Molecular Genetic Techniques

Genome Sequencing

DNA sequencing techniques can reveal nucleotide sequences of an organism's genome.

DNA sequencing is the process by which the precise order of nucleotides within DNA is determined. Nucleotide bases pair according to specific rules: adenine (A) always pairs with thymine (T) or uracil (U), and cytosine (C) always pairs with guanine (G), which is called complementary base pairing. Understanding the complementary base pairing rule of nucleic acids enables scientists to sequence an organism's entire set of DNA, or genome, but this pairing rule (which is also fundamental to PCR) is only a small part of the process of sequencing DNA.

To sequence an organism's genome, geneticists developed a process called dideoxy sequencing, which is a method used to obtain the nucleotide sequence of a strand of DNA. The process involves making many copies of a target DNA region and uses materials similar to those needed for DNA replication. These include a DNA polymerase enzyme, a primer, four DNA nucleotides, and the template strand of DNA to be sequenced. In addition, dideoxy sequencing uses special chain-terminating versions of the four DNA nucleotides, each of which is labeled with a dye of a different color to identify where each DNA chain ends. The modified nucleotides can be detected during or after binding. By recording the order of the nucleotides that are bound to the unknown DNA strand, researchers can deduce the nucleotide sequence of the original piece of DNA. Shortly after sequencing the first virus in 1976, a species of yeast became the first eukaryotic genome to be sequenced. A eukaryotic organism is one with a nucleus and whose DNA is bound together by proteins.

Initially, the process of sequencing was labor-intensive because much of the work of scientists had to be done by hand and thus was time-consuming. However, many advances in sequencing techniques have emerged, including the use of technology to sequence thousands of pieces of DNA at the same time. As the sequencing process becomes faster and cheaper, large quantities of genetic information are being collected and made available to researchers. Keeping track and making sense of vast quantities of sequence data is one practical application of the field of study known as bioinformatics, which uses computers and mathematics to store and analyze biological data. These data have countless uses, including finding genetic markers for diseases, determining patterns that might not otherwise be visible or clear, and testing for genetic diseases to improve and individualize medical treatments.

While current techniques of gene sequencing are efficient, new techniques are frequently being developed to improve the process. One newly developed technique involves viewing DNA using a very fast movie camera attached to a microscope. This technique adds a colored dye to the DNA strand to highlight each of the four nitrogen bases (A, T, C, G). This enables the sequences to be seen at a higher resolution and identified more easily.

Improvements to genetic sequencing techniques have broad applications in the field of preventive medicine. Long pieces of DNA, upward of one million bases or more, can be sequenced quite rapidly for relatively little money. Knowing a person's genetic code can provide a lot of information about how genetic diseases may be transmitted to offspring and how those diseases may be expressed or influenced by the environment. This has shown particular utility in cancer research, sometimes enabling doctors to be able to use genetic information to identify the particular type of cancer a patient has. This helps doctors to make better choices for patient treatments.

Comparison of Human Genetic Sequence

The genetic sequences of many organisms have been determined. This allows for a comparison of the nucleotide sequences of each species to help determine evolutionary histories. The shaded regions represent mutations that are used to understand evolutionary relatedness.