Chapter 2
Enzymatic Modification of DNA and RNA
KAI-OLAF NETZER
lntroduction
Enzymes represent the tools of the molecular biologist. They serve to manipulate DNA and RNA in vitro. Today, more than 200 different enzyrnes
with various uses areaffered by commercial manufacturers. Figure 1 shows
a rough classification of enzyrnes by their principal actions. This chapter
will provide examples from each group and protocols for their use.
Subprotocol 1
Restrietion Endonucleases
The first observations that bacteria such as E. coli contain site-specific DNA
degrading enzyrnes date back to the late 1960s. The so-called restriction
endonucleases (or restriction enzyrnes) are part of a system that protects
resident DNA against contamination by sequences offoreign origin. None
of the early restriction enzyrnes lived up to the hopes of potentiallaboratory
use. They recognized the DNA at specific sites, but cleaved at random sites
far removed. Today, restriction enzymes have been isolated that fall into
three different groups according to their action (Figure 2). Both restriction
enzyrnes with site specific recognition and cleavage are routinely used in
molecular biology.
Kai-OlafNetzer, Krankenhaus Köln-Merheim, Medizinische Klinik I, Ostmerheimer Str.
200, Köln, 51109, Germany (phone +49-221-8907-2200;fax +49-221-8907-2335; e-mail
kai.netzer@uni-koeln.de)
Enzymatic Modification of DNA and RNA
KAI-OLAF NETZER
lntroduction
Enzymes represent the tools of the molecular biologist. They serve to manipulate DNA and RNA in vitro. Today, more than 200 different enzyrnes
with various uses areaffered by commercial manufacturers. Figure 1 shows
a rough classification of enzyrnes by their principal actions. This chapter
will provide examples from each group and protocols for their use.
Subprotocol 1
Restrietion Endonucleases
The first observations that bacteria such as E. coli contain site-specific DNA
degrading enzyrnes date back to the late 1960s. The so-called restriction
endonucleases (or restriction enzyrnes) are part of a system that protects
resident DNA against contamination by sequences offoreign origin. None
of the early restriction enzyrnes lived up to the hopes of potentiallaboratory
use. They recognized the DNA at specific sites, but cleaved at random sites
far removed. Today, restriction enzymes have been isolated that fall into
three different groups according to their action (Figure 2). Both restriction
enzyrnes with site specific recognition and cleavage are routinely used in
molecular biology.
Kai-OlafNetzer, Krankenhaus Köln-Merheim, Medizinische Klinik I, Ostmerheimer Str.
200, Köln, 51109, Germany (phone +49-221-8907-2200;fax +49-221-8907-2335; e-mail
kai.netzer@uni-koeln.de)