PreparingAntibioticsStockSolutionandAmpicillinAgarPlates
AMPICILLIN Beta-lactam-antibiotics are not very stable when dissolved. Slow but steady degradation happens even when frozen to -20°C. Therefore commercial beta-lactam-antibiotics are sold as dried powder and reconstituted just before use! If possible adjust stock solution pH to stability optimum and choose suitable storage temperature! Due to enhanced solubility Ampicillin-Na is used for cell culture instead of......阅读全文
Preparing－Antibiotics－Stock－Solution－and－Ampicillin－Agar－Plates
AMPICILLIN Beta-lactam-antibiotics are not very stable when dissolved. Slow but steady degradation happens even when frozen to -20°C. Therefore comme
Preparation－of－Agar－plates
Prepare media and add 1.5 agar before autoclaving it (15g per liter). After autoclavation, cool the media in a 55 degree waterbath. Do not allow the s
Agar－Plates－for－Selection－of－Clones－in－Bacteria
Cloning of PCR productsStocks:LB Agar: Luria Broth after Lennox:per LiterTryptone10 gYeast Extract5 gSodium Chloride5 gBact. Agar15 g pH 7.0Autoclave
PREPARATION－OF－2％－FORMALDEHYDE－STOCK－SOLUTION－(2－METHODS)
METHOD 1:Formaldehyde preservative – 2% formaldehyde solution in protein-free phosphate-buffered saline (PBS).Prepare as follows:Add 2 g paraformaldeh
Expression－Library－Screening－(Procaryotic)－Using－APFusion－Proteins
Outline:Bacteriophage lambda is a linear double stranded DNA, approximately 50 kB in size. The two sticky ends help the phage to recircularize after e
Bacterial－Media－Solutions－and－Stocks
3 agar (200 ml)Add 6 grams agar to 200 ml deionized water. Autoclave to sterilize.1.6 agar (200 ml)Add 3.2 grams agar to 200 ml deionized water.Autocl
Protein－Expression－and－Purification－Protocol
Step 1: Transform appropriate DNA plasmid into BL21(DE3) E. coli cells. These cells must be competent. (Protocol for how to make competent cells.)a) T
Maintenance－of－Probes－in－bacteria－including－Escherichia－coli
Plasmid (pUC series) containing genomic DNA fragments are maintained in E. coli strain DH5aTM. The E. coli cultures are routinely cultured at 37 C on
Antibiotic－concentrations－for－E.－coli－selection
Antibiotic ConcentrationsThe following list shows recommended antibiotic concentrations for LB media or agar plates.AntibioticConcentrationAmpicillin1
基本实验技术
I. Safety ProceduresA. ChemicalsA number of chemicals used in this laboratory are hazardous. All manufacturers of hazardous materials are required by
Preparing－Lambda－DNA
Preparing Lambda DNA1- Coliphage lambda DNA is a widely used vector for recombinant DNA. The middle third of its 48,000 bp contains no genes required
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
Streptomyces:Protocols/Spore－Prep
Spore Prep - Inoculating & HarvestingDescription A spore prep is a method of preserving a sporulating strain of Streptomyces. The stock is stored in 2
McKinney:TransformByElectroporation
Materials50mL 7H9 mycobacterial medium + 3mL per transformation102mL 10% glycerol (possibly a few mL more if you are doing many transformations)400mL
Preparation－of－Bacterial－Proteins－for－Analysis－by－2DPAGE
The following protocol has been developed for preparing soluble bacterial proteins in a form suitable for analysis by 2D-PAGE. The procedure was princ
Phage－Titer
IntroductionLambda phage is a commonly used vector for transgenes. Its very high rate of infectivity makes it ideal for creating large numbers of clon
Expression－Protocol－in－M9－Minimal－Media－via－T7－Promoter
The following protocol has been used successfully to 15N or 13C/15N label our proteins using our pET1120/BL21(DE3) expression system: Preparing M9 min
Expression－Protocol－in－M9－Minimal－Media－via－T7－Promoter
Expression Protocol in M9 Minimal Media via T7 PromoterThe following protocol has been used successfully to 15N or 13C/15N label our proteins using ou
Inoue法制备大肠杆菌超级感受态细胞
实验步骤： 1、Inoculate from an overnight grown in LB.从培养过夜的LB平板上挑取单菌落 。2、Grow in 250 ml "SOB" at 18℃ until OD600 = 0.6.(0.3)接种于250ml SOB，18度培养至OD＝0.6。3、On
Easy－Way－to－Clone－Genes－From－a－Phage－Library
Easy Way to Clone The protocol is oriented towards a C. albicans genomic library I made in Lambda Zap II on 7/97.The overall sequence of events is: •
cDNA－LIBRARY－SCREENING
PREPARE SOLUTIONS1. 10mM MgSO4, 0.2% Maltose LB (100 mL):Mix 1.0 g of Bacto-Tryptone, 1.0 g of NaCl, 0.5 g of Yeast Extract, and 1.0 mL of 1M MgSO4. A
Hints－and－precautions－for－the－care,－feeding－and－breeding－of－Neurospora2
9. Spontaneous germination. Some genotypes result in spontaneous ascospore germination (for example, the unpigmented ascospores of per-1 Type I). A si
Expression－L19－using－Pichia－pastoris
Pichia pastoris is a methylotropic yeast used to express high amounts of protein. Secreted expression is achieved with a cleavalable faktor. To yield
Preparation－of－Broth－and－Plates,－etc.
Recipes: 1) LB BrothMake 16 gm of LB Broth Base (Gibco #M27800C) up to 800 ml in ddH2O. Swirl to dissolve, then add 110 µl of 10 N NaOH. Autoclave. 2
Yeast－Media,－Solutions－and－Stocks
Yeast Media:Note: Synthetic complete medium can be prepared by adding media supplements (see below).Medium using 6.7 g yeast nitrogen base without ami
Dropout－plates－for－yeast
Materials(Solutions are all available from the media room)200ml bottle of 2x SD200ml bottle of 4% agar -- make sure to sign it out40% glucoseCSM minus
Preparing－Overnight－Bacterial－Culture
Materials:Sterile LB medium (Luria-Bertani Medium) with or without antibiotic:water 500 mlbacto-tryptone 10 gbacto-yeast extracts 5 gsodium chloride 1
常用试剂和培养基
Sterile water bottles200 ml, 500 ml, or 950 ml water. Autoclave.0.05 M CaCl2 (per 200 ml)CaCl2·2 H2O 1.5 g Add 198 ml water.Autoclave.0.5 M CaCl2 (p
Colony－Hybridization
ProcedurePrepare serial 10-fold dilutions of transformed bacteria in LB and spread 100 µL onto LB/Amp plates, as described in the Electroporation prot
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