IGEM:IMPERIAL/2007/Tutorials/Guide for Engineers/Standard Lab Techniques

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Contents

Introduction

The Molecular Biologist Toolbox Kit

Restriction Enzymes

Restriction enzymes are a class of enzyme that can causes specific breaks in a DNA molecule.The mechanism by which restriction enzymes cut DNA is by causing hydrolysis of the phosphodiester bonds between the nucleotides that make up DNA.
Restriction enzymes can be broadly split into two types:

  • Exonucleases cleave DNA from the ends of the DNA molecule. The action of exonucleases will remove nucleotides from the ends of DNA.
  • Endonucleases cleave within the interior of the DNA molecule. The action of endonucleases will cause fragmentation of DNA molecules.

Endonucleases

Some examples of restriction sites
Some examples of restriction sites

Endonucleases are essential for recombinant DNA technology because they allow specific genes to be cut out and released from within a DNA molecule. The specificity of endonucleases is determined by a specific sequence of nucleotides that they can recognise and cleave. This sequence of nucleotides is called the restriction site and will vary between different types of endonucleases.

DNA polymerase

Vectors and Plasmids

Gene Cloning

Gel Electrophoresis

Blotting Techniques

Western Blotting

Southern Blotting

Northern Blotting

PCR (Polymerase Chain Reaction)

DNA Sequencing

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