- Email: [email protected]
Fusion Gene
738
Fungal Genetics
fitnesses relate to fertility differences among couples. Frank (1997) discusses the relation of the FTNS with Price's equation.
References
Bennett JH (1983) Natural Selection, Heredity, and Eugenics. Oxford: Clarendon Press. Ewens WJ (1989) An interpretation and proof of the Fundamental Theorem of Natural Selection. Theoretical Population Biology 36: 167±180. Fisher RA (1958) The Genetical Theory of Natural Selection. New York: Dover. Frank SA (1997) The Price equation, Fisher's Fundamental Theorem, kin selection, and casual analysis. Evolution 51(6): 1712±1729. Lessard S (1997) Fisher's fundamental theorem of natural selection revisited. Theoretical Population Biology 52: 119±136. Lessard S and Castilloux AM (1995) The fundamental theorem of natural selection in Ewens' sense: case of fertility selection. Genetics 141: 733±742. Nagylaki T (1991) Error bounds for the primary and secondary theorems of natural selection. Proceedings of the National Academy of Sciences, USA 88: 2402±2406. Nagylaki T (1992) Introduction to Theoretical Population Genetics. New York: Springer-Verlag. Price GR (1972) Fisher's `Fundamental Theorem' made clear. Annals of Human Genetics 36: 129±140.
See also: Additive Genetic Variance; Fisher, R.A.; Fitness; Wright, Sewall
Fungal Genetics D Stadler Copyright ß 2001 Academic Press doi: 10.1006/rwgn.2001.0484
Fungal genetics is the experimental study of the properties of genes and chromosomes carried out with filamentous fungi (such as Neurospora, Aspergillus, and Ascobolus) or with yeasts (such as Saccharomyces, Schizosaccharomyces, and Candida). These organisms have been important in basic genetics because they are eukaryotes but are also amenable to the elegant methods of bacteriology. See also: Ascobolus; Aspergillus nidulans; Neurospora crassa; Saccharomyces cerevisiae (Brewer's Yeast); Schizosaccharomyces pombe, the Principal Subject of Fission Yeast Genetics
Fungi D Stadler Copyright ß 2001 Academic Press doi: 006/rwgn.2001.0485
A group of simple, nongreen plants that includes molds, mushrooms, rusts and smuts, and sometimes yeasts. See also: Ascobolus; Aspergillus nidulans; Neurospora crassa
FUS-CHOP Fusion See: Myxoid Liposarcoma and FUS/TLS-CHOP Fusion Genes
Fusion Gene P Riggs Copyright ß 2001 Academic Press doi: 10.1006/rwgn.2001.0486
A gene fusion is defined as two genes that are joined so that they are transcribed and translated as a single unit. Gene fusions can occur in vivo, both naturally and as a result of genetic manipulations, and can be constructed in vitro using recombinant DNA techniques. They occur in nature over the course of evolution, for example, where two genes whose products are part of a metabolic pathway fuse, giving rise to a fusion protein that carries out both steps of the pathway.
History The first gene fusions created by design were between the rIIA and rIIB genes of phage T4, studied by Champe and Benzer. They used the effects of missense, nonsense and frameshift mutations in the rIIA gene on RIIB activity to elucidate the properties of the genetic code. Subsequently, fusions were created in Escherichia coli using in vivo genetic techniques to join various genes to the lacZ gene, which codes for the easily assayed enzyme b-galactosidase. These fusions were used as a way to examine the expression level and regulation of the gene fused to lacZ. Fusions were originally limited to genes that were located near the b-galactosidase gene, but later Casadaban and coworkers pioneered in vivo and in vitro techniques that allowed fusion to virtually any gene.
Fusion P roteins 739
Current Uses The major current use of gene fusions is still the study of gene expression, including levels of expression and location of gene products. Both gene fusions and reporter constructs (where the gene of interest is replaced by a `reporter' gene instead of being fused to it) are used for this purpose. Fusions to lacZ are common, but any gene whose product is active as a fusion and can be assayed is suitable for this purpose. In this method, an extract of a cell or tissue containing a gene fusion is prepared and the level of gene expression is measured by assaying the fusion. Gene fusions can also be used to study the differential expression of a gene in different tissues of an organism, by histochemical staining for the fused gene in sections, tissues, or the whole organism. Two genes commonly used for this technique are the lacZ and gfp genes. The lacZ gene has been used primarily because of the vast experience researchers have with b-galactosidase fusions, and the many substrates available for this enzyme. One of these substrates, X-gal, produces a dark-blue insoluble product when cleaved by bgalactosidase. Thus, the blue color does not diffuse away from the site of cleavage, and one can infer the location and level of expression from the intensity of the blue color. The gfp gene codes for green fluorescent protein, which fluoresces green when excited by blue or UV light. This allows visualization, and in many cases can be used on intact, live organisms.
Further Reading
Casadaban M J, Martinez-Avias A, Shapiro D K and Chou J (1983) b-galactosidase gene fusion for analyzing gene expression in Escherichia coli and yeast. Methods Enzymology 100: 293±307. Champe S P and Benzer S (1962) An active cistron fragment. Journal of Molecular Biology 4: 288±292.
See also: Beta (b)-Galactosidase; Fusion Proteins
Fusion Proteins P Riggs Copyright ß 2001 Academic Press doi: 10.1006/rwgn.2001.0487
A fusion protein is a protein consisting of at least two domains that are encoded by separate genes that have been joined so that they are transcribed and translated as a single unit, producing a single polypeptide. Fusion proteins can be created in vivo, but are usually created using recombinant DNA techniques. The fusion often consists of the protein that is being studied joined to
one of a small number of proteins that have useful properties to aid in the study.
History Some of the first fusion proteins were created in Escherichia coli using in vivo genetic techniques to join various proteins to the b-galactosidase enzyme. These fusions were used initially as a way to assay the expression level of the protein of interest. Fusions were originally limited to proteins whose genes were located near the b-galactosidase gene, but later, Casadaban and coworkers pioneered in vivo and in vitro techniques that allowed fusion to virtually any protein. Researchers were originally surprised that some of the fusions were bifunctional, i.e., when the C-terminus of a protein was fused to the amino terminus of b-galactosidase, both the proteins retained activity. As more and more fusions to b-galactosidase were obtained and found to have activity, researchers began to make fusions to other proteins besides b-galactosidase and found that they could be bifunctional as well.
Uses of Fusion Proteins The technique of creating fusion proteins has been extended to other fusion partners, and additional uses have been developed for the fusion partner. Three of the most important uses of fusion proteins are: as aids in the purification of cloned genes, as reporters of expression level, and as histochemical tags to enable visualization of the location of proteins in a cell, tissue, or organism. For purification, a protein that can be easily and conveniently purified by affinity chromatography is fused to a protein that the researcher wishes to study. A number of proteins and peptides have been used for this purpose, including staphylococcus protein A, glutathione-S-transferase, maltose-binding protein, cellulose-binding protein, chitin-binding domain, thioredoxin, strepavidin, RNaseI, polyhistidine, human growth hormone, ubiquitin, and antibody epitopes. The proteins used most often as fusion partners for reporter constructs are b-galactosidase, luciferase, and green fluorescent protein (GFP). b-galactosidase has the advantage of numerous commercially available substrates, including some that produce a colored product and some that lead to the production of light. Luciferase and GFP both produce light, and can be visualized directly or quantitated using a luminometer or a fluorometer, respectively. GFP has an advantage in that it does not require a substrate, whereas luciferase requires its substrate, luciferin, as
Fungal Genetics
fitnesses relate to fertility differences among couples. Frank (1997) discusses the relation of the FTNS with Price's equation.
References
Bennett JH (1983) Natural Selection, Heredity, and Eugenics. Oxford: Clarendon Press. Ewens WJ (1989) An interpretation and proof of the Fundamental Theorem of Natural Selection. Theoretical Population Biology 36: 167±180. Fisher RA (1958) The Genetical Theory of Natural Selection. New York: Dover. Frank SA (1997) The Price equation, Fisher's Fundamental Theorem, kin selection, and casual analysis. Evolution 51(6): 1712±1729. Lessard S (1997) Fisher's fundamental theorem of natural selection revisited. Theoretical Population Biology 52: 119±136. Lessard S and Castilloux AM (1995) The fundamental theorem of natural selection in Ewens' sense: case of fertility selection. Genetics 141: 733±742. Nagylaki T (1991) Error bounds for the primary and secondary theorems of natural selection. Proceedings of the National Academy of Sciences, USA 88: 2402±2406. Nagylaki T (1992) Introduction to Theoretical Population Genetics. New York: Springer-Verlag. Price GR (1972) Fisher's `Fundamental Theorem' made clear. Annals of Human Genetics 36: 129±140.
See also: Additive Genetic Variance; Fisher, R.A.; Fitness; Wright, Sewall
Fungal Genetics D Stadler Copyright ß 2001 Academic Press doi: 10.1006/rwgn.2001.0484
Fungal genetics is the experimental study of the properties of genes and chromosomes carried out with filamentous fungi (such as Neurospora, Aspergillus, and Ascobolus) or with yeasts (such as Saccharomyces, Schizosaccharomyces, and Candida). These organisms have been important in basic genetics because they are eukaryotes but are also amenable to the elegant methods of bacteriology. See also: Ascobolus; Aspergillus nidulans; Neurospora crassa; Saccharomyces cerevisiae (Brewer's Yeast); Schizosaccharomyces pombe, the Principal Subject of Fission Yeast Genetics
Fungi D Stadler Copyright ß 2001 Academic Press doi: 006/rwgn.2001.0485
A group of simple, nongreen plants that includes molds, mushrooms, rusts and smuts, and sometimes yeasts. See also: Ascobolus; Aspergillus nidulans; Neurospora crassa
FUS-CHOP Fusion See: Myxoid Liposarcoma and FUS/TLS-CHOP Fusion Genes
Fusion Gene P Riggs Copyright ß 2001 Academic Press doi: 10.1006/rwgn.2001.0486
A gene fusion is defined as two genes that are joined so that they are transcribed and translated as a single unit. Gene fusions can occur in vivo, both naturally and as a result of genetic manipulations, and can be constructed in vitro using recombinant DNA techniques. They occur in nature over the course of evolution, for example, where two genes whose products are part of a metabolic pathway fuse, giving rise to a fusion protein that carries out both steps of the pathway.
History The first gene fusions created by design were between the rIIA and rIIB genes of phage T4, studied by Champe and Benzer. They used the effects of missense, nonsense and frameshift mutations in the rIIA gene on RIIB activity to elucidate the properties of the genetic code. Subsequently, fusions were created in Escherichia coli using in vivo genetic techniques to join various genes to the lacZ gene, which codes for the easily assayed enzyme b-galactosidase. These fusions were used as a way to examine the expression level and regulation of the gene fused to lacZ. Fusions were originally limited to genes that were located near the b-galactosidase gene, but later Casadaban and coworkers pioneered in vivo and in vitro techniques that allowed fusion to virtually any gene.
Fusion P roteins 739
Current Uses The major current use of gene fusions is still the study of gene expression, including levels of expression and location of gene products. Both gene fusions and reporter constructs (where the gene of interest is replaced by a `reporter' gene instead of being fused to it) are used for this purpose. Fusions to lacZ are common, but any gene whose product is active as a fusion and can be assayed is suitable for this purpose. In this method, an extract of a cell or tissue containing a gene fusion is prepared and the level of gene expression is measured by assaying the fusion. Gene fusions can also be used to study the differential expression of a gene in different tissues of an organism, by histochemical staining for the fused gene in sections, tissues, or the whole organism. Two genes commonly used for this technique are the lacZ and gfp genes. The lacZ gene has been used primarily because of the vast experience researchers have with b-galactosidase fusions, and the many substrates available for this enzyme. One of these substrates, X-gal, produces a dark-blue insoluble product when cleaved by bgalactosidase. Thus, the blue color does not diffuse away from the site of cleavage, and one can infer the location and level of expression from the intensity of the blue color. The gfp gene codes for green fluorescent protein, which fluoresces green when excited by blue or UV light. This allows visualization, and in many cases can be used on intact, live organisms.
Further Reading
Casadaban M J, Martinez-Avias A, Shapiro D K and Chou J (1983) b-galactosidase gene fusion for analyzing gene expression in Escherichia coli and yeast. Methods Enzymology 100: 293±307. Champe S P and Benzer S (1962) An active cistron fragment. Journal of Molecular Biology 4: 288±292.
See also: Beta (b)-Galactosidase; Fusion Proteins
Fusion Proteins P Riggs Copyright ß 2001 Academic Press doi: 10.1006/rwgn.2001.0487
A fusion protein is a protein consisting of at least two domains that are encoded by separate genes that have been joined so that they are transcribed and translated as a single unit, producing a single polypeptide. Fusion proteins can be created in vivo, but are usually created using recombinant DNA techniques. The fusion often consists of the protein that is being studied joined to
one of a small number of proteins that have useful properties to aid in the study.
History Some of the first fusion proteins were created in Escherichia coli using in vivo genetic techniques to join various proteins to the b-galactosidase enzyme. These fusions were used initially as a way to assay the expression level of the protein of interest. Fusions were originally limited to proteins whose genes were located near the b-galactosidase gene, but later, Casadaban and coworkers pioneered in vivo and in vitro techniques that allowed fusion to virtually any protein. Researchers were originally surprised that some of the fusions were bifunctional, i.e., when the C-terminus of a protein was fused to the amino terminus of b-galactosidase, both the proteins retained activity. As more and more fusions to b-galactosidase were obtained and found to have activity, researchers began to make fusions to other proteins besides b-galactosidase and found that they could be bifunctional as well.
Uses of Fusion Proteins The technique of creating fusion proteins has been extended to other fusion partners, and additional uses have been developed for the fusion partner. Three of the most important uses of fusion proteins are: as aids in the purification of cloned genes, as reporters of expression level, and as histochemical tags to enable visualization of the location of proteins in a cell, tissue, or organism. For purification, a protein that can be easily and conveniently purified by affinity chromatography is fused to a protein that the researcher wishes to study. A number of proteins and peptides have been used for this purpose, including staphylococcus protein A, glutathione-S-transferase, maltose-binding protein, cellulose-binding protein, chitin-binding domain, thioredoxin, strepavidin, RNaseI, polyhistidine, human growth hormone, ubiquitin, and antibody epitopes. The proteins used most often as fusion partners for reporter constructs are b-galactosidase, luciferase, and green fluorescent protein (GFP). b-galactosidase has the advantage of numerous commercially available substrates, including some that produce a colored product and some that lead to the production of light. Luciferase and GFP both produce light, and can be visualized directly or quantitated using a luminometer or a fluorometer, respectively. GFP has an advantage in that it does not require a substrate, whereas luciferase requires its substrate, luciferin, as