DNA

DNA

DNA contains genes.

DNA, or deoxyribonucleic acid, differs from RNA in that it is missing a hydroxyl group on its 2 Carbon. This makes the DNA more stable than RNA.

The two strands of DNA, of opposing polarity, are held together by hydrogen bonding between base pairs.

DNA contains 4 bases: adenine, guanine, cytosine, and thyamine. A and T bond together, as do C and G.

Hydrogen bonds can be broken through a process called denaturation, which involves the addition of heat or chemicals. As the the C-G pair contains 3 H bonds, and A-T contains only 2, the proportion of each base will determine the melting temperature (T_M) of a sequence. Denatured DNA will anneal with specific, complementary fragments of DNA such as probes or primers. This allows for various techniques such as hybridization or PCR.

Nucleosomes

Negatively charged DNA is coiled around positively charged proteins called histones, forming a structure analagous to beads on a string. Nucleosomes form nucelofilaments, or 30 nm fibres, that are anchored to scaffold proteins to form chromosomes.

DNA Replication

DNA replication is semi-conservative, meaning the parental strands are both replicated and split between the daughter strands. RNA primers are recognized by DNA Polymerase III, which elongates the DNA. DNA Pol III also has proofreading ability. DNA Pol I fills in the gaps.

Restriction Enzymes

DNA is cleaved at certain sequences by restriction endonucleases - enzymes that act with high specifity at restriction sites. These sites are usually 4-8 bases and produce similar cut ends, which can be blunt or sticky. Restriction enzymes are bacterial proteins used to degrade foreign DNA and are used in basic and clinical research to identify specific sequences of DNA.

Southern Blotting

Southern Blotting is used to detect specific DNA sequences by using gel electrophoresis and probe hybridization.

DNA Sequencing

DNA was first sequenced using the Sanger Method, which uses labeled dideoxynucleotides that prevent further elongation. Using the 4 ddNTPs produces DNA fragments of varying lengths that can be electrophoresed and examined to show the DNA sequence.

Automated sequencing now uses fluorescently labeled ddNTPs and separates the fragments using capillary electrophoresis. It is capable of analyzing over 1000 bases/sec.