Single Nucleotide Polymorphism
8.3 POLYMORPHISMS DETECTED BY PCR
Without a doubt, the polymerase chain reaction (PCR) represents the single most important technique in the field of molecular biology today. What PCR accomplishes in technical terms can be described very simply — it allows the rapid and unlimited amplification of specific nucleic acid sequences that may be present at very low concentrations in very complex mixtures. Within less than a decade after its initial development, it has become a critical tool for all practicing molecular biologists, and it has served to bring molecular biology into the practice of many other fields in the biomedical sciences and beyond. The reasons are several fold. First, PCR provides the ultimate in sensitivity — single DNA molecules can be detected and analyzed for sequence content (Li et al., 1988; Arnheim et al., 1990). Second, it provides the ultimate in resolution — all polymorphisms, from single base changes to large rearrangements, can be distinguished by an appropriate PCR-based assay. Third, it is extremely rapid — for many applications, it is possible to go from crude tissue samples to results within the confines of a single workday. Finally, the technique is an agent of democracy — once the sequences of the pair of oligonucleotides that define a particular PCR reaction are published, anyone anywhere with the funds to buy the oligonucleotides can reproduce the same reaction on samples of his or her choosing; this stands in contrast to RFLP analyses in which investigators are often dependent upon the generosity of others to provide clones to be used as probes. Numerous books and thousands of journal articles have been published on the principles and applications of the technique [Erlich (1989) and Innis et al. (1990) are two early examples].
Although the applications of PCR are as varied as the laboratories in which the technique is practiced, this section will focus entirely on six general applications that are relevant to the detection and
Without a doubt, the polymerase chain reaction (PCR) represents the single most important technique in the field of molecular biology today. What PCR accomplishes in technical terms can be described very simply — it allows the rapid and unlimited amplification of specific nucleic acid sequences that may be present at very low concentrations in very complex mixtures. Within less than a decade after its initial development, it has become a critical tool for all practicing molecular biologists, and it has served to bring molecular biology into the practice of many other fields in the biomedical sciences and beyond. The reasons are several fold. First, PCR provides the ultimate in sensitivity — single DNA molecules can be detected and analyzed for sequence content (Li et al., 1988; Arnheim et al., 1990). Second, it provides the ultimate in resolution — all polymorphisms, from single base changes to large rearrangements, can be distinguished by an appropriate PCR-based assay. Third, it is extremely rapid — for many applications, it is possible to go from crude tissue samples to results within the confines of a single workday. Finally, the technique is an agent of democracy — once the sequences of the pair of oligonucleotides that define a particular PCR reaction are published, anyone anywhere with the funds to buy the oligonucleotides can reproduce the same reaction on samples of his or her choosing; this stands in contrast to RFLP analyses in which investigators are often dependent upon the generosity of others to provide clones to be used as probes. Numerous books and thousands of journal articles have been published on the principles and applications of the technique [Erlich (1989) and Innis et al. (1990) are two early examples].
Although the applications of PCR are as varied as the laboratories in which the technique is practiced, this section will focus entirely on six general applications that are relevant to the detection and