一种经济高效的用Silica提取纯化质粒DNA的方法

目的：为了达到批量提取质粒DNA的目的,在多次实验的基础上,建立一种经济、高效的质粒提取方法。方法：以pUC18、pET28b、pCAMBIA1304等3种质粒为材料,分别采用silica法和碱裂解法提取质粒DNA,通过质粒DNA浓度的紫外分光光度法定量测定、电泳分析和HindⅢ酶切鉴定,对两种质粒提取方法的效果进行了比较与评价;对silica法进行了改进和优化,进行大批量重组子的提取和验证。结果：silica法和碱裂解法提取质粒DNA效果相当,都可进行后续实验,但silica法具有经济、高效、无毒的优势。结论：silica法是一种简单、经济、高效的质粒提取方法,可用于批量质粒DNA提取。     

应用ABI PRISMTM310测序仪进行快速且经济的DNA测序

目的 使DNA测序更快速且经济。方法 根据测序模板的不同采取相应的测序方法。视具体情况调整测序反应条件,改进仪器的操作方法,减少测序反应体系及合理使用试剂,结果 对测序模板,测序条件和仪器操作的优化,可缩短测序时间,充分利用试剂,可降低测序成本。结论 在保证测序质量的前提下,不仅快速而且经济。     

A Rapid and Simple Method for DNA Engineering Using Cycled Ligation Assembly

DNA assembly techniques have developed rapidly, enabling efficient construction of complex constructs that would be prohibitively difficult using traditional restriction-digest based methods. Most of the recent methods for assembling multiple DNA fragments in vitro suffer from high costs, complex set-ups, and diminishing efficiency when used for more than a few DNA segments. Here we present a cycled ligation-based DNA assembly protocol that is simple, cheap, efficient, and powerful. The method employs a thermostable ligase and short Scaffold Oligonucleotide Connectors (SOCs) that are homologous to the ends and beginnings of two adjacent DNA sequences. These SOCs direct an exponential increase in the amount of correctly assembled product during a reaction that cycles between denaturing and annealing/ligating temperatures. Products of early cycles serve as templates for later cycles, allowing the assembly of many sequences in a single reaction. To demonstrate the method’s utility, we directed the assembly of twelve inserts, in one reaction, into a transformable plasmid. All the joints were precise, and assembly was scarless in the sense that no nucleotides were added or missing at junctions. Simple, efficient, and low-cost cycled ligation assemblies will facilitate wider use of complex genetic constructs in biomedical research.     

A Simple and Rapid Aqueous Method for the Purification of DNA from Wheat Chloroplast

We report here a simple and rapid method for the purification of chloroplast DNA (ctDNA) from wheat (Triticum aestivum). It utilizes an aqueous procedure, which does not involve at any stage running of gradients. Due to use of DEPC which inactivates DNases activated by EDTA, the DNase action on crude chloroplast preparation containing ctDNA is avoided.     

Cytokinins: A Rapid Extraction and Purification Method

A method is described for the isolation, purification and quantitation of free cytokinin bases and ribosides using ethyl acetate at pH 7.7 for the extraction. The extraction is almost complete (97.7%) as determined by using N⁶-(Δ²-isopentenyl)adenine-8-¹⁴C. The subsequent fast purification by chromatography on a standardized silica gel column in chloroform-methanol (7:3 v/v) is followed by thin layer chromatography (silica gel 60 F²⁵⁴) in chloroform-acetic acid (8:2 v/v). The recovery of N⁶-(Δ²-isopentenyl)adenine-8-¹⁴C after this two step purification was 78–82%. The efficiency of the method was determined by applying this procedure to N⁶-(Δ²-isopentenyl)adenine and N⁶(Δ²-isopentenyl)adenosine. Using gas liquid chromatography the recovery for N⁶-(Δ²-isopentenyl)adenosine was determined to be 61% and compared to 43% for N⁶-(Δ²-isopentenyl)adenosine, showing the suitability of the described method for gas liquid chromatography.     

A Rapid Method for Purification of Oligonucleotides

Abstract

A simple, rapid and novel method for purification of the oligonucleotide using silica gel matrix is described.     

大肠杆菌质粒的快速提取

质粒的提取是生物学中的常规技术之一 ,但常规法提取质粒十分复杂 ,繁琐并且费时。国外公司的试剂盒大大简化了此过程。但价格使国内大多数实验室难以承受。我们利用自制的玻璃粉提取质粒 ,大大简化了提取过程 ,同时相对于进口试剂盒而言 ,实验成本也大大降低     

A Rapid Method for Purification of d-Glucoside 3-Dehydrogenase

The stability of palm oil was tested by subjecting it to elevated temperatures for different durations of time, viz; at 80°C for 150 hr and 60°C for 400 hr.

The following results were obtained.

(1) The absorption spectrum resembled that of carotenoid and this changed progressively with a rise in peroxide and carbonyl values during the first 80 hr at 80°C.

(2) Peroxide values of Sabah palm oil were higher compared to Sumatra oil, there were marked increases in peroxide and carbonyl values of alkali refined oil as compared to crude oil. On the contrary, the residual color of crude Sumatra oil decreased considerably. Moreover, the steam emulsion number of alkali refined Sumatra oil was double the initial value after 400 hr.     

A Rapid Method for Purification of Myelin Basic Protein

A rapid procedure for purification of myelin basic protein has been developed. White matter is delipidated with 2-butanol, and the residue is extracted at pH 7.5 and 8.5. Myelin basic protein is solubilized by extraction in acetate buffer, pH 4.5. The entire procedure requires less than 4 h, and gives homogeneous protein essentially free of protease activity. This procedure can be scaled down to process milligram amounts of white matter; thus it can be very useful for purification of myelin basic protein from very limited amounts of human white matter obtained during surgery.     

LTSE—新的大肠菌群快速检测法

本文报道快速简易、结果准确、应用广泛、效益明显,总大肠菌群和粪大肠菌群在15～18小时内同时检出,费用比部颁三步发酵法(GB)节省75～84%的大肠菌群检测新方法。笔者研究出 Lactose-Tryptone-Sodium Dodecyl sulfate-Trace Elements[简称 LTSE]培养基,加上几种鉴定验证试验的 LTSE 法,用来快速检测水质、食品、冷饮、餐具等各类样品703件,结果与国标法(GB)对照总符合率为99.3%;同时与美国《水和废水标准检验法》(16版1985年)新发展的现行标准检测粪大肠菌群的正式 A-1法相比较特异性强,灵敏度高,能提高检出率,并节约经费38.1%.     

一种快速提取葱属植物 DNA 的方法

目的：针对分子育种工作的繁琐性及葱属植物次生代谢物较多的特点,研究一种利用普通试剂及简单仪器高效快速提取葱属植物DNA的方法。方法：对碱处理法稍加改进,在碱性环境中高温裂解细胞,将释放的DNA保存在Tris缓冲液中,用PCR检测提取质量并分析DNA的保存时间。结果：与用试剂盒提取的DNA相比,扩增产物无显著差异,利用此方法提取DNA并分析了13个洋葱品种的细胞质雄性不育类型;保存时间分析显示,DNA在常温下只能保存5d,在4℃或-20℃可至少保存2个月。结论：该方法方便快速、成本低、适于田间操作,得到的DNA可满足分子生物学实验的基本要求,可用于以PCR为基础的分子标记辅助育种。     

A Simple and Rapid Method for the Purification of Ovine Pineal Arylalkylamine N-Acetyltransferase

A two-step chromatographic procedure has been developed for the purification of ovine pineal arylalkylamine N-acetyltransferase (EC 2.3.1.87), based on the principles of disulfide exchange and anion exchange. The enzyme from 20 ovine pineal glands can be purified about 500-fold in a day; recovery is about 5%. Polyacrylamide gel electrophoretic analysis of the final preparation shows four major bands; one appears to be arylalkylamine N-acetyltransferase.     

一种快速握取植物总DNA的良好材料与方法

本文介绍以种子植物幼嫩胚乳为材料,用改良的氯仿－异戊醇快速提取方法提取植物总ＤＮＡ。这种材料和方法,细胞磨碎容易,ＤＮＡ收取量大,操作简便,适用于遗传操作及教学实验等方面的ＤＮＡ快速制备。     

A Rapid Chromosome-Mapping Method for Cloned Fragments of Yeast DNA

A rapid and generally applicable method is described for mapping a cloned yeast DNA segment to the chromosome(s) from which it originated. The method is based upon the recent finding that the integration into a yeast chromosome of a segment of the 2 mu plasmid DNA results, in heterozygous diploids, in the specific loss of genetic information from the chromosome into which the 2 mu DNA was integrated (Falco et al. 1982). After verification of the accuracy of the method using several genes whose position was known in advance, the method was used to locate the yeast actin gene, which lies on the left arm of chromosome VI, about 50 cM distal to CDC4.     

A Rapid DNA Minipreparation Method Suitable for AFLP and Other PCR Applications

A rapid DNA minipreparation method was developed for rice and other plant species. This method uses an Eppendorf tube and 1-ml pipette tip to grind plant tissues, and requires only one transfer for DNA isolation. In a single day, one person can complete DNA isolation from more than 120 leaf samples. The yields of the DNA samples ranged from 2.3 to 5.2 g from 25–50 mg fresh leaf tissue. DNA samples extracted using this method from rice were completely digested with five restriction enzymes (EcoR I, EcoR V, Hind III, Mse I and Pst I) and were successfully used for AFLP and other PCR applications.     

简单快速分离无RNA的大肠杆菌质粒DNA的方法

本文介绍一种简单快速分离质粒ＤＮＡ方法。此方法有两个主要步骤。用这种方法分离的质粒ＤＮＡ纯度高、无ＲＮＡ,并可用于酶切、连接等操作。     

从非变性聚丙烯酰胺凝胶回收纯化DNA样本

介绍一种从非变性聚丙烯酰胺凝胶中回收和纯化目标DNA片段的方法,经比较回收纯化前后PCR产物的电泳结果,表明该方法具有简单快捷和效率高的优点。     

A Rapid, High-Yield Purification of L-Alanine:4,5-Dioxovalerate Transaminase from Rat Kidney Mitochondria Using an Improved Enzyme Assay Method

The present report documents an improved enzyme assay method for the mammalian L-alanine:4,5-dioxovalerate transaminase which is of significant utility in work with crude tissue homogenates, cell cultures, or purified enzyme preparations. We also describe a new and rapid purification procedure for this enzyme from rat kidney mitochondria. The three-step procedure involves the use of digitonin and lubrol for mitochondrial matrix preparation and L-alanine-Sepharose 4B column chromatography followed by gel filtration on Sepharose 6B. By this procedure it is possible to obtain a highly purified enzyme preparation in a relatively short time with a 37.5% yield.     

利用寡核苷酸探针快速筛选重组克隆

提出了利用寡核苷酸探针快速筛选重组克隆的方法,在数小时内即可完成杂交和自显影的过程。以克隆羊β－乳球蛋白基因（ＢＬＧ）第一内含子为例进行了说明。     

A Simple Method for the Purification of Organelle DNA of Plants

We report here a simple procedure for the purification of the organelle DNA. Mitochondrial DNA from Sorghum and the chloroplast DNA from Populus and spinach were purified using this protocol. The method utilizes a quick centrifugation of the isolated organelle DNA through a two step CsCl density gradient for removal of small molecular weight nucleic acids which pose a major problem for getting clean restriction patterns. This method of purification can be adopted with any isolation procedure for organelle DNA.