AN INTRODUCTION TO TEE USE OF GENE PROBES IN THE BACTERIOLOGICAL ANALYSIS OF WATER A special report produced for DoE to support Contract ET 9418 DoE 2357-M
Report No DWI0029

Dec 1989

SUMMARY

I OBJECTIVES

To provide an introduction to the techniques and theory currently used in developing gene probes to detect and quantify total coliforms or Escherichia coli in water.

II REASONS

This report has been prepared by WRc at the request of DoE to aid in understanding and provide background knowledge to work being carried out at the University of Leicester in connection with DoE contract "Development of Gene Probes For Coliform Bacteria" (PECD 7/7/302).

III RESUME OF CONTENTS

To understand the mechanism by which gene probes may be deployed to detect specific bacteria in a water sample requires some background knowledge of molecular biology. This is introduced in Section 2. In particular the structure of DNA, which is the genetic material, is described along with how the sequence of bases contains the genetic information. In Section 3, the structure of gene probes and how they recognise and bind to their specific target sites within the bacterial genetic material is discussed. The strategy to develop gene probes for the bacterial analysis of water is outlined in Section 4. To develop a gene probe that is specific for a particular species (eg E. coli) or group (eg total coliforms) of bacteria requires the identification of sequences within the genetic material that are unique to that species or group. Gene probes designed to target these sequences will thus detect that particular species or group of bacteria. Two potential target sites for coliform or E. coli specific gene probes are the lac operon and the 16S rRNA. The advantages and disadvantages of these two target sites are summarised. One disadvantage of using gene probes is the sensitivity of the detection method. In Section 5, a method, called the polymerase chain reaction, which may overcome the problem is introduced. Much of the research and development of gene probes requires techniques in genetic engineering. Some of these, in particular cloning and sequencing of DNA fragments are presented in the Appendices.

Copies of this report may be available as an Acrobat pdf download under the 'Pre 2000 Reports' heading on the DWI website.