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DNA的克隆是指在体外将含有目的基因或其它有意义的DNA片段同能够自我复制的载体DNA连接，然后将其转入宿主细胞或受体生物进行表达或进一步研究的分子操作的过程，因此DNA克隆又称分子克隆，基因操作或重组DNA技术。DNA克隆涉及一系列的分子生物学技术，如目的DNA片段的获得、载体的选择、各种工具酶的选用、体外重组、导入宿主细胞技术和重组子筛选技术等等。 一 目的DNA片段的获得 DNA克隆的第一步 ...

一．原理 ⑴ 5’-RACE法 ① 以目的mRNA 为模板，使用5’末端P 标记的RT 引物进行反转录反应，合成1ststrand cDNA。 ② 使用RNase H 分解 Hybrid DNA-RNA中的RNA链。 ③ 使用T4 RNA Ligase 使单链cDNA进行环化或形成首尾连接物。 ④ 进行PCR扩增。 图 4－2 5’-RACE 法原理 ⑵ 引物 ...

This procedure allows the concentration of DNA samples from dilute solution and the removal of unwanted salts from DNA samples. Materials •3M Sodium Acetate buffer pH 5.2 (store at 4 ℃) •Col ...

Dr. William H. Heidcamp Biology Department Gustavus Adolphus College http://homepages.gac.edu/~cellab/chpts/chpt14/ex14-6.html Materials •DNA •CsCl •0.3 N NaOH •0.2 M Tris-HCl buf ...

Equilibriation of Phenol Caution: Phenol and chlorofom are very toxic. Wear gloves protective eye wear and lab coat. Use hood. Phenol dissolves and denatures proteins. It dissolves protein that is ti ...

Hi This is my first post and I am desparately looking for some help recovering DNA from CsCl preps. I have recently run into some problems precipitating DNA using sodium acetate (3M 1:30 dilution) and ...

I'm looking for a methode to demethyle DNA in vivo during more than 5 days . All I've read about 5-aza-CdT is for maximum 5 days. Why ? Is there another drug and if yes how does it work ? thanks Elea ...

Description This method was suggested in a BRL "Focus" article several years ago (Zeugin & Hartley 1985). It is a useful way to avoid coprecipitation of proteins and accumulation of salt ...

Abstract: The “Freeze squeeze” method is a cost effective and technically simple procedure to isolate and prepare PCR products for DNA sequencing. Reagents •70% ethanol •3 M so ...

Amberg Lab Upstate Medical University http://www.upstate.edu/biochem/amberg/protocols/transformation.html 1.Grow cells to 1X10E8 or OD600 of 1.2-1.3. 2.Spin cells at 5000rpm for 5 min and wash pelle ...

Hi....I need a prot℃ol for extracting DNA from hair/roots...(specifically horse)...and any additional information such as ?storage pr℃edure/timing/etc....thanks.... -tree- --------------------------- ...

Submission Details: *Dr. Simon DawsoundefinedDepartment of BiochemistrundefinedUniversity of NottinghaundefinedU.K.*17:0undefined22/1/96. Database: Method/Protocol-->Spin column purification can be used to change buffers with ...

In the hood: 96 well dish with bacteria titertech microtubes glass pipette Remove colonies from each well using the titertech and place them into the cover Pipette up and down to thoroughly mix the co ...

Rapid elution of DNA from agarose gels James Movius Hahn Lab Last modified Sun Nov 1 1998 A method of quickly purifying agarose gel DNA fragments for use in subsequent reactions such as further restr ...

Author: Suzanne Gerttula Date: March 141994 1.Digest between 1 and 10 μg Genomic DNA. I digest at 37 C for about six hours over the course of a day. Spin down briefly heat to 65℃ for 10 min ...

Abstract: It is very difficult to isolate Rhizobium DNA due to the gum production by the organism. Hence we have designed a protocol for efficient isolation of DNA from the organism for further gene a ...

Hi I have a 9kb fragment already cloned into bluescript and I am trying to insert an extra 3.5kb. I've been trying for some time now with no success - when I do get colonies very few have any of the n ...

双元载体的农杆菌转化 1.1农杆菌感受态细胞的制备 1.1.1. 取-70℃保存的EHA105于含50μg/ml链霉素平板划线，28℃培养。 1.1.2. 挑取单菌落接种于5ml YM液体培养基中，220rpm 28℃振荡培养12-16 hr。 1.1.3. 取2ml菌液转接于100ml YM液体培养基中，28℃220rpm振荡培养至OD600=0.5。 1.1.4. 转入无菌离心管，500 ...

试验准备： 1、LB液体培养基(Luria-Bertani) ：称取蛋白胨(Tryptone)10 g，酵母提取物(Yeast extract) 5 g，NaCl 10 g，溶于800ml去离子水中，用NaOH调pH至7.5 加去离子水至总体积1升高压下蒸气灭菌20分钟。 2、LB固体培养基：液体培养基中每升加12g琼脂粉高压灭菌。 3、氨苄青霉素(Ampicillin Amp)母液：配成 50 ...

In situ immunodetection of 5-methylcytosine (5-mC) on squashed cells using anti-methylcytosine Cross-references are to Schwarzacher & Heslop-Harrison 2000. Practical in situ Hybridization. Bios Ox ...