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人类基因组计划的主要任务之一就是要从大片段基因组区域或整条染色体DNA 上鉴定出基因表达序列(gene expressed sequences)或转录单位(transcription units)。在人类基因组30亿个碱基对中，发生转录的表达序列(即基因)仅占总序列的3～5％。基因组中 绝大部分是基因间隔序列(intergenic seguences)或 内含子(intron)和各种各样的重复序列 ...

基因打靶技术是一种定向改变生物活体遗传信息的实验手段，它的产生和发展建立在胚胎干(ES) 细胞技术和同源重组技术成就的基础之上，并促进了相关技术的进一步发展。基因打靶技术将广泛应用于基因功能研究、人类疾病动物模型的研制以及经济动物遗传物质的改良等方面。 1 ．原理 首先获得ES 细胞系，利用同源重组技术获得带有研究者预先设计突变的中靶ES 细胞。通过显微注射或者胚胎融合的方法将经过遗传修饰的ES ...

DNA片段的克隆需要合适的载体，载体或是质粒，或是噬菌体，或是病毒，通常大多经过人工改造一、质粒 常用的有pBR322，pUC系列质粒等。 （一）pBR322质粒是4362bp的环状双链DNA载体，有2个抗药性基因（四环素和氨苄青霉素），一个复制起始点和多个用于克隆的限制酶切点。当缺失抗药性基因的大肠杆菌被pBR322成功地转化时，它便从该质粒获得了抗生素抗性。两个抗生素基因中均含供插入外源DN ...

Simple and Efficient Method (SEM) to Make Competent Cells Preparation of Frozen Competent of DH5α 1) 250 ml TB solution 10mM Pipes or 10 mM Hepes 0.65g 15 mM CaCl2 0.55g 250 mM KCl 4.66g ~undefined55 ...

Transformation DNA fragments (or plasmid DNA) into competent E. coli ~undefined Caution: Use aerosol protecting tips if selection of transformed cells is not based on X-gal strategy. 1.Remove competent cells ...

Does anyone have expereince of extracting DNA from urine? I am having problems with this. I have previously extracted the DNA but then it degraded very quickly and since then I havent even been able t ...

I am planning to perform sodium bisulphite sequencing. Currently I am trying to design primers of the promoter region of my target genes. However some of these promoters have not been well-characteriz ...

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(adapted from Bruce A. Roe Department of Chemistry and Biochemistry The University of Oklahoma Norman Oklahoma 73019 broe@ou.edu) Typically 2.5 - 3 volumes of an ethanol/acetate solution is added to t ...

Hello I have been trying over the last few months to introduce a single point mutation in a looped portion of an RNA using the dut-ung method of mutagenesis. So far I have had little success.&nb ...

A practical method for the preparation of total DNA from filamentous fungi M.I. Borges M.O. Azevedo R. Bonatelli Jr. M.S.S. Felipe and S. Astolfi-Filho Departamento de Biologia Celular Universidade de ...

Electroelution of agarose fragments Electroelution buffer 1 M Tris pH 7.5 12.0 mls 0.5 M EDTA 0.24 mls 1 M NaCl 3.0 mls qs to 600 mls dH2O Acetate cushion 3 M NaAcetate pH 4.8 480 μl 0.1 % Bromphe ...

I have tried to purify my PCR products by using 3 different approaches: 1) QIAQuick PCR purification kit: but when I run the gel to see its efficacy there was simply nothing!! it cleaned everything in ...

Kitto Lab The University of Texas at Austin http://research.cm.utexas.edu/bkitto/Kittolabpage/Protocols/Microbiology/electroporation.html This procedure prepares glycerol stock cultures of bacteria f ...

Kitto Lab The University of Texas at Austin http://research.cm.utexas.edu/bkitto/Kittolabpage/Protocols/Microbiology/ethanolPpt.html This procedure allows the concentration of DNA samples from dilute ...

Mitochondrial DNA Isolation from Somatic Embryogenic Cell Cultures of Larix (Excerpt from Thesis). Mitochondrial DNA is isolated by a modification of the methods described by Wilson and Chourey (1984) ...

Preparation of Sonicated Salmon Sperm DNA Procedure: 1) Use Pharmacia #27-4564-01. With clean flamed scissors and forceps weigh 0.25 g/50 ml conical and add 50 ml/conical of 0.02 M Tris pH 7.6. Allo ...

The Preuss LabThe Division of Biological SciencesThe University of Chicago http://preuss.bsd.uchicago.edu/protocols/topo.html ...

Contributed by Dr. A. Gratchev Single Nucleotide Primer Extension is a powerful method which can be used for the precise analysis of methylation in a certain position. The procedure is shown on the fi ...

Hi all! We are trying to extract genomic DNA from the leaves of an Ericacea (we suppose it is rich in tanins and phenolics) and we can't do it! We have used two different kits which work OK for other ...