LargeScalePlasmid,Cosmid,BAC,PAC,andFosmidDNAIsolation
DNA Isolation by a Cleared Lysate Method Followed by Double Acetate Precipitation Version 3b - updated September 26, 1999The Most Recent Roe Lab Implementation (by Feng Chen, Hau-Qin Pan and Fu Ying) Adapted from the Genome Sequencing Center, Washington Univ. ProtocolA smear of colonies of cosmid/BAC/PAC were picked and transferred into a 12X75 mm Falcon tube containing 3 ml of LB medium (10 g Bacto-Tryptone, 5 g Bac......阅读全文
Isolation－and－growth－of－mouse－primary－myoblasts
Isolation of limb muscle from neonatal mice1. Neonatal mice by decapitation or CO2 inhalation.2. Rinse the limbs with 70% ethanol and remove them wi
Isolation－and－culture－of－porcine－muller－cells
1. Porcine eyes were harvested from pentobarbital-anesthetized animals, each weighing 30 to 40 pounds, and transported to the laboratory in ice-co
Isolation－of－human－endometrial－epithelial－cells
Tissue collection1. Endometrial biopsies were collected from women undergoing gynaecological procedures for benign conditions. 2. All women reported
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
Isolation－of－Nonmuscle－Actin
General Preparation1. Prepare buffers and have them cold. Make 50X stock of Buffer G and dilute as needed from frozen aliquots of 50X Buffer G.1X Extr
Isolation－of－human－atrial－or－ventricular－cells
1. Human tissue was derived from atrial or ventricular biopsy specimens belonging to patients undergoing heart surgery.2. Isolated myocardial tissue w
Isolation－of－dermal－mast－cell－(DMC)
1. Human DMC were isolated from skin by enzymatic digestion.2. Dermal tissue was obtained from patients with skin after informed consent was given.3.
Isolation－and－culture－of－smooth－muscle－cell
1.Male Wistar rats of about 250g body wt were anesthetized with ether and were decapitated. 2.Kidneys and thoracic aorta were removed, rinsed with
Nucleolar－Isolation－Protocol2
7. Resuspend the pellet with 3 ml S1 solution (Figure 3). The pellet should be resuspended readily by pipetting up and down. A pellet that cannot be r
Isolation－of－microRNA－(miRNA)
实验概要 This protocol utilizes the powerful guanidine isothiocyanate–phenol:chloroform extraction method which allows the rapid isolation of t
ISOLATION－OF－PRIMARY－MOUSE－EMBRYO－FIBROBLASTS
You will need:13.5 day pregnant mouse (we use MTK NEO inbred white mice)2 sets sterile instrumentsone containing a pair of curved forceps and a pair o
Primary－cardiac－fibroblast－and－cardiomyocyte－isolation
1. Ventricles were removed under sterile conditions. 2. Ventricles were placed in cold sterile primary cell culture medium, minced into approximat
细胞分离技术（Cell－Isolation）
一、离心技术离心是研究如细胞核、线粒体、高尔基体、溶酶体和微体，以及各种大分子基本手段。一般认为，转速为10~25Kr/min的离心机称为高速离心机；转速超过25Kr/min，离心力大于89Kｇ者称为超速离心机。目前超速离心机的最高转速可达100Kr/min，离心力超过500Kg。（一）、差速离心（
Isolation－and－culture－of－pancreatic－stellate－cells
Pancreatic stellate cells (PaSCs or PSCs) are myofibroblast-like cells found in the areas of the pancreas that have exocrine function. PaSCs are
Isolation－of－kidney－glomeruli－from－mice
Isolation of mice glomeruli1. Mice were anesthetized by an intraperitoneal injection of Avertin (2,2,2-tribromoethyl and tertiary amyl alcohol; 17
Human－B－cell－isolation－and－culture
实验概要This protocol provides a general protocol for human B cell isolation and culture.实验步骤B cell isolation1. Donor blood was obtained with informed con
Isolation－of－rat－cardiac－fibroblasts－and－cardiomyocytes
1. Hearts were removed from newborn rats (day 0), put into calcium- and bicarbonate-free HEPES-buffered Hanks’ medium, cut into pieces and digeste
Isolation－of－human－prostatic－epithelial－cells
1. A small piece of tissue from each specimen was removed and minced. 2. The tissue was digested with collagenase overnight. 3. To remove the collagen
Follicular－thyroid－cell－isolation－and－culture
Thyroid epithelial cells (also called follicular cells or principal cells) are cells of thyroid gland that are responsible for the production and
Twohybrid－analysis－of－genetic－regulatory－networks2
2.2 Interaction mating - large scaleWith a few modifications, the procedure described above can be used to test for interactions between a single prey
Intestinal－intraepithelial－lymphocytes
Intestinal intraepithelial lymphocytes Intestinal intraepithelial lymphocytes (IELs) are mostly T cells, which are continuously associated with gut
酵母人工染色体
· Easy YAC Preparation Method (Andrew Davies,Shaw lab)· Screening YAC libraries (Donis Keller Lab)This is a method for screening YAC l
A－novel－method－of－growing－fungi－for－DNA－extraction
Preparation of fungi for DNA extraction typically involves growing cultures in liquid culture in Erlenmeyer flasks, Roux bottles or even microfuge tub
Ligation－Optimization
The following protocol can be used to optimize ligation conditions for difficult to clone (e.g. very large) fragments. The principle is to independent
SOUTHERN－BLOTTING
Materials:Whatman 3 mm Blotting Papernitrocellulose (Schleicher & Schuell, Amersham) or nylon membrane filter (Amersham).Paper towels (preferably C-fo
Performing－a－hunt－by－interaction－mating
AbstractWhen more than one bait will be used to screen a single library, significant time and resources can be saved by performing the interactor hunt
Multicolour－3DFISH－in－vertebrate－cells1
IntroductionMulticolour 3D-FISH in combination with confocal microscopy, 3D image reconstruction and quantitative image analysis is an efficient tool
TritonPrep－Method－for－bacterial－DNA－Purification
Triton-Prep Method for bacterial DNA PurificationGrow 5 ( large scale-15ml culture). Harvest in single eppendorf tube (or 15 ml disposable tube).Resus
An－Ultrafast－method－of－DNA－extraction－from－Neurospora
We have found that the DNA extraction procedure of Metzenberg and Baitch (Neurospora Newsl. 28:20)/Stevens and Metzenberg (Neurospora Newsl. 29:27) wh
基因工程的载体3
⑷基因组成 lDNA至少包括61个基因，大多基因按功能相似性成簇排列，其中一部分为噬菌体生命活动的必须基因，另一部分约1／3为非必须区段。 3. l噬菌体载体的类型 插入型 （Insertion vectors ）