一种新型双向启动子——棉花曲叶病毒启动子的分离,序列分析与功？…

以棉花曲叶病毒（ＣＬＣｕＶ）侵染的烟草叶片组织ＤＮＡ为模板,通过聚合酶链反应扩增ＣＬＣｕＶ双向启动子片段并子片段并插入克隆载体。序列分析和同源性比较表明,克隆的启动子长４３６ｂｐ,与目前发现的９种ＣＬＣｕＶ株系的启动子序列均不相同,同源性最高达９９．３２％。将启动子片段分别以不同方向与ＧＵＳ报告基因和ｎｏｓ终止子融合,构建了瞬时表达载体。通过基因枪法将质粒载体导入烟草和棉花叶片细胞中进行瞬时表达。     

人血小板生成素（hTPO）的cDNA克隆与序列测定

由于人血小板生成素ｃＤＮＡ的阅读框架较长而且序列中无合适的酶切位点，因此，我们利用一个同义突变创造的ＫｐｎＩ酶切位点人为将ｈＴＰＯ ｃＤＮＡ分成Ｎ－端和Ｃ－端两个片段。首先利用反转录聚合酶链式反应从人胎肝ｍＲＮＡ中分别扩增出两个ｃＤＮＡ片段，于ＥｃｏＲＩ，ＫｐｎＩ位点分别克隆入测序载体ｐＵＣ１９中。     

DNA指纹图谱在农作物品种鉴定中的应用

ＤＮＡ(脱氧核糖核酸 )是细胞染色体中的遗传基因 ,其分子各个片段就代表各个遗传信息。由于ＤＮＡ指纹图谱直接反映ＤＮＡ水平上的差异 ,它成为当今最先进的遗传标记系统。ＤＮＡ指纹图谱技术主要有 :限制性片段长度多态性(Ｒｅｓｔｒｉｃｔｉｏｎｆｒａｇｍｅｎｔｌｅｎｇｔｈｐｏｌｙｍｏｒ ｐｈｉｓｍ ,简称ＲＦＬＰ)、随机扩增多态性 (ＲａｎｄｏｍａｍｐｌｉｆｉｅｄｐｏｌｙｍｏｒｐｈｉｃＤＮＡ ,简称ＲＡＰＤ )、小卫星ＤＮＡ(Ｍｉｎｉ ｓａｔｅｌｌｉｔｅＤＮＡ)、微卫星ＤＮＡ(ＭｉｃｒｏｓａｔｅｌｌｉｔｅＤＮＡ)、内部简单重…     

PLA单体--丙交酯合成方法的研究进展

只有高分子量聚乳酸才能满足组织工程对其强度及其它性能的要求，而丙交酯是制备高分子量聚乳酸的重要原料，其纯度和成本的高低决定了高分子量聚乳酸的质量和价格，20世纪70年代以来，在提高丙交酯的产率，纯度，降低丙交酯的成本方面做了大量的工作，并取得一定的进展，文中综述了制备丙交酯的各种方法和工艺，并讨论了各自的优缺点，提出了制备丙交酯和高分子量聚乳酸的可能发展方向。     

小麦品种“Lee”中抗条锈病基因的RAPD标记

共用３８０个１０碱基随机引物，对抗条锈病基因ＹｒＬｅｅ的近等基因系进行了ＲＡＰＤ分析，其中有两个引物在抗病的近等基因系ＹｒＬｅｅ／６＊Ｔａｉｃｈｕｎｇ２９Ｆ３和基因供体亲本“Ｌｅｅ”中扩增出了特异的ＤＮＡ片段，而在轮回亲本“Ｔａｉｃｈｕｎｇ２９”中没有扩增出同样的ＤＮＡ片段，引物ＯＰＲ－７扩增出的ＤＮＡ片段长度约为４６５ｂｐ，引物ＯＰＲ－９扩增出的ＤＮＡ片段长度约国６６７ｂｐ。用基因供体亲本“Ｌｅ     

小麦品种“Lee”中抗条锈病基因的 PAPD 标记

共用３８０个１０碱基随机引物，对抗条锈病基因ＹｒＬｅｅ的近等基因系进行了ＲＡＰＤ分析，其中有两个引物在抗病的近等基因系ＹｒＬｅｅ／６＊Ｔａｉｃｈｕｎｇ２９Ｆ３和基因供体亲本“Ｌｅｅ”中扩增出了特异的ＤＮＡ片段，而在轮回亲本“Ｔａｉｃｈｕｎｇ２９”中没有扩增出同样的ＤＮＡ片段。引物ＯＰＲ－７扩增出的ＤＮＡ片段长度约为４６５ｂｐ，引物ＯＰＲ－９扩增出的ＤＮＡ片段长度约为６６７ｂｐ。用基因供体亲本“Ｌｅｅ”与感病品种“铭贤１６９”杂交的Ｆ２代分离群体（１８９株植株）进行的遗传连锁性分析表明，引物ＯＰＲ－７扩增出的特异ＤＮＡ片段与ＹｒＬｅｅ基因紧密连锁。     

乙肝病毒反义DNA的合成

为了深入研究反义ＤＮＡ对乙肝病毒的抑制作用，设计合成了针对乙肝病毒基因不同功能区域的反义片段２０个，无关序列４个，长度为１５聚到１８聚，分为无修饰反义ＤＮＡ、反义ＤＮＡ－多聚赖氨酸共阶连接物、部分硫代和全硫代反义ＤＮＡ四种。合成反应是在ＡＢＩ３８１型ＤＮＡ合成仪上以１μｍｏｌ或１０μｍｏｌ的规模进行的。但在计算机程序上作了一些修改以适应于部分硫代和全硫代ＤＮＡ合成。抑制实验的结果表明，所有合成的反     

骨骼肌膜DNA结合蛋白

应用差速及蔗糖密度梯度离心法提取了骨骼肌非核的肌膜蛋白,发现在肌膜上有一种ＤＮＡ结合蛋白质,其分子量为２９ｋＤａ,能与环状质粒ＤＮＡ和双链线状ＤＮＡ发生特异性结合,它可能在肌肉介导的基因转移中起作用     

乳酸-氨基酸共聚物的制备及研究发展

制备乳酸-氨基酸共聚物是改性聚乳酸的有效方法之一。综述了近年来乳酸-氨基酸共聚物的制备方法与特点,指出了较高分子量和高氨基酸含量聚合物的制备工艺是研发的重点。     

热休克后表达受抑基因的mRNA差别显示和克隆

以人成骨肉瘤细胞株（ＨＯＳ－８６０３）作为热休克模型，用差别显示ｍＲＮＡ法（ｄｄｍＲＮＡ）筛选，发现和克隆了一个热休克后表达受抑的ｃＤＮＡ片段。在４３℃、４０分钟热处理后１小时，ＨＯＳ－８６０３细胞总ｃＤＮＡ图谱未见明显变化，但以该ｃＤＮＡ片段作为探针，ＲＮＡｄｏｔｂｌｏｔ和Ｎｏｒｔｈｅｒｎｂｌｏｔ证实，热休克后该基因的ｍＲＮＡ的量明显下降，而ｍＲＮＡ未见明显降解。我们将这个因热休克而选择性的表达受抑的基因命名为ＨＳＳＧ－１。对全长３１２ｂＰ的ｃＤＮＡ片段的序列分析表明，ＨＳＳＧ－１可能是一个未知的新基因。     

有机-无机转化法制备石墨烯薄膜带的探索研究

分析了石墨烯的制备方法及其优缺点,为了实现大量生产石墨烯薄膜,提出了采用有机-无机转化方法制备石墨烯。将高分子量、低分子量分布的聚丙烯腈与二甲基亚砜配制成溶液,并制成分子结构为螺旋线性的截面形状为扁平的石墨烯薄膜带前驱体,前驱体经过热水、蒸汽牵伸(取向度高)、预氧化(环化和交联)和碳化(芳构化)后制成了石墨烯薄膜带。     

Limitations of electrospray ionization of fulvic and humic acids as visible from size exclusion chromatography with organic carbon and mass spectrometric detection

A method was developed to link size exclusion chromatography electrospray ionization mass spectrometry (SEC-ESI-MS) analyses of fulvic and humic acids with SEC and organic carbon detection (SEC-OCD), the latter providing an absolute measure of the amount of organic matter eluting from the SEC column. This approach allows us to determine which molecular weight fraction of the complex polydisperse mixtures is detectable by ESI-MS. It could be shown that the cone voltage setting for the ESI interface has strong impact on ESI-MS detection. Using conventional settings for low molecular weight compounds, the high molecular weight (HMW) compounds are hardly amenable to ESI-MS. With increasing cone voltage, an increasing signal intensity is obtained for the HMW fraction that elutes at shorter retention times. However, mostly fragment ions are obtained under these conditions. Thus, the range of compounds amenable to ESI-MS analysis is restricted by the limited stability of the fulvic and humic acid molecules of higher molecular weight in the electrospray process rather than by the mass spectrometer used. Compounds above 1000 amu are hardly visible as intact ions. However, insight into structural characteristics of these compounds can be gained by investigating their fragment ions by SEC-ESI-MS. The use of SEC-OCD parallel to SEC-MS helps to assess and optimize the detection potential of ESI-MS for polydisperse mixtures.     

On-line coupling of size exclusion chromatography with electrospray ionization-tandem mass spectrometry for the analysis of aquatic fulvic and humic acids

A method was developed for the analysis of humic and fulvic acids by size-exclusion chromatography-electrospray ionization-tandem mass spectrometry using a completely volatile eluent. Humic and fulvic acids were separated into three peaks. These fractions occupied different mass ranges and showed differences in the fine structure of their mass spectra. The low-molecular-weight (LMW) fraction of fulvic acids is most sensitively determined by ESI-MS, and it appears that previous results obtained by infusion-ESI-MS were primarily determined by this fulvic acid fraction. The average molecular weight of this fractions turned out to be lower than that reported from infusion-ESI-MS measurements. Its scan spectra and the product ion spectra of some of its molecular anions perfectly match those previously obtained from whole fulvic acid mixtures. Obviously, a class of well-defined polycarboxylated molecules exist that occurs in all fulvic acid fractions thus far investigated. With decreasing elution time and increasing molecular weight, detection by ESI-MS loses sensitivity as compared to the parallel UV recording, and the fine structure of the scan spectra becomes increasingly uniform for both fulvic and humic acids. The average molecular weight of the HMW fraction exceeds those values calculated from infusion experiments. Scan spectra and product ion spectra of the high-molecular-weight (HMW) fraction of both the humic and the fulvic acids suggest that the HMW fraction consists of several subunits that originate from the LMW fraction.     

Catalyst-free synthesis of polyesters via conventional melt polycondensation

Most commodity polyesters are synthesized via melt polycondensation of dicarboxylic acid and diol using metallic catalysts; however, the resultant metal residues can pose toxic effects on human and environment. Although polyesters can be synthesized through autocatalysis of dicarboxylic acid without additional catalysts, high molecular weight (HMW) products cannot be obtained by this strategy, which was previously attributed to the low equilibrium constant of esterification and the difficulty of removing water. Herein, we get a new understanding that the kinetic deviation of dicarboxylic acid/diol monomers is the only reason for the low molecular weight of polyesters by autocatalysis. Accordingly, we introduce a dynamic stoichiometric strategy to overcome this difficulty using anhydride-formable dicarboxylic acids as monomers through a tandem mechanism involving proton transfer, anhydride formation and re-esterification. A series of catalyst-free HMW polyesters, including poly(butylene succinate) (PBS), poly(ethylene succinate) (PES), poly(butylene succinate-co-butylene adipate) (PBSA), and poly(ethylene succinate-co-ethylene terephthalate) (PEST), were thereby synthesized. This new approach not only enables large-scale production of HMW polyesters comparable to commercial products, but also avoids the problems associated with catalysts, which is very promising for the applications with high safety requirements.     

Ionic Strength Effects in Aqueous Solutions of Gelatin and Sodium Dodecylsulphate

Abstract

The interaction between the anionic surfactant sodium dodecylsulphate and the polyampholyte gelatin in aqueous solution has been studied by viscosity and fluorescence. Three different gelatin molecular iveights, a polydisperse gelatin and two fractionated samples derived from it, have been examined at two ionic strengths. On addition of sodium dodecylsulphate SDS, the viscosity of a low molecular weight fractionated (o.) gelatin increases smoothly with SDS concentration, unlike the polydisperse (standard) and high molecular zveight (HMW) fractionated gelatins which show maxima. The viscosity increase at the maximum increases with gelatin molecular weight. On addition of 0.1 M salt, the viscosities of the gelatin-surfactant mixtures are lower than the equivalent no-salt cases. Fluorescence studies suggest that the SDS micelles adsorbed onto these various gelatins under two different ionic strengths are remarkably similar.     

嗜冷杆菌EastSeaG5-1415脂肪酶基因的克隆和序列测定

从东海深海底泥中筛选到一株产低温碱性脂肪酶的海洋细菌EastSeaG5-1415,经鉴定为嗜冷杆菌Psychrobacter glacincola。将其染色体DNA用Sau3AI部分酶切后,低熔点琼脂糖回收2～10kb的。DNA片段,用Klenow大片段酶半补齐,与用Sal I酶切半补齐的质粒pUC19连接后,转化E．coli．JM109构建基因文库。用平板测活法从基因文库初步筛选到两株产碱性脂肪酶的阳性克隆,采用ELISA反应进一步鉴定。序列测定分析表明,两个重组质粒中都包含长度为951bp的碱性脂肪酶基因的完整开放阅读框架(ORF)和上游基因调控序列。此片段编码有317个氨基酸组成的酶,计算分子量为35000 Dal。通过Southem杂交证实了此片段来自于嗜冷杆菌Psychrobacter glacincola EastSeaG5-1415基因细。     

High‐Throughput Block Optical DNA Sequence Identification

Optical techniques for molecular diagnostics or DNA sequencing generally rely on small molecule fluorescent labels, which utilize light with a wavelength of several hundred nanometers for detection. Developing a label‐free optical DNA sequencing technique will require nanoscale focusing of light, a high‐throughput and multiplexed identification method, and a data compression technique to rapidly identify sequences and analyze genomic heterogeneity for big datasets. Such a method should identify characteristic molecular vibrations using optical spectroscopy, especially in the “fingerprinting region” from ≈400–1400 cm^?1. Here, surface‐enhanced Raman spectroscopy is used to demonstrate label‐free identification of DNA nucleobases with multiplexed 3D plasmonic nanofocusing. While nanometer‐scale mode volumes prevent identification of single nucleobases within a DNA sequence, the block optical technique can identify A, T, G, and C content in DNA k‐mers. The content of each nucleotide in a DNA block can be a unique and high‐throughput method for identifying sequences, genes, and other biomarkers as an alternative to single‐letter sequencing. Additionally, coupling two complementary vibrational spectroscopy techniques (infrared and Raman) can improve block characterization. These results pave the way for developing a novel, high‐throughput block optical sequencing method with lossy genomic data compression using k‐mer identification from multiplexed optical data acquisition.     

茶树基因组DNA测试方法概述

在查阅国内外关于茶树DNA测试方法研究文献的基础上,结合茶树DNA测试方法的现状,对茶树基因组DNA的主要测试方法进行了归纳,指出了DNA的提取、定量及其分子标记三类测试方法研究中的主要进展,并对DNA提取过程中的主要影响因素、分子标记方法的优缺点进行了评述,提出对茶树中DNA测试方法实行标准化,是实现茶树的种质资源鉴定、优良品种选育的重要途径。     

果树树皮浸提物的紫外光谱分析

本文阐述了果树树皮浸提物的紫外光谱分析方法,实验中对30种不同树种、同一树种不同产地果树树皮浸提物的紫外光谱、一阶导数光谱特征峰的异同进行了比较,确定了利用紫外光谱法鉴别果树树皮种类的依据。     

Fluorescence lifetime for on-the-fly multiplex detection of DNA restriction fragments in capillary electrophoresis

This paper describes the first use of frequency-domain fluorescence lifetime for multiplex detection of DNA restriction fragments in capillary electrophoresis (CE). The fragments were labeled with monomeric intercalating dyes that can be excited by either the 488- or 514-nm line of an argon ion laser and have lifetimes in the range of 0.5-2.5 ns. We were able to achieve multiplex lifetime detection in the CE separation of a restriction fragment digest and a DNA size ladder in the same run, for fragments shorter than 700 bp. Different gel buffer systems, including a modified polyacrylamide gel and several tris-borate-EDTA/hydroxyethylcellulose (TBE/HEC) gels, were investigated for separation and detection of the dye-labeled DNA fragments. Best results for both electrophoretic resolution and lifetime detection were obtained using a gel containing 1% high molecular weight (90,000-105,000) HEC and 0.3% low molecular weight (24,000-27,000) HEC in TBE buffer.