DNA Fragment Analysis

Before a gene can be manipulated using gene technology, it first has to be isolated from the entire genome. Trying to study a particular gene or groups of genes presents a challenge. In eukaryotes, a DNA molecule is very long, so it usually carries thousands of genes, which are surrounded by thousands more noncoding nucleotide sequences. The noncoding nucleotide sequences are stretches of DNA that do not translate genetic information into proteins. The human genome, for example, carries between 20,000 and 25,000 genes. This makes finding any specific gene very difficult. The time frames for DNA analysis may be lengthy. The first step, polymerase chain reaction (PCR), can typically take up to a couple of hours. Once PCR is completed, the next step, gel electrophoresis, separates the DNA fragments into different sizes.The DNA of interest can then be isolated if its size is known.

DNA evidence is extremely useful in placing certain individuals at crime scenes because each individual has a unique DNA sequence, and certain parts of the genome are highly variable among individuals. DNA evidence collected from a crime scene can be exposed to restriction enzymes, which cut DNA at a specific sequence and are designed to target some of those highly variable regions of DNA. The resulting mixture of DNA fragments is then subjected to gel electrophoresis, a technique that separates DNA and other molecules based on their size using electric charge. This technique separates the DNA into a unique pattern of bands visible in agarose or polyacrylamide, which are two of a number of gelatinous substrates used for growing biological specimens. The technique employs an electrical field to separate molecules based on their size. The same person's DNA yields the same pattern every time that a particular restriction enzyme is used. These results undergo a full analysis, where comparisons are made to at least 13 short tandem repeat (STR) locations. This takes a large amount of time. Similar technology is employed in cases of questioned parentage. A sample of the mixture of DNA fragments is added to wells in one end of an agarose sheet that sits in an electrolyte solution between two electrodes. When electric current is passed through the bath, the DNA fragments move through the gel toward the positive end of the chamber because DNA carries a negative charge. Smaller pieces of DNA are able to move through the gel more easily because they have less resistance than large pieces. After some time, separate bands appear on the gel, showing where the small pieces of DNA have moved farther ahead and longer fragments have lagged behind. The DNA of multiple sources can be put through the same agar-gel sheet for easy comparison. If the bands match, the conclusion is that the DNA sample came from the same person. Partially matching bands may indicate a relationship between two people, such as parent-child, grandparent-child, or more distant relationships. A relationship is based on the presence of an allele, or version of a gene, that appears in the genome of a relative. For example, for the alleles responsible for hair color, it is possible that they may not be expressed for a generation or more and may appear in later progeny, such as in a grandchild or great-grandchild.