Chemicon公司的CpGenomeDNAModificationKit的protocol修正

我用了快4个月这个试剂盒了,下面是一些总结出来的经验,我现在诱变每次都很成功了,呵呵:)1. 我溶NaOH的水PH值大概在6.5左右,所以我每次调Reagent I的时候3MNaO0H基本都要加150uL左右。2. 解链时用50度水浴15分钟。3. 加入Reagent I后,50度水浴16小时。4. 加入Reagent II和III后,室温孵育10分钟5. 第一次离心用9000 rpm 5分钟。6. 70%乙醇清洗3次,每次9000rpm 1 分钟,最后一次13000rpm2min。7. 加入20mM NaOH/90%EtOH后,反映5分钟。8. 不要去掉20mM NaOH/90%EtOH,直接加入90%乙醇后离心,9000rpm 1min9. 90%乙醇清洗2次,每次9000 rpm 1 min,最后一次13000rpm 3min10. PH 7.5 TE buffer 65度 溶解 15 minPS: 如果发现PCR结果中M......阅读全文

PCR产物纯化方法

Purification of PCR Products in Preparation for CloningJoseph SambrookPeter Maccallum Cancer Institute and The University of Melbourne, AustraliaDavid

The-PRC2-Complex-Sets-Long-杢erm-Gene-Silencing-Through-Modification

Packaging of DNA into chromatin allows the cell to store its genetic information efficiently and has an important role in regulating gene expression.

Detection-of-Intracellular-Antigens-by-Flow-Cytometry

实验概要Fix and Perm  reagents are designed for use with all commercially available flow  cytometers. Alignment and compensation should be performed accor

Guide-to-Cell-Proliferation-and-Apoptosis-Methods

Chapter 1: Cell Death - Apoptosis and Necrosis1.1Introduction21.1.1Terminology of cell death21.1.2Differences between necrosis and apoptosis31.1.3Apop

RNA-FISH-on-cultured-cells-in-interphase

IntroductionFluorescence in situ hybridization (FISH) has become a widely used method in genome and molecular genetic studies. The technique is highly

Automated-Genomic-DNA-Extraction

实验概要This section  provides a general protocol for automated isolation of genomic DNA from  10-20 µl blood samples in a 96-well format using the Charge

Transient-Transfection-of-Cos1-Cells

Transient Transfection of Cos-1 CellsNote: This protocol has been optimized for Cos-1 cells. Successful transfection of each cell type requires optimi

Protocol-to-Count-Cell-Number-of-Preimplantation-Embryos

Protocol to Count Cell Number of Preimplantation Embryos using Nuclear Staining with Hoechst 33342 or DAPI  Introduction The following is a simple pro

修正活络止痛丸的成分

  鸡血藤、何首乌、过岗龙、牛大力、豨莶草、豆豉姜、半枫荷、两面针、臭屎茉莉、走马胎、威灵仙、连钱草、千斤拔、独活、穿破石、薏苡仁、土五加、钩藤、山白芷、宽筋藤。辅料为:炼蜜。[1]

外显子基因为什么保守?《Nature-Genetics》出新答案了!

   基因的基本结构分外显子和内含子,比较不同物种的基因发现外显子序列通常是保守的,而内含子序列则很少保守。外显子的种间差异不大,换句话说,人和老鼠、线虫的外显子基因都很相似。  这种相似性的科学解释是,外显子是生物体生存的关键区域,这部分区域的许多变异都会导致死亡表型,所以未存于世。内含子由于没有

流式细胞仪技术专辑

Flow Cytometry Analysis (Springer Lab, Harvard University) Flow cytometry employs instrumentation that scans single cells flowing past excitation sour

Cell-Extraction-Protocol

实验概要Primary tissues  are valuable tools for the study of intracellular and extracellular  markers which characterize disease states. We have developed

Tissue-Harvest-Protocol

TISSUES TO BE PROCURED(minimally and preferably within 5-8 hours after death):1. Brain2. Liver3. Muscle4. Skin5: Others as the specific case dictatesP

Microarray-Hybridization-Protocol

Introduction:One microarray set consists of 7 nylon membranes with 2.5 x 7.5 cm dimension. 2304 genes were spotted onto nylon membranes (Schleicher an

Western-Blotting-Protocol

实验概要The western blot  (sometimes called the protein immunoblot) is a widely used analytical  technique used to detect specific proteins in the given s

Dot-Blot-Protocol

a. Label nitrocellulose membrane (using a pencil) to identify protein elution fractions.b . Pipette 2μl from each fraction onto the membrane, allow th

Protocol-for-dsRNA-Synthesis

实验概要        We routinely produce dsRNA by in vitro transcription of a PCR generated DNA template containing the T7 promoter sequence on both ends

Migration-Assay-Protocol

Materials to be prepared beforehand:1) FBS free medium2) 10% FBS medium3) Cell migration filter insert ( Transwell®, 12mm Diameter, 12 μm Pore Size.)P

RNA-Isolation-Protocol

Stabilize RNAStart with 15 ml E. coli Culture containing 7.5* 109 cells (OD600= 0.2 Dilute cells or scale up)Pipet 30 ml of RNAProtect Bacteria Reagen

Sandwich-ELISA-Protocol

实验概要The  Sandwich ELISA measures the amount of antigen between two layers of  antibodies (i.e. capture and detection antibody). The antigen to be  mea

cDNA/AFLP-Protocol

Preparation of Para-magnetic beads from Promega cat#Z5482:a) suspend magnetic particles in bottle - transfer 200 ul (200 ug) of beads per sampleof RNA

RNA-Isolation-Protocol

RNA Isolation Protocol(Revised 5-15-2003)Stabilize RNAStart with 15 ml E. coli Culture containing 7.5* 109 cells (OD600= 0.2 Dilute cells or scale up)

Immunofluorescence-Microscopy-Protocol

实验概要Immunofluorescence  allows the imaging of a specific factor in cells or tissue sections  through the use of a specific antibody chemically which i

Adhesion-Assay-Protocol

Materials to be prepared beforehand:1) Washing Buffer--0.1% BSA in medium (DMEM or RPMI)2) Blocking Buffer--0.5% BSA in medium (DMEM or RPMI)3) Lamini

Bacteria-Culture-Protocol

Bacteria Culture ProtocolBy 徐晓政1、TBS Medium Preparation:Prepare 1L of TBS medium contains:Tryptone 12gYeast extract 24gNaCl 5gSodium Succinate 5gGlyce

Protocol-for-Cell-Fusion

Healthy Sp2/0 cells should be rapidly growing by this time. Sp 2/0 cells should be started about two weeks before the cell fusion. Every two days, the

Western-Blot-Protocol

一、提取抗原蛋白将提取RNA途中留存的样品,加入150μl 100%酒精充分混匀,静置5min(RT), 2000×g , 4℃离心5min, 吸取上清至新管中, 加入750μl异丙醇, 混匀, 静置10min(RT), 12000×g, 4℃离心10min, 弃上清, 加入1ml 0.3mol/L

Colony-PCR-Protocol

1. Pull out eight glycerol stock plates from the –80oC freezer and set on bench top to thaw. Be sure to remove the foil seal before leaving the plates

TAIL-PCR-Protocol

TAIL is a series of reactions that are intended to map where a T-DNA (transfer DNA) has inserted within the genome. The main components of the 3 react

Yale-Immunofluorescence-Protocol

实验概要We provide a protocol for fixation, immunostaining, and imaging in 384-well Plates.主要试剂Reagents1. 384-well view plates (Aurora)2. HUVEC (pooled, L