Genetic Engineering and DNA Technology
Plasmid
a small, circular, double-stranded DNA molecule that carries accessory genes separate from those of a bacterial chromosome; in DNA cloning, used as vectors carrying up to about 10,000 base pairs (10 kb) of DNA. Plasmids are also found in some eukaryotes, such as yeasts.
cDNA Library
A gene library containing clones that carry complementary DNA (cDNA) inserts. The library includes only the genes that were transcribed in the cells whose mRNA was isolated to make the cDNA.
DNA Microarray Assay
A method to detect and measure the expression of thousands of genes at one time. Tiny amounts of a large number of single-stranded DNA fragments representing different genes are fixed to a glass slide and tested for hybridization with samples of labeled cDNA.
Totipotent
describing a cell that can give rise to all parts of the embryo and adult, as well as extraembryonic membranes in species that have them.
Pluripotent
describing a cell that can give rise to many, but not all, parts of an organism.
Cloning Vector
In genetic engineering, a DNA molecule that can carry foreign DNA into a host cell and replicate there. Cloning vectors include plasmids and bacterial artificial chromosomes (BACs), which move recombinant DNA from a test tube back into a cell, and viruses that transfer recombinant DNA by infection.
In Situ Hybridization
a technique using nucleic acid hybridization with a labeled probe to detect the location of a specific mRNA in an intact organism.
In Vitro Mutagenesis
a technique used to discover the function of a gene by cloning it, introducing specific changes into the cloned gene’s sequence, reinserting the mutated gene into a cell, and studying the phenotype of the mutant.
Gel Electrophoresis
a technique for separating nucleic acids or proteins on the basis of size and electrical charge, both of which affect their rate of movement through an electric field in a gel made of agarose or another polymer.
Single Nucleotide Polymorphism
a single base-pair site in a genome where nucleotide variation is found in at least 1% of the population.
Gene Cloning
the production of multiple copies of a gene.
Genetic Engineering
The direct manipulation of genes for practical purposes.
Restriction Fragment
a DNA segment that results from the cutting of DNA by a restriction enzyme.
Genomic Library
A set of cell clones containing all the DNA segments from a genome, each within a plasmid BAC, or other cloning vector.
Gene Therapy
The introduction of genes into an afflicted individual for therapeutic purposes.
Restriction Enzyme
An endonuclease (type of enzyme) that recognizes and cuts DNA molecules foreign to a bacterium (such as phage genomes). The enzyme cuts at specific nucleotide sequences (restriction sites).
Expression Vector
a cloning vector that contains a highly active bacterial promoter just upstream of a restriction site where a eukaryotic gene can be inserted, allowing the gene to be expressed in a bacterial cell. Expression vectors are also available that have been genetically engineered for use in specific types of eukaryotic cells.
Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR)
a technique for determining expression of a particular gene. It uses reverse transcriptase and DNA polymerase to synthesize cDNA from all the mRNA in a sample and then subjects the cDNA to PCR amplification using primers specific for the gene of interest.
Transgenic
pertaining to an organism whose genome contains a gene introduced from another organism of the same or a different species.
Deoxyribonucleic Acid
a nucleic acid molecule, usually a double-stranded helix, in which each polynucleotide strand consists of nucleotide monomers with a deoxyribose sugar and the nitrogenous bases adenine (A), cytosine (C), guanine (G), and thymine (T); capable of being replicated and determining the inherited structure of a cell’s proteins.
Polymerase Chain Reaction (PCR)
a technique for amplifying DNA in vitro by incubating it with specific primers, a heat-resistant DNA polymerase, and nucleotides.
Bacterial Artificial Chromosomes (BAC)
a large plasmid that acts as a bacterial chromosome and can carry inserts of 100,000 to 300,000 base pairs (100 – 300 kb).
Restriction Fragment Length Polymorphism (RFLP)
a single nucleotide polymorphism (SNP) that exists in the restriction site for a particular enzyme, thus making the site unrecognizable by that enzyme and changing the lengths of the restriction fragments formed by digestion with that enzyme. A RFLP can be coding or non-coding DNA.
Stem Cell
any relatively unspecified cell that can produce, during a single division, one identical daughter cell and one more specialized daughter cell that can undergo further differentiation.
Restriction Site
a specific sequence on a DNA strand that is recognized and cut by a restriction enzyme.
Northern Blotting
a technique that enables specific nucleotide sequences to be detected in samples of mRNA. It involves gel electrophoresis of RNA molecules and their transfer to a membrane (blotting), followed by nucleic acid hybridization with a labeled probe.
RNA Interference (RNAi)
a technique used to silence the expression of selected genes. RNAi uses synthetic double-stranded RNA molecules that match the sequence of a particular gene to trigger the breakdown of the gene’s messenger RNA.
Nucleic Acid Hybridization
the process of base pairing between a gene and a complementary sequence on another nucleic acid molecule.
Short Tandem Repeats (STR)
simple sequence DNA containing multiple tandemly repeated units of 2-5 nucleotides. Variations in STRs act as genetic markers in STR analysis, used to prepare genetic profiles.
Recombinant DNA
A DNA molecule made in vitro with segments from different sources.
Complementary DNA (cDNA)
a double-stranded DNA molecule made in vitro using mRNA as a template and the enzymes reverse transcriptase and DNA polymerase. A cDNA molecule corresponds to the exons of a gene.
Electroporation
a technique to introduce recombinant into cells by applying a brief electrical pulse to a solution containing the cells. The pulse creates temporary holes in the cells’ plasma membranes, through which DNA can enter.
Nucleic Acid Probe
In DNA technology, a labeled single-stranded nucleic acid molecule used to locate a specific nucleotide sequence wherever it occurs; radioactive, fluorescent, or other labeling of the probe allows its location to be detected.
Genetic Profile
an individual’s unique set of genetic markers, detected most often today by PCR or, previously, by electrophoresis and nucleic acid probes.
Sticky End
a single-stranded end of a double-stranded restriction fragment.
Ti Plasmid
a plasmid of a tumor-inducing bacterium (the plant pathogen Agrobacterium) that integrates a segment of its DNA (T DNA) into a chromosome of a host plant. The Ti plasmid is frequently used as a vector for genetic engineering in plants.
Hybridization
In genetics, the mating or crossing of two true-breeding varieties.
Southern Blotting
a technique that enables specific nucleotide sequences to be detected in samples of DNA. It involves gel electrophoresis of DNA molecules and their transfer to a membrane (blotting), followed by nucleic acid hybridization with a labeled probe.