A High-Throughput Screening System Based on Fluorescence-Activated Cell Sorting for the Directed Evolution of Chitinase A

Chitinases catalyze the degradation of chitin, a polymer of N-acetylglucosamine found in crustacean shells, insect cuticles, and fungal cell walls. There is great interest in the development of improved chitinases to address the environmental burden of chitin waste from the food processing industry as well as the potential medical, agricultural, and industrial uses of partially deacetylated chitin (chitosan) and its products (chito-oligosaccharides). The depolymerization of chitin can be achieved using chemical and physical treatments, but an enzymatic process would be more environmentally friendly and more sustainable. However, chitinases are slow-acting enzymes, limiting their biotechnological exploitation, although this can be overcome by molecular evolution approaches to enhance the features required for specific applications. The two main goals of this study were the development of a high-throughput screening system for chitinase activity (which could be extrapolated to other hydrolytic enzymes), and the deployment of this new method to select improved chitinase variants. We therefore cloned and expressed the Bacillus licheniformis DSM8785 chitinase A (chiA) gene in Escherichia coli BL21 (DE3) cells and generated a mutant library by error-prone PCR. We then developed a screening method based on fluorescence-activated cell sorting (FACS) using the model substrate 4-methylumbelliferyl β-d-N,N′,N″-triacetyl chitotrioside to identify improved enzymes. We prevented cross-talk between emulsion compartments caused by the hydrophobicity of 4-methylumbelliferone, the fluorescent product of the enzymatic reaction, by incorporating cyclodextrins into the aqueous phases. We also addressed the toxicity of long-term chiA expression in E. coli by limiting the reaction time. We identified 12 mutants containing 2–8 mutations per gene resulting in up to twofold higher activity than wild-type ChiA.     

江西省水系连通及水美乡村建设试点县实施方案编制存在的问题及对策

为提高水系连通及水美乡村建设试点县实施方案编制质量，更好地参加全国竞争性立项，本文分析研究了江西省55个水系连通及水美乡村建设试点县实施方案和相应的专家评审意见，对实施方案编制过程中存在的符合性、完整性不够，实施范围情况调查不准等七个方面突出问题进行梳理剖析，并提出相应对策，可为实施方案编制提供参考。     

超高层劲性结构及大型机械设备立体协同施工技术

基于某超高层建筑群项目的主体施工阶段,对超高层劲性结构建筑群项目施工过程中可能遇到的问题,诸如大型机械立体协同、钢结构与钢筋连接节点优化、钢结构桁架层施工等重难点进行总结分析,并提出相应的解决方案以供探讨,可为同类型工程提供参考经验。     

基于深度学习的彩色以及近红外图像去马赛克

单传感器捕获的彩色-近红外（RGB-NIR）图像存在光谱干扰，从而导致重建出的标准彩色图像（RGB）图像与近红外（NIR）图像存在色彩失真以及细节信息模糊。针对这个问题提出一种基于深度学习的去马赛克方法，通过引入跳远连接与稠密连接解决了梯度消失和梯度弥散问题，使得网络更容易训练，并且提升了网络的拟合能力。首先，用浅层特征提取层提取了马赛克图像的像素相关性以及通道相关性等低级特征；然后，将得到的浅层特征图输入到连续多个的残差稠密块以提取专门针对去马赛克的高级语义特征；其次，为充分利用低级特征与高级特征，将多个残差稠密块提取到的特征进行组合；最后，通过全局跳远连接恢复最终的RGB-NIR图像。在深度学习框架Tensorflow上使用公共的图像与视觉表示组（IVRG）数据集、有植被的户外多光谱图像（OMSIV）数据集和森林（Forest）三个公开数据集进行实验。实验结果表明，所提方法优于基于多级自适应残差插值、基于卷积卷积和神经神经网络以及基于深度残差U型网络的主流的RGB-NIR图像去马赛克方法。     

复杂地形下风力机尾流数值模拟研究和激光雷达实验对比

以CFD数值计算和实验相结合的方法,对处于中国西南某多山地区陆上风电场的尾流特性进行研究,验证不同数值方法在复杂地形的适用性。首先采用2台激光雷达,测量目标风力机一个月内的自由来流风速和尾流廓线,在地形上坡加速效应下,不同大气稳定度下目标风力机的自由来流风速廓线均呈负梯度。然后分别采用经典致动盘和改进致动盘法,模拟目标风力机在主风向下的尾流发展。不同于只有风速与压降关系的经典致动盘法,改进致动盘法更考虑了叶片几何和气动参数(尺寸信息、攻角、桨距角、升阻力系数等)。通过与后置激光雷达尾流测试结果对比,这2种基于CFD技术的数值模拟方法,计算网格相同,计算时间相当,且均能较好地模拟因为复杂地形而引起的尾流偏转;其中改进致动盘的尾流形状与激光雷达相似,速度亏损也更接近激光雷达结果。因此,改进致动盘法更适合于复杂地形条件下风场模拟,较好平衡了计算的效率与精度。     

定子分段式永磁直线同步电机速度波动抑制方法研究

为了解决定子分段式永磁直线同步电机(PMLSM)存在的因磁阻效应、负载阻力、摩擦力、参数摄动以及动子进出定子时耦合面积变化所造成的控制性能下降的难题,根据动子和定子的耦合状态,提出一种在完全耦合阶段和动子退出定子阶段的分段式控制方法。首先,在动子与定子完全耦合阶段采用改进滑模控制器减小推力波动导致的速度波动,再通过加入扰动观测器降低滑模切换项所带来的抖振现象;在动子退出定子阶段,建立相关电磁参数与动子位置的函数关系,实时补偿由耦合面积变化引起的动子失速,使动子速度在退出时接近给定值。仿真及实验结果表明动定子完全耦合过程中的速度稳态误差为0.005 m/s,收敛时间为0.3 s,动子退出定子阶段的速度波动不超过0.04 m/s,满足定子分段式PMLSM用于长行程自动运输系统对平稳性及快速性的需求。     

Effect of paddy variety,milling status and aeration on the progeny emergence of Sitophilus oryzae L. (Coleoptera: Curculionidae)

In Sri Lanka, insects cause enormous losses in stored paddy; the rice weevil Sitophilus oryzae is a devastating insect. The magnitude of losses in stored paddy caused by insects varies with characteristics of paddy but has not yet been fully investigated. Due to the increased concerns on the use of synthetic chemicals, safer alternative pest management strategies for stored paddy are needed. Objectives of this study were to determine the effect of paddy variety, milling status and nature of aeration on the progeny emergence of S. oryzae. The experimental design was a three-factor factorial, completely randomized design (CRD).Freshly emerged S. oryzae adults were introduced to un-milled paddy or milled/polished rice belonging to traditional and improved varieties. The samples were maintained either open or airtight, and the progeny adults emerged was determined at monthly intervals. Progeny emergence was lower in the traditional varieties, un-milled paddy and under air-tight condition compared to improved varieties, milled/polished rice and aerated samples, respectively. Overall, the improved variety Bg 300 exhibited the maximum resistance to infestation by S. oryzae. Furthermore, white-colour and long-grain varieties produced lower progeny of S. oryzae than red-colour and short-grain varieties, respectively. It is concluded that the maximum protection in paddy/rice from insect infestation during storage can be achieved by using traditional varieties, stored as un-milled paddy (without dehusked) under airtight condition.     

火灾条件下拱顶油罐弱连接结构的失效分析

拱顶油罐受外部火焰烘烤发生失效爆炸，易造成严重人员伤亡和巨大经济损失。为对火灾条件下拱顶油罐的失效爆炸进行预警，通过高温材料力学性能实验平台对拱顶油罐弱连接结构进行破坏测试，得出不同火灾温度条件下弱连接结构失效的拉断力和应力值。实验结果表明，当温度高于300℃时，拉断力和失效应力值变化显著，其中450℃为变化峰值温度。在此基础上，构建了拱顶油罐失效压力计算模型，推导出了不同容积拱顶油罐在不同火灾温度下的失效压力。所得结果可作为拱顶油罐失效的判定依据，为油罐的爆炸预警提供数据支撑。     

Asynchronous control for two‐dimensional hidden Markovian jump systems with partly known mode observation conditional probabilities

This study focuses on the asynchronous control problem for two‐dimensional discrete‐time hidden Markovian jump systems where the mode observation conditional probability matrix is partly known. Considering the original system modes are invisible, the observed modes emitted from an observer serve as an alternative for stability analysis and controller design where a mode observation conditional probability matrix is constructed to characterize the emission between system modes and observed modes. Specially, only partly known information of the mode observation conditional probability matrix is accessible. With the introduction of the free‐connection weighting matrices, the asymptotic mean square stability criterion is firstly derived based on Lyapunov method. This introduction provides a further degree of relaxation and less conservatism is therefore achieved. Secondly, we present synthesis conditions for asynchronous state feedback controller design given in terms of a set of interconnected linear matrix inequalities. Moreover, cluster concept based on the partitions of observed modes is adopted which helps to decrease the number of controllers and simplify the design complexity. A numerical example, regarding the cases with and without clustering of the observed modes, is presented to illustrate the effectiveness of the proposed method.     

基于稳定连接的配电网边缘划分方法及应用研究

随着能源互联网的建设,为了应对配电主站所面临的数据接入和计算压力,文章提出了二级配电网边缘架构,以分布式边缘智能架构缓解主站压力。主要从边缘定义、划分方法、边缘功能描述与交互描述4个方面对所提边缘架构进行介绍。提出电网稳定连接区域概念及其生成方法,并结合南京地区电网实例,说明了以电网稳定连接区域作为边缘划分的合理性。最后,对所提边缘架构在配电网中的实际应用场景进行了设计和展望。