DryTransfer干法蛋白轉膜
Trans-Bot SD Assembly1. Prepare the transfer buffer.2. Following electrophoresis, equilibrate the gels in transfer buffer. Equilibration facilitates the removal of electrophoresis buffer salts and detergents. The length of time required for equilibration is dependent on the gel thickness. For example, 15 min. for a 0.75 mm SDS-PAGE gel.3. Cut the membrane to the dimensions of the gel. Wet the ......閱讀全文
Dry－Transfer－干法蛋白轉膜
Trans-Bot SD Assembly1. Prepare the transfer buffer.2. Following electrophoresis, equilibrate the gels in transfer buffer.? Equilibration facilitates
SEMIDRY－ELECTROPHORETlC－TRANSFER－(WESTERN－BLOTS)
?Introduction??? After proteins have been separated by electrophoresis, individual protein bands can often be identified by using an antibody that is
Southern－Blotting:－DNA－Transfer
Southern Blotting: DNA Transfer1. Depurination of DNA fragments: Wash gel in 0.25 M HCl 5' (small gels), 7' (large gels)2. Denaturation: Wash
Transfer－of－Eukaryote－Suspension－Cultures
MaterialsFibroblast suspension cultureTissue culture laminar flow hoodMedia appropriate to culture line usedDisposable pipettes (10 ml and 1.0 ml)Disp
Horizontal－Transfer－of－Supernumerary－Chromosomes－in－Fungi
Several species of filamentous fungi contain so-called dispensable or supernumerary chromosomes. These chromosomes are dispensable for the fungus
Shuttle－for－transfer－of－acetyl－groups－from－mitochondria－to－the－cytosol
Acetyl-CoA is produced in mitochondria through the metabolism of fatty acids and the oxidation of pyruvate to acetyl-CoA. When ATP is needed, this ace
Noninvasive－Human－Nuclear－Transfer－with－Embryonic－Stem－Cells
Noninvasive Human Nuclear Transfer with Embryonic Stem CellsSohyun L. McElroy1?and?Renee A. Reijo PeraCenter for Human Embryonic Stem Cell Research an
COLONY－HYBRIDIZATION
COLONY HYBRIDIZATION1) CUT 4 PIECES OF 3MM WHATMAN PAPER AND PLACE EACH ONE IN A SEPARATE CONTAINER(A CAFETERIA TRAY WILL PROBABLY WORK PERFECTLY).2)
Western－雜交
Western?雜交（主要內容如下）Preparing of Protein LysatesWestern BlottingFar Western BlottingSemi Dry BlottingStripping MembranesTrouble Shooting and OthersPrepa
SOUTHERN－BLOT的步驟
1. Run the gel as normal. Often for genomic southerns it is desirable to run long gels (18cm) over 4-6hrs.2. Photograph the gel with a ruler adjacent
Southern雜交技術
?SOUTHERN BLOT1. Run the gel as normal. Often for genomic southerns it is desirable to run long gels (18cm) over 4-6hrs.2. Photograph the gel with a r
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
Immunostaining－of－Paraffin－Sections
Procedure:?1)?Fix tissues for 3 hr on ice in 4 formaldehyde (2.5 ml of Polysciences #18814 made up to 10 ml in 80 mM NaPO4 [3.2 ml of 1 M NaPO4] pH 6.
Pouring－Plates
1. For rich media, weigh out appropriate ingredients and place into a flask. Add water until appropriate volume. Use a flask at least 2 times larger t
Acid－Phenol－Yeast－RNA－Prep
This is the preferred method for yeast RNA preparationuse Gloves and RNAse free solutions throughout.1. Use a YPD overnight culture to innoculate fres
Harvesting－Hematopoietic－Cells－from－Mice
Materials4 mice from each genotype4 Ly5 miceBuckets with wet ice 3xBucket with dry ice 1xDewar flask with liquid nitrogen100 mL beakers with 95% ethan
Quick－and－Easy－Isolation－of－Genomic－DNA－from－Yeast
ProcedureTransfer 1.5 ml of liquid culture of yeast grown for 20 - 24 h at 30°C in YPD (1% yeast extract, 2% peptone, 2% dextrose) into a microcentrif
Sphingomyelin－Mass－Measurement
Protocol:Bligh & Dyer extraction1) Pellet approximately 1 X 107 cells.2) Resuspend pellet in 3 ml CHCl3: CH3OH (1:2) and vortex hard.3) Add 0.8 ml H2O
CGH－Protocols－（二）
DNA preparation by cryotom tissue dissectionPreparations/Materials:?Cool cryostat down to -20 to -30°C about 3 hours prior to dissection?Label eppendo
Sea－Urchin－Fertilization
Gametes?are collected as?described. To remove some of their jelly coat,wash eggs several times in ASW by allowing them to settle and gently pouring of
Smolke:Protocols/Western
OverviewBlotting for large V5-tagged proteins in?S. cerevisiaeMaterialsY-PER (Pierce)Halt EDTA-free Protease Inhibitor (Pierce)NuPAGE Novex Bis-Tris 4
病毒冷凍保藏技術
實驗概要Snap freezing, or flash freezing, is the process by which samples are lowered to temperatures below -70°C very rapidly using dry ice or liquid
western－blotting操作手冊
Running Protein GelsSolutions10X Running Buffer (0.25 M Tris, 1.92 M glycine, 1% SDS)121 g Tris577 g glycine40 g SDSddh20 to 4 L (check pH at 1:10 dil
BioRad（伯樂）Western－Blot半干法轉膜的十大注意事項
首先講轉膜儀的清洗：Do not immerse the unit in liquid. Use special care when cleaning the anode plate to avoid scratching or marring the platinum. Do not use
Protocols－for－LCM－preparation－and－analysis
Protocols for LCM preparation and analysis?I. Preparation, LCM and RNA/DNA extraction of Frozen Tissue SectionsA.?EmbeddingB.?CuttingC.?StainingII. Pr
Immunoblotting－(Western－Blotting)
實驗概要We provide a protocol for SDS-PAGE, Protein Blotting, Immuno-Detection.主要試劑1.?0.3 M TRIZMA? base (Product No. T1503), 20% methanol.2.?0.025 M TRIZ
Metaphase－chromosome－preparation
Materials:?RPMI 1640 medium?fetal calf serum (FCS), 20%?Colcemid (e.g. Boehringer Mannheim cell biology reagents, Best.-Nr. 295892)?cell cuture flask?
Large－Scale－Plasmid－Preps:－PEG－method
1. Grow 250 - 500 mL of bacteria overnight in LB with 50 μg/mL of ampicillin.2. Transfer culture to Nalgene bottles. Spin in SGA rotor at 6000 rpm, at
Quick－Yeast－DNA－Prep:－Isolation－of－Total－DNA－(genomic－and－plasmid)
Grow a 5 ml YPD O/N culture inoculated with a single yeast colony at 30 deg.Transfer culture to a small 13 x 100 glass tube. Spin down cells 2 min. in
Arabidopsis－gDNA－isolation
This is a simple and fast protocol for the extraction of genomic DNA from Arabidopsis thaliana that works fine in PCR for simple amplicons. We only us