Polymerase Chain Reaction (PCR) is a fundamental technique in molecular biology. It creates millions of copies of a specific DNA segment from a small initial sample. Understanding “highly variable” in PCR is key to its diverse applications.
Genetic Variation Basics
Deoxyribonucleic acid (DNA) is the blueprint for life, carrying genetic instructions for an organism’s growth, development, and reproduction. Genes are specific DNA segments containing instructions for building proteins or functional RNA molecules, responsible for an organism’s traits.
While DNA’s fundamental structure is consistent across individuals of the same species, slight differences exist in the sequence of its building blocks, called nucleotides. These variations are known as genetic variation. Different versions of the same gene are called alleles, contributing to individual diversity.
Many variations are single nucleotide polymorphisms (SNPs), where a single nucleotide differs between individuals. Other forms of genetic variation involve insertions, deletions, or duplications of larger DNA segments. This inherent variability distinguishes one individual from another, even within the same family or species.
Identifying Highly Variable Regions
Highly variable regions are specific DNA sequences within the genome that show a greater degree of difference among individuals compared to more conserved regions. These segments change frequently, making them exceptionally useful for distinguishing between individuals or even closely related species.
One example of highly variable regions includes Short Tandem Repeats (STRs), also known as microsatellites. STRs are short sequences of 2 to 6 nucleotides repeated multiple times. The number of repeats varies considerably from person to person. For example, one individual might have 10 repeats of a specific sequence, while another might have 15 repeats at the same location.
Another type is Variable Number Tandem Repeats (VNTRs), or minisatellites, similar to STRs but with longer repeat units (10 to 60 nucleotides). The variability in both STRs and VNTRs stems from the differing number of repeat units at specific chromosomal locations. These differences make these regions highly distinctive, acting like genetic barcodes unique to each individual.
Utilizing Variability in PCR
PCR specifically leverages these highly variable regions for various applications by targeting flanking DNA sequences. Primers, short synthetic DNA strands, are meticulously designed to bind to stable, non-variable DNA sequences adjacent to a highly variable region. This precise binding ensures only the desired variable segment is amplified.
Once primers bind, PCR makes millions of copies of the specific highly variable region. The significance of amplified products lies in their length. Because the number of repeat units within an STR or VNTR varies between individuals, the length of the amplified DNA fragment also varies. If one person has more repeats at a given STR locus, their amplified PCR product will be longer than someone with fewer repeats.
Analyzing these different lengths, typically through gel electrophoresis or capillary electrophoresis, allows scientists to differentiate individuals. This method is crucial in forensic DNA profiling, matching DNA from a crime scene to a suspect based on the unique pattern of their STR lengths across multiple loci. Paternity testing relies on comparing the STR profiles of a child and potential father to establish biological relationships. The analysis of these regions via PCR also plays a role in human identification and distinguishing between species.