GrowingOvernightCultures
1. Place 2 mL of the appropriate sterile medium in a 13 mm yellow-capped culture tube. If more culture is needed, place up to 5 mL in a 16 mm green-capped culture tube. Large amount of culture should be made in growth flasks, using up to 25 flask volume in medium (e.g., no more than 30 mL in a 125 mL flask, or 250 mL in a 1000 mL flask).2. Touch a single colony with the end of an applicator stick and hold it in your ......阅读全文
General－Cloning－Protocols
Large Scale Preps: (See Large scale plsasmid prep protocol for more details)Cultures: Inoculate a 5 mL LB/Amp (50 - 100 µg/mL) culture in early a.m. w
粘粒
Preparation of Cosmid DNAPreparation of Cosmid DNA from 50 ml Cultures (Donis Keller Lab)Cosmid vectors containing foreign DNA inserts are known to re
粘粒
Preparation of Cosmid DNAPreparation of Cosmid DNA from 50 ml Cultures (Donis Keller Lab)Cosmid vectors containing foreign DNA inserts are known to re
Modified－Yeast－Transformation
Inoculate cells from an overnight culture into 50 ml YEPD and incubate at 30°C with shaking. Typically, add 0.1 to 0.2 ml saturated culture in the eve
Transformation－of－Magnaporthe－grisea－to－phosphinothricin－resistance
Three transformation systems have been reported for the rice blast fungus Magnaporthe grisea (Parsons et al. 1987 Proc. Natl. Acad. Sci. USA 84:4161-4
Tissue－Culture－Methods2
IV. MAINTENANCECultures should be examined daily, observing the morphology, the color of the medium and the density of the cells. A tissue culture log
MITOMYCIN－C－TREATMENT－OF－PMEFs
Cultures to be treated should be sub confluent ie actively growing.1. Add 1/20 volume Mitomycin C (200 ug/ml 10 ug/ml), to culture and incubate at 37
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
Identification－and－expansion－of－the－tumorigenic－lung－cancer－stem－cell－...
Identification and expansion of the tumorigenic lung cancer stem cell populationLung cancer contains a rare population of CD133+ cancer stem-like ce
Agrobacterium－growth－and－transformation
Growth and storage of Agrobacterium tumefaciensStrain GV3101: resistant to gentamycin and rifampicin so add 25-50 ug/ml Gentamycin, 10 ug/ ml rifampic
Heterogeneity－of－SingleCell－Gene－Expression－Across－Phenotypically（一）
Introduction Multi-cellular populations are fundamentally driven by the collective properties of individual cells. However, our understanding of ge
Primary－type－II－pneumonocyte－Isolation－and－culture
Pulmonary alveolar type II cells carry out highly specialized functions that include the synthesis, secretion, and reutilization of surfactant, a
GST融合蛋白纯化方法
Abstract: Many people have vented out frustration over insoluble GST-fused proteins. This is a protocol for enzymatically active soluble GST-fused pro
BAC－DNA分离方法－Isolation－of－BAC－DNA－from－Largescale－Cultures
Isolation of BAC DNA from Large-scale CulturesJoseph SambrookPeter Maccallum Cancer Institute and The University of Melbourne, AustraliaDavid W. Russe
Bacterial－cell－culture
MaterialsGlass culture tubes with metal caps and labelsGrowth medium, from media room or customizedGlass pipette tubesParafilmEquipmentVortexerFireboy
Method:－Cell－Counts－Using－a－Hemacytometer
Method: Cell Counts Using a HemacytometerJune 1, 1990Rosalie VeilePurpose:The purpose of this procedure is to determine the cell density of the cultur
酵母转化的几种方法
Modified Yeast Transformation Inoculate cells from an overnight culture into 50 ml YEPD and incubate at 30°C with shaking. Typically, add 0.1 to 0.2 m
Yeast－Cell－Cycle－by－Flow－Cytometry
ReagentsCold absolute ethanol.0.5 M Na citrate stock (filtered), 50mM diluted stock.10 mg/ml RNase A (Boil 10 mins, cool, filter and store at -20°C).4
ELECTROPORATION－OF－ES－CELLS－AND－ISOLATION－OF－H/R－CLONES
Need 1.5-2 x 107 cells from a 2 day culture.1. Cells are harvested as normal, washed x 1 in PBS then taken up at conc. of 1.2 x 10 7 cells/ml in cold
原代神经细胞培养方法－Neuron－Cell－Culture
1. Preparation of coverslips1.1- Mass cultureOur standard mass cultures are plated on astrocytes. Those, in turn, are plated on glass coverslips pre-
酵母GST蛋白纯化方法
GST Fusion Protein Purification from Yeast5 ml overnight culture of your favorite yeast in your favorite medium.Inoculate 50 ml and grow 30o C shaking
TISSUE－CULTURE－ON－COVERSLIPS
I. Purpose:A. Skin tissue may be used for chromosome analysis in special cases when the results from peripheral blood are inconclusive, e.g. suspected
全细胞靶点筛选抗生素新药的方法
A target-specific whole cell assay for antibacterial drug discoveryLorraine HernandezSrinivas KodaliDoris CullySheo SinghJun Wang , jun_wang2@merck.co
TOP10－chemically－competent－cells
OverviewThis protocol is a variant of the Hanahan protocol [1] using CCMB80 buffer for DH10B, TOP10 and MachI strains. It builds on Example 2 of the B
Subculture－of－Adherent－Cell－Lines
AimAdherent cell lines will grow in vitro until they have covered the surface area available or the medium is depleted of nutrients. At this point the
Aseptic－Technique－and－Good－Cell－Culture－Practice
AimTo ensure all cell culture procedures are performed to a standard that will prevent contamination from bacteria, fungi and mycoplasma and cross con
Midiprep－preparation－of－Plasmid－DNA
实验概要The PureLink™ HiPure Plasmid DNA Midiprep Kit allows purification of 100–350 μg of high-quality plasmid DNA from 15–25 mL overnight E. coli cul
Largescale－Immunocytology
This protocol describes our method for preparing cells for immunofluorescence, in which all incubations and washes are performed in microtiter dishes.
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
Human－thyroid－endothelial－cell－isolation
1. Human thyroid glands were obtained at surgery from multinodular goiters.2. The tissue was incubated in 0.1% type II collagenase (Worthington) overn