极大螺旋藻藻蓝蛋白基因在巴斯德毕赤酵母X-33中表达的研究

通过PCR技术克隆了极大螺旋藻藻蓝蛋白基因，构建了克隆载体和表达载体，并将该基因导入巴斯德毕赤酵母X-33基因组，筛选了能够有效表达藻蓝蛋白基因的毕赤酵母工程菌株PC8，从而为发酵法生产藻蓝蛋白奠定了基础。     

High-yield production of the human lysozyme by Pichia pastoris SMD1168 using response surface methodology and high-cell-density fermentation

BackgroundLysozyme plays a crucial role in innate immunity with its well-recognized bacteriolytic activity. In this study, the influence of expression parameters (inoculation volume, culture volume, growth time, induction temperature and time, initial pH and methanol concentration) on human lysozyme (HLZ) production in recombinant P. pastoris SMD1168 was investigated through Plackett–Burman (PB) design and response surface methodology (RSM).ResultsIt was revealed that induction temperature, induction time and culture volume had significant influence (P < 0.01) on HLZ expression level, which were elected for further optimization with three-dimensional response surface designs for enhanced HLZ production. The highest lysozyme activity reached 3301 U/mL under optimized conditions (at 23.5°C for 90 h with culture volume of 48 mL) in shake flask, which increased 2.2 fold compared with that achieved with the standard protocol (Invitrogen). When high-cell-density fermentation of the recombinant Pichia pastoris was performed in a 15 L fermenter under optimized conditions, the extracellular lysozyme activity reached 47,680 U/mL. SDS-PAGE analysis of the product demonstrated that HLZ was produced as a single major protein with a molecular weight of approximately 14.7 kDa, consistent with its expected size.ConclusionsThe results indicated that the optimized culture conditions using PB design and RSM significantly enhanced the expression level of HLZ, and the Pichia expression system for HLZ production was successful and industrially promising.     

南极假丝酵母(Candida antarctica)肪酶A在毕赤酵母中的表达、纯化及性质

将编码南极假丝酵母脂肪酶A的基因进行克隆,与pPIC9K连接构建成重组载体.并将其电转化入毕赤酵母GS115中.通过MD和MM平板及PCR扩增,筛选和鉴定重组子.重组细胞经过120 h培养后.发酵液上清酶活达到13 U/mL.聚丙烯酰胺凝胶电泳显示,重组脂肪酶分子量约为50 kD.比原始脂肪酶略大.粗酶液经中空纤维素膜超滤浓缩、硫酸铵分级沉淀和离子柱交换层析后.得到在SDS-PAGE上显示为单一条带的重组脂肪酶.该酶最适反应温度为75 ℃,在70℃保持30 min后,仍具90%以上的活性,最适反应pH值为7.0,在pH6.0～8.0的范围内,具有一定的稳定性.     

南极假丝酵母(Candida antarctica)脂肪酶A在毕赤酵母中的表达、纯化及性质

将编码南极假丝酵母脂肪酶A的基因进行克隆,与pPIC9K连接构建成重组载体,并将其电转化入毕赤酵母GS115中。通过MD和MM平板及PCR扩增,筛选和鉴定重组子。重组细胞经过120 h培养后,发酵液上清酶活达到13 U/mL。聚丙烯酰胺凝胶电泳显示,重组脂肪酶分子量约为50 kD,比原始脂肪酶略大。粗酶液经中空纤维素膜超滤浓缩、硫酸铵分级沉淀和离子柱交换层析后,得到在SDS-PAGE上显示为单一条带的重组脂肪酶。该酶最适反应温度为75℃,在70℃保持30 min后,仍具90%以上的活性,最适反应pH值为7.0,在pH6.0～8.0的范围内,具有一定的稳定性。     

Release of formaldehyde during the biofiltration of methanol vapors in a peat biofilter inoculated with Pichia pastoris GS115

BackgroundMethanol can be effectively removed from air by biofiltration. However, formaldehyde is one of the first metabolic intermediates in the consumption of methanol in methylotrophic microorganisms, and it can be released out of the cell constituting a secondary emission.ResultsThe total removal of methanol was achieved up to input loads of 263 g m⁻³ h⁻¹ and the maximum elimination capacity of the system was obtained at an empty bed residence times of 90 s and reached 330 g m⁻³ h⁻¹ at an input methanol load of 414 g m⁻³ h⁻¹ and 80% of removal efficiency. Formaldehyde was detected inside the biofilter when the input methanol load was above 212 g m⁻³ h⁻¹. Biomass in the filter bed was able to degrade the formaldehyde generated, but with the increase of the methanol input load, the unconsumed formaldehyde was released outside the biofilter. The maximum concentration registered at the output of the system was 3.98 g m⁻³ when the methanol load was 672 g m⁻³ h⁻¹ in an empty bed residence times of 60 s.ConclusionsFormaldehyde is produced inside a biofilter when methanol is treated in a biofiltration system inoculated with Pichia pastoris. Biomass present in the reactor is capable of degrading the formaldehyde generated as the concentration of methanol decreases. However, high methanol loads can lead to the generation and release of formaldehyde into the environment.How to cite: Guerrero K, Arancibia A, Cáceres M, et al. Release of formaldehyde during the biofiltration of methanol vapors in a peat biofilter inoculated with Pichia pastoris GS115. Electron J Biotechnol 2019;40. https://doi.org/10.1016/j.ejbt.2019.04.003.     

Release of formaldehyde during the biofiltration of methanol vapors in a peat biofilter inoculated with Pichia pastoris GS115

BackgroundMethanol can be effectively removed from air by biofiltration (Shareefdeen et al., 1993; Babbitt et al., 2009 [1,2]). However, formaldehyde is one of the first metabolic intermediates in the consumption of methanol in methylotrophic microorganisms (Negruţa et al., 2010 [3]), and it can be released out of the cell constituting a secondary emission.ResultsThe total removal of methanol was achieved up to input loads of 263 g m⁻³ h⁻¹ and the maximum elimination capacity of the system was obtained at an empty bed residence times of 90 s and reached 330 g m⁻³ h⁻¹ at an input methanol load of 414 g m⁻³ h⁻¹ and 80% of removal efficiency. Formaldehyde was detected inside the biofilter when the input methanol load was above 212 g m⁻³ h⁻¹. Biomass in the filter bed was able to degrade the formaldehyde generated, but with the increase of the methanol input load, the unconsumed formaldehyde was released outside the biofilter. The maximum concentration registered at the output of the system was 3.98 g m⁻³ when the methanol load was 672 g m⁻³ h⁻¹ in an empty bed residence times of 60 s.ConclusionsFormaldehyde is produced inside a biofilter when methanol is treated in a biofiltration system inoculated with Pichia pastoris. Biomass present in the reactor is capable of degrading the formaldehyde generated as the concentration of methanol decreases. However, high methanol loads can lead to the generation and release of formaldehyde into the environment.How to cite: Guerrero K, Arancibia A, Caceres M, et al. Release of formaldehyde during the biofiltration of methanol vapors in a peat biofilter inoculated with Pichia pastoris GS115. Electron J Biotechnol 2019;40. https://doi.org/10.1016/j.ejbt.2019.04.003.     

我国南北方制造业工业设计应用现状对比分析

在对国内工业设计发展状况的资料调查中发现,南北方工业设计的发展不均衡,南方地区对工业设计应用的广度和深度明显优于北方;但北方制造业在对工业设计的认识上与南方制造业相当,大多数企业在接触工业设计的时间上甚至远远早于南方企业。本文对这种现状的形成及原因进行了探讨。     

研究生"链式"培养模式与传统模式的比较及构建障碍解决方法

在对比研究生“链式”培养模式与传统培养模式的基础上,阐述了“链式”培养模式的优势和特点,并对构建研究生“链式”培养模式实践中出现的观念、定位和中间层培养问题展开了一些探讨。     

The characterization of four gene expression analysis in circulating tumor cells made by Multiplex-PCR from the AdnaTest kit on the lab-on-a-chip Agilent DNA 1000 platform

Introduction

Nowadays, on-a-chip capillary electrophoresis is a routine method for the detection of PCR fragments. The Agilent 2100 Bioanalyzer was one of the first commercial devices in this field. Our project was designed to study the characteristics of Agilent DNA 1000 kit in PCR fragment analysis as a part of circulating tumour cell (CTC) detection technique. Despite the common use of this kit a complex analysis of the results from a long-term project is still missing.

Materials and methods

A commercially available Agilent DNA 1000 kit was used as a final step in the CTC detection (AdnaTest) for the determination of the presence of PCR fragments generated by Multiplex PCR. Data from 30 prostate cancer patients obtained during two years of research were analyzed to determine the trueness and precision of the PCR fragment size determination. Additional experiments were performed to demonstrate the precision (repeatability, reproducibility) and robustness of PCR fragment concentration determination.

Results

The trueness and precision of the size determination was below 3% and 2% respectively. The repeatability of the concentration determination was below 15%. The difference in concentration determination increases when Multiplex-PCR/storage step is added between the two measurements of one sample.

Conclusions

The characteristics established in our study are in concordance with the manufacturer’s specifications established for a ladder as a sample. However, the concentration determination may vary depending on chip preparation, sample storage and concentration. The 15% variation of concentration determination repeatability was shown to be partly proportional and can be suppressed by proper normalization.Key words: lab-on-a-chip devices, capillary electrophoresis, multiplex PCR, circulating tumour cells, Agilent DNA 1000 kit     

DNA Barcoding and the International Barcode of Life Project in China

<正>1.Scientific and Social Benefits of DNA Barcoding Along with the accelerated global trade and climate change,the needs for sustainable development and for understanding biodiversity are increasing.Rapid and accurate species identification and sustainable utility of biodiversity resources have become a great need for the world.     

The nature of indexing: how humans and machines analyze messages and texts for retrieval. Part I: Research,and the nature of human indexing

Does human intellectual indexing have a continuing role to play in the face of increasingly sophisticated automatic indexing techniques? In this two-part essay, a computer scientist and long-time TREC participant (Pérez-Carballo) and a practitioner and teacher of human cataloging and indexing (Anderson) pursue this question by reviewing the opinions and research of leading experts on both sides of this divide. We conclude that human analysis should be used on a much more selective basis, and we offer suggestions on how these two types of indexing might be allocated to best advantage. Part one of the essay critiques the comparative research, then explores the nature of human analysis of messages or texts and efforts to formulate rules to make human practice more rigorous and predictable. We find that research comparing human vs automatic approaches has done little to change strongly held beliefs, in large part because many associated variables have not been isolated or controlled.Part II focuses on current methods in automatic indexing, its gradual adoption by major indexing and abstracting services, and ways for allocating human and machine approaches. Overall, we conclude that both approaches to indexing have been found to be effective by researchers and searchers, each with particular advantages and disadvantages. However automatic indexing has the over-arching advantage of decreasing cost, as human indexing becomes ever more expensive.     

旋花科植物雄性细胞的细胞质DNA存在状况及其系统学意义

用DAPI荧光染色方法检测了旋花科4属4种植物——空心菜Ipomoea aquatica Forsk. 、茑萝 Qua- moclit pennata (Desr．)Boj．、月光花 Calonyction aculeatum(L．)House和日本菟丝子Cuscuta japonica Choisy 中生殖细胞和精细胞中细胞质DNA存在的状况。证明除日本菟丝子外，其余3种植物的雄性细胞中都 存在细胞质DNA。此外，在我们曾研究过的5种旋花科种植物的生殖细胞和精细胞中亦显示含有细胞质 DNA，因而可认为这是旋花科较普遍的特性，日本菟丝子精细胞中缺少细胞质DNA，可作为—个新的胚胎学证据，支持此属从旋花科分出独立为菟丝子科。     

西藏人体寄生虫病的流行与防治探讨

本文从西藏寄生虫病流行的三个基本环节与三个影响因素入手,对西藏业已调查清楚的人体寄生虫病的流行进行分析研究,旨在探索出西藏人体寄生虫病流行的三个基本环节的特点及其规律,影响因素,重点研究社会因素的影响,并从西藏人体寄生虫病防治政策制定,医疗预防、卫生宣传等方面深入分析,结合实际,提出有价值的防治措施。     

高原地区小儿甲流流行特征及防控对策

文章就西藏地区小儿甲流流行特征及诊疗经验作一阐述,同时提出了医疗机构如何应对和防控甲流疫情。     

Policy entrepreneurship in the co-evolution of institutions, preferences, and technology: Comparing the diffusion of totally chlorine free pulp bleaching technologies in the US and Sweden

This paper uses the concept of policy entrepreneurship to explain the social processes underlying sustainable technical change in the pulp and paper industry in the 1990s. It presents a detailed case study on the differences in the transition to and diffusion of totally chlorine free (TCF) pulp bleaching methods in Sweden and the USA. This change was sustained in the former, while it was almost absent in the latter. The paper studies the causes for this difference. It looks at the complex coevolution of technology, institutions, and consumer preferences that has taken place in both countries and draws conclusions on how transitions to sustainable paths in production and consumption may be supported by economic and social policy.     

On the role of human and machine metadata in relevance judgment tasks

In order to evaluate the effectiveness of Information Retrieval (IR) systems it is key to collect relevance judgments from human assessors. Crowdsourcing has successfully been used as a method to scale-up the collection of manual relevance judgments, and previous research has investigated the impact of different judgment task design elements (e.g., highlighting query keywords in the document) on judgment quality and efficiency. In this work we investigate the positive and negative impacts of presenting crowd human assessors with more than just the topic and the document to be judged. We deploy different variants of crowdsourced relevance judgment tasks following a between-subjects design in which we present different types of metadata to the human assessor. Specifically, we investigate the effect of human metadata (e.g., what other human assessors think of the current document, as in which relevance level has already been selected by the majority crowd workers), machine metadata (e.g., how IR systems scored this document such as its average position in ranked lists, statistics about the document such as term frequencies). We look at the impact of metadata on judgment quality (i.e., the level of agreement with trained assessors) and cost (i.e., the time it takes for workers to complete the judgments) as well as at how metadata quality positively or negatively impact the collected judgments.