息,约简预测模型结构;采用SVR-ARMA构建狮头鹅禽养殖舍气温组合预测模型,先通过SVR对气温进行预测,再由基于ARMA模型的残差预测值修正气温预测结果。利用该模型对广东省汕尾市2018年7月21日至2018年7月30日期间的狮头鹅养殖禽舍气温进行预测。结果表明,该组合预测模型取得了良好的预测性能,与标准BP神经网络、标准SVR、PCA-BPNN(反向传播神经网络,BackPropagationNeuralNetwork)、PCA-SVR和PCA-BPNN-ARMA等模型对比分析,其评价指标平均绝对误差、均方根误差和平均绝对百分比误差分别为0.183 2℃、0.454 0℃和0.005 9,均表明所提出的组合模型具有更高的预测效果,不仅能够满足狮头鹅养殖禽舍气温实际精准调控的需要,还为狮头鹅健康养殖和种苗繁育环境精细化管理提供决策。     

利用CRISPR/Cas9系统编辑日本结缕草ZjSGR基因技术研究

日本结缕草(Zoysia japonica)是一种常见的具有众多优良性状的暖季型草种,秋冬季节在我国北方地区会较早出现叶片脱绿现象,在一定程度上限制了日本结缕草在北方的大面积使用。ZjSGR是从结缕草中分离出来的一种衰老诱导基因,负调控叶片滞绿性状。本研究利用CRISPR/Cas9系统,通过基因枪介导的遗传转化方法实现对日本结缕草ZjSGR基因的编辑,经测序验证获得1株单碱基缺失突变植株。突变植株叶片深绿,在正常生长温度(28~30℃)条件下,和对照相比,叶绿素含量更高(P<0.05)。本研究为进一步阐述ZjSGR基因在日本结缕草滞绿性状调控中的作用机理提供了基础研究材料,也为培育日本结缕草滞绿品种育种工作奠定了基础。     

草果AtNCED基因克隆、表达和植物表达载体构建

9-顺式环氧类胡萝卜素双加氧酶(9-cis-epoxycarotenoid dioxygenase, NCED)是ABA生物合成的限速酶。诸多研究表明NCED基因表达量的变化与ABA含量显著相关,在种子休眠过程中发挥重要作用。为探究草果AtNCED基因的序列特征和功能,本研究以草果(Amomum tsaoko Crevost et Lemarie)种子转录组为基础,采用RT-PCR技术克隆了一个AtNCED基因。该基因全长2 372 bp,包含一个1 830 bp的开放阅读框(open reading frame, ORF),编码610个氨基酸。生物信息学分析显示,AtNCED基因编码的蛋白在N-末端包含典型的叶绿体转运肽序列,在催化结构域含有4个高度保守的组氨酸残基。AtNCED蛋白与芭蕉科马来西亚蕉同源性较高,与其他单子叶植物处在同一进化分支。实时荧光定量PCR分析结果表明,AtNCED基因表达量随着层积时间的增加逐渐下降,该基因可能正向调控种子休眠,在草果种子休眠诱导与维持中发挥重要作用。进一步地,利用同源重组方法,成功构建了pBI121-AtNCED过表达载体,并转化至农杆菌EHA105。该研究结果为弄清ABA激素信号在种子休眠中的调控作用提供了帮助。