Thermo Scientific Shake 'n Stack 6244, Operating And Maintenance Manual

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© Thermo
Scientific
, May 2003. Issue 7 i
HYBRIDISATION GUIDE
Contents
CHAPTER 1: INTRODUCTION............................................................................. 1
CHAPTER 2: NUCLEIC ACID TRANSFER PROCEDURES ................................ 3
Dot/Slot Blotting............................................................................... 3
Colony/Plaque Blotting .................................................................... 4
Plaque Blotting ................................................................................ 5
Southern Blotting............................................................................. 5
Capillary Blotting ............................................................................. 6
Figure 2.1: Capillary Blot Assembly ............................................ 7
Northern Blotting ........................................................................................ 8
CHAPTER 3: HYBRIDISATION PROCEDURES .................................................. 9
Southern Blot DNA Hybridisations................................................... 9
Northern Blot RNA Hybridisation ..................................................... 11
Notes for Nucleic Acid Hybridisations using the Thermo Range of
Equipment ....................................................................................... 12
Placing Membranes in a Bottle........................................................ 12
Figure 3: Inserting Membranes in a Bottle .................................. 14
Prehybridisation............................................................................... 15
Hybridisation.................................................................................... 15
Washing .......................................................................................... 16
CHAPTER 4: NUCLEIC ACID PROBES............................................................... 18
Double-Stranded DNA Probes ........................................................ 18
Single-Stranded
DNA Probes.......................................................... 18
RNA Probes .................................................................................... 19
Synthetic Oligonucleotides .............................................................. 19
CHAPTER 5: NON-RADIOACTIVE SYSTEM………………………………………..20
The DIG System in Thermo Hybridisation Ovens............................ 20
Electrophoresis & Transfer.............................................................. 21
DNA Hybridisation ........................................................................... 21
CHAPTER 6: HYBRIDISATION USING OLIGONUCLEOTIDE PROBES ............ 23