Bio-Rad CHEF-DR II System - Manual (Page 3)

Table of Contents
Page
Section 1 Introduction ..................................................................................................1
1.1 Safety ..........................................................................................................................1
1.2 Overview ....................................................................................................................1
1.3 Specifications .............................................................................................................2
1.4
Description of Major Components ............................................................................3
Section 2 Operation ......................................................................................................6
2.1 Instrument Set-up .......................................................................................................6
2.2 Electrophoresis Chamber Operation..........................................................................6
2.3 CHEF-DR II Operation ..............................................................................................7
Section 3 Sample Preparation ...................................................................................11
3.1 Agarose Block ..........................................................................................................11
3.2 Liquid Samples.........................................................................................................11
3.3 Preparation of Agarose Embedded Mammalian DNA............................................11
3.4 Preparation of Agarose Embedded Bacterial DNA.................................................12
3.5 Preparation of Agarose Embedded Yeast DNA ......................................................14
3.6 Accelerated Sample Preparation ..............................................................................15
3.7 Restriction Enzyme Digestion of Plugs ...................................................................15
3.8 Hemocytometer Usage.............................................................................................16
3.9 Estimation of Agarose Plug DNA Concentration ...................................................17
Section 4 Gel Electrophoresis ....................................................................................19
4.1 Casting the Gel .........................................................................................................19
4.2 Buffer Circulation and Temperature........................................................................20
4.3 Sample Loading........................................................................................................21
4.4 DNA Size Standards ................................................................................................22
4.5 Electrophoresis .........................................................................................................22
4.6
Separations at Room Temperature...........................................................................23
4.7 Removing and Staining the Gel ...............................................................................23
Section 5 Applications ................................................................................................23
5.1 Strategies for Electrophoretic Separations...............................................................24
5.2 Pulsed Field Conditions by DNA Size ....................................................................25
5.3 Pulsed Field Conditions by Organism .....................................................................25
5.4 Blotting Megabase DNAs ........................................................................................26
5.5 Separation of DNA Size Standards..........................................................................28
Section 6 Maintenance ...............................................................................................29
6.1 Electrode Replacement.............................................................................................29
6.2 Fuses .........................................................................................................................30
6.3
Maintenance of the Electrophoresis Cell .................................................................30
Section 7 Troubleshooting Guide ..............................................................................30
Section 8 References ...................................................................................................32
Section 9 Instruments and Reagents for Pulsed Field Electrophoresis ................38