Bio-Rad CHEF-DR® III Variable Angle System - Manual (Page 3)

Table of Contents
Page
Section 1 General Information .....................................................................................1
1.1 Safety............................................................................................................................1
1.2 Overview......................................................................................................................1
1.3 Specifications ...............................................................................................................2
1.4
Description of Major Components ..............................................................................3
Section 2 Operation........................................................................................................5
2.1 Instrument Set-up.........................................................................................................5
2.2 Electrophoresis Chamber Operation ...........................................................................6
2.3 CHEF-DR III Operation ..............................................................................................7
Section 3 Sample Preparation.......................................................................................9
3.1 Agarose Block..............................................................................................................9
3.2 Liquid Samples ..........................................................................................................10
3.3 Preparation of Agarose Embedded Mammalian DNA .............................................10
3.4 Preparation of Agarose Embedded Bacterial DNA ..................................................11
3.5 Preparation of Agarose Embedded Yeast DNA........................................................12
3.6 Restriction Enzyme Digestion of Plugs ....................................................................13
3.7 Hemocytometer Usage ..............................................................................................13
3.8 Estimation of Agarose Plug DNA Concentration .....................................................15
Section 4 Gel Electrophoresis .....................................................................................16
4.1 Casting the Gel...........................................................................................................16
4.2 Buffer Circulation and Temperature .........................................................................17
4.3 Loading the Samples..................................................................................................18
4.4 DNA Size Standards ..................................................................................................18
4.5 Electrophoresis...........................................................................................................19
4.6
Separations at Room Temperature ............................................................................19
4.7 Removing and Staining the Gel.................................................................................20
Section 5 Applications..................................................................................................20
5.1 Strategies for Electrophoretic Separations ................................................................20
5.2 Pulsed Field Conditions by DNA Size ......................................................................22
5.3 Pulsed Field Conditions by Organism.......................................................................22
5.4 Blotting Megabase DNAs..........................................................................................23
5.5 Separation of DNA Size Standards ...........................................................................25
Section 6 Maintenance.................................................................................................27
6.1 Replacing Electrodes .................................................................................................27
6.2 Fuses...........................................................................................................................27
6.3
Maintenance of the Electrophoresis Cell...................................................................27
Section 7 Troubleshooting Guide ...............................................................................28
Section 8 References.....................................................................................................30
Section 9 Systems, Accessories and Reagents for
Pulsed Field Electrophoresis ......................................................................38