Restriction endonuclease: any of a large family of enzymes that recognize, bind to, and hydrolyze specific nucleic acid sequences in double-stranded DNA
Cloning: in molecular biology this term refers to the production of large quantities of identical DNA molecules
Blotting: a molecular biological technique that involves the transfer of proteins, DNA, or RNA, out of the size-separating gel onto a solid support such as filter paper
DNA sequencing: the process of determining the precise order of nucleotides within a DNA molecule
Microarray: commonly known as DNA chip, is a collection of microscopic DNA spots attached to a solid surface
Gene chip: is the solid medium of a microarray containing the DNA spots
Transgenesis: the process of inserting a gene from one source into a living organism that would not normally contain it
Gene therapy: a technique involving the insertion of genes into an individual's cells and tissues to treat a disease, such as a hereditary disease in which a deleterious mutant allele is replaced with a functional one
Modern molecular medicine encompasses the utilization of many molecular biological techniques in the analysis of disease, disease genes, and disease gene function. The study of disease genes and their function in an unaffected individual has been possible by the development of recombinant DNA and cloning techniques. The basis of the term recombinant DNA refers to the recombining of different segments of DNA. Cloning refers to the process of preparing multiple copies of an individual type of recombinant DNA molecule. The classical mechanisms for producing recombinant molecules involves the insertion of exogenous fragments of DNA into either bacterially derived plasmid (circular double-stranded autonomously replicating DNAs found in bacteria) vectors or bacteriophage (viruses that infect bacteria)-based vectors. The term vector refers to the DNA molecule used to carry or transport DNA of interest into cells.
The technologies of molecular biology are critical to modern medical diagnosis and treatment. For example, cloning allows large quantities of therapeutically beneficial human proteins to be produced in pure form. In addition the cloning of genes allows for the utilization of the DNA for treatments referred to as gene therapy which involves the introduction of a normal copy of a gene into an individual harboring a defect gene. These molecular biology techniques involve the use of a wide array of different enzymes, many of which are purified from bacteria (Table 41-1).
TABLE 41-1:Common Enzymes Used in Molecular Biology ||Download (.pdf) TABLE 41-1: Common Enzymes Used in Molecular Biology
|Enzyme(s) ||Activity ||Comments |
|Restriction endonucleases ||Recognize specific nucleotide sequences and cleaves the DNA within or near to the recognition sequences ||See Table 41-2 |
|Reverse transcriptase (RT) ||Retrovirally encoded RNA-dependent DNA polymerase ||Used to convert mRNA into a complimentary DNA (cDNA) copy for the purpose of cloning cDNAs |
|RNase H ||Recognizes RNA-DNA duplexes and randomly cleaves the phosphodiester backbone of the RNA...|