DNA Fingerprinting and Restriction Fragment Length Polymorphism Analysis

Abstract

DNA fingerprinting and restriction fragment length polymorphism (RFLP) analyses have proven extremely useful for strain identification, epidemiological studies, and the taxonomic analysis of prokaryotic and eukaryotic organisms. Both techniques require the isolation of relatively high-molecular-weight genomic or plasmid DNA, enzymatic cleavage of the isolated nucleic acids using restriction endonucleases, and electrophoretic separation of the resulting DNA fragments. The two techniques differ with respect to the means of examining the resultant restriction fragments: DNA fingerprinting uses ethidium bromide staining and visualizes all restriction fragments, whereas RFLP analysis used DNA or RNA probes that selectively bind (hybridize) to a few restriction fragments. In either case, the resulting banding patterns are generally unique to one or a few strains of a particular microbe and as such, can serve as a "fingerprint" for strain identification. Figure 31-1 shows the general scheme for DNA fingerprinting and RFLP analysis of bacterial genomic DNA. While DNA fingerprinting is relatively rapid, routine, and inexpensive to perform, RFLP analysis is more complex, expensive, and time consuming. However, RFLP analysis can show small differences between the genomic DNAs of organisms that is not evidenced by DNA fingerprinting techniques. In addition, RFLP analyses can also be useful for the construction of genetic maps and for map-based cloning in eukaryotic organisms (Young, 1990). More recently, it has been shown that DNA primers corresponding to repetitive extragenic palindromic (REP) and enterobacterial repetitive intergenic consensus (ERIC) sequences,coupled with the polymerase chain reaction (peR) technique can be used to fingerprint the genomes of a large number of different gram-negative soil bacteria (de Bruijn, 1992; Hulton et aI., 1991; Judd et aI., 1993; Stem et aI., 1984; Versalovic et aI., 1991).