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Covalent bond: a relatively strong chemical bond in which a single pair of electrons is shared by two different atoms.
DNA enzymology: The study of enzymes used to modify, ligate, cleave and/or synthesize DNA molecules.
DNA ligase: Enzyme involved in the replication and repair of DNA molecules. Ligase seals nicks in the backbone of one strand of a double stranded DNA molecule. It catalyses the formation of a (phosphodiester) bond between the 5' and 3' ends of the nicked DNA backbone.
DNA polymerase: Enzymes that catalyze the synthesis of new DNA molecules from single stranded DNA templates. There are many different DNA polymerases, each having a different function. For example: Escherichia coli bacterium have three different DNA polymerases called pol I, pol II, pol III. Pol III is mainly responsible for DNA replication. The other two function primarily in DNA repair.
Enzyme: A protein that acts as a catalyst which speeds up, enables, or controls chemical reactions in living organisms. Enzymes change the rate of the reactions without being used up in the reaction. Enzyme function is highly dependent on environmental characteristics such as temperature and pH.
Escherichia coli (E. coli): A Gram negative, rod-shaped bacterium which inhabits the lower intestines of most mammals. Because it grows at a wide range of temperatures with minimal nutrient requirements, it is the bacterium of choice for studies in genetics, biochemistry and molecular biology. In addition, E. coli has yielded a wide variety of restriction endonucleases including EcoRI, EcoRI', EcoRII, EcoB, EcoK, EcoPI, and EcoPI5. Although E. coli is ubiquitous among humans, some strains of this bacterium do cause human disease.
Haemophilus (or Hemophilus) influenzae bacteria: A Gram negative, rod-shaped bacterium which inhabits the upper respiratory tract of humans. H. influenzae has yielded a variety of restriction endonucleases including HindI, HindII, HindIII, and HindIV. Some strains of H. influenzae cause disease in humans.
Ligase: see DNA ligase
Methylase: An enzyme that attaches methyl (CH3-) groups to base residues in DNA and/or RNA molecules via covalent chemical bonds.
Methylation: The process of chemically attaching a methyl group to a base residue in DNA and/or RNA molecules. Methylation can protect DNA from cleavage by a restriction nuclease if the methylated base(s) is located within the nuclease's recognition sequence.
Nucleases: Enzymes that cleave or cut DNA and/or RNA molecules.
There are two types of nucleases: Endonucleases do not require a "free end" on the DNA strand and can thus cut along the length of the molecule. Exonucleases require a free end, and thus can only cut single base residues from the ends of the DNA strand.
Phage: (from the Greek word "phagein" meaning "to eat.") A virus that infects bacteria, also known as a bacteriophage.
Phage DNA: DNA that is packaged into a phage particle.
Phage particles: A complete phage virus with all its parts. Phages generally are composed of complex capsids containing the nucleic acid, and a tail through which the nucleic acid is injected into the infected host. Phage particles are unable to replicate without the aid of the host cell.
Prokaryotic cell: A cell whose genetic material is not enclosed in a nuclear envelope. For example: bacteria are prokaryotic cells.
Recognition sequence: The specific nucleotide sequence to which a restriction enzyme binds prior to cutting the DNA backbones. Recognition sequences are generally 4, 5, or 6 base pairs in length, and are often palindromic--that is, they read the same backwards and forwards when they are read in the 5' - 3' direction
EcoRI 5' GAATTC 3'
3' CTTAAG 5'
Restriction nuclease: (See restriction enzyme.) Enzymes that cleave DNA molecules after recognition of specific sequences of bases.
Restriction enzyme: An endonuclease which recognizes a specific sequence of bases in a DNA molecule. Type 1 restriction enzymes bind to the recognition site and then cut randomly somewhere along the length of the molecule. Type II restriction enzymes bind at a recognition site and then cleave the molecule by clipping the DNA backbones somewhere within this sequence of bases. Each restriction enzyme has a single, specific recognition sequence, and clips the DNA molecule at a specific site. Thus, treatment of a particular DNA molecule with a particular restriction enzyme will always produce the same set of DNA fragments. It is type II restriction enzymes that have been used extensively in recombinant DNA technology.
Strain of bacteria: A group of bacterial cells all derived from a single cell. Strains are identified by information which follows the name of the genus and species.