Genetic Engineering Between a Tree and a Firefly

It sounds like a child’s riddle: What do you get when you cross a firefly with a tobacco plant? Answer: tobacco that lights itself. That is essentially what a team of scientists at the University of California at San Diego has done. By outfitting a fragment of a plant virus with the gene that tells firefly cells to produce a protein central to generating light, the researchers have created a plant that literally glows in the dark. The technique, reported in last week’s issue of the journal Science, is significant not so much as a demonstration of virtuoso genetic engineering, but…

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The luciferase gene from the firefly, Photinus pyralis, was used as a reporter of gene expression by light production in transfected plant cells and transgenic plants. A complementary DNA clone of the firefly luciferase gene under the control of a plant virus promoter (cauliflower mosaic virus 35S RNA promoter) was introduced into plant protoplast cells (Daucus carota) by electroporation and into plants (Nicotiana tabacum) by use of the Arobacterum tumefaciens tumor-inducing plasmid.

Extracts from electroporated celLs (24 hours after the introduction of DNA) and from transgenic plants produce light when mixed with the substrates luciferin and adenosine triphosphate. Light produced by the action of luciferase was also detected in undisrupted leaves or cells in culture from transgenic plants incubated in luciferin and in whole transgenic plants “watered” with luciferin.

Although light was detected in most organs in intact, transgenic plants (leaves, stems, and roots), the pattern of luminescence appeared to reflect both the organ-specific distribution of luciferase and the pathway for uptake of luciferin through the vasculaure of the plant.. Luciferase Luciferase is a generic name for enzymes commonly used in nature for bioluminescence. The most famous one is firefly luciferase (EC 1. 13. 12. 7). In luminescent reactions, light is produced by the oxidation of a luciferin (a pigment), sometimes involving Adenosine triphosphate (ATP) .

The rates of this reaction between luciferin and oxygen are extremely slow until they are triggered by the presence of luciferase. The reaction takes place in two steps: luciferin + ATP –> luciferyl adenylate + PPi luciferyl adenylate + O2 –> oxyluciferin + AMP + light The reaction is very energy efficient: nearly all of the energy input into the reaction is transformed into light. As a comparison, the incandescent light bulb loses about 90% of its energy to heat. Luciferin and luciferase do not refer to a particular molecule.

They are generic terms for a light-producing chemical and its associated regulatory compound, usually a protein. A wide variety of species regulate their light production using a luciferase. The most famous is the firefly, although it even exists in organisms as different as the Jack-O-Lantern mushroom and many marine creatures. In the firefly, the oxygen required is supplied through a tube in the abdomen called the abdominal trachea. Some organisms, notably the click beetles, have several different luciferase enzymes which each can produce different colors from the same luciferin.

Luciferase can be produced in the lab through genetic techniques, and has a wide variety of uses. Genes for luciferase can be genetically engineered into organisms so that they glow when exposed to the right luciferin. This allows visualization of certain biological processes, stages of infection, and provides other valuable sources of information. Mice, silkworms, and potatoes are just a few organisms that have already been engineered to produce the chemical. Luciferase can be used in blood banks to determine if red blood cells are starting to break down.

Laboratories can use luciferase to produce light in the presence of certain diseases. The possibilities for uses for luciferase continue to expand. One peculiar application of luciferase was when a team of students from the University of Hertfordshire made news with their proposal to genetically engineer christmas trees to glow without lights, using luciferase (in addition to other bioluminescent chemicals). Luciferin would be added to a fertilizer used for the tree, which would react with the luciferase that the tree produces on its own.

Genetically modified organisms utilizing bioluminescence include transgenic mice, fish, and microbes such as bacteria and fungi. Genetic modification is surrounded by controversy, as our ability to create new genes, and new organisms, has outstripped our knowledge of their long-term effect on broader ecosystems. It is also a fascinating and exciting field, as fluorescent genes open up new perspectives on the inner workings of cells themselves. Fluorescent genes, such as the gene GFP found in jellyfish, or luciferase from fireflies, can be directed to target a specific mammalian gene, revealing the exact ocations of the particular protein associated with that gene within a cell. This is an invaluable tool in medical research, pharmaceutical development and drug screening, gene therapy, and improving our understanding of biological processes. There are many possible applications for engineering bioluminescence in a variety of forms. Glowing fish could make for exotic pets, while trees that include bioluminescent genes could be used to illuminate roads and highways, or to provide Christmas trees that are truly green and don’t require electricity.

Bioluminescence could also be used as a warning or flag, to show when action is required. Crops could glow to indicate they need water, bad bacteria in meat could be located, glowing tags could identify escaped criminal offenders, and contaminated water could be flagged and monitored. Genetic engineering is any process by which genetic material (the building blocks of heredity) is changed in such a way as to make possible the production of new substances or new functions. As an example, biologists have now learned how to transplant the gene that produces light in a firefly into tobacco plants.

The function of that gene—the production of light—has been added to the normal list of functions of the tobacco plants. Words to Know Amino acid: An organic compound from which proteins are made. DNA (deoxyribonucleic acid): A large, complex chemical compound that makes up the core of a chromosome and whose segments consist of genes. Gene: A segment of a DNA molecule that acts as a kind of code for the production of some specific protein. Genes carry instructions for the formation, functioning, and transmission of specific traits from one generation to another.

Genetic code: A set of nitrogen base combinations that act as a code for the production of certain amino acids. Host cell: The cell into which a new gene is transplanted in genetic engineering. Human gene therapy (HGT): The application of genetic engineering technology for the cure of genetic disorders. Nitrogen base: An organic compound consisting of carbon, hydrogen, oxygen, and nitrogen arranged in a ring that plays an essential role in the structure of DNA molecules. Plasmid: A circular form of DNA often used as a vector in genetic engineering.

Protein: Large molecules that are essential to the structure and functioning of all living cells. Recombinant DNA research (rDNA research): Genetic engineering; a technique for adding new instructions to the DNA of a host cell by combining genes from two different sources. Vector: An organism or chemical used to transport a gene into a new host cell. Gene splicing Gene splicing: The process by which genes are cut apart and put back together to provide them with some new function. Scientists have discovered that cells contain certain kinds of enzymes that take DNA molecules apart and put them back together again.

Endonucleases, for example, are enzymes that cut a DNA molecule at some given location. Exonucleases are enzymes that remove one nitrogen base unit at a time. Ligases are enzymes that join two DNA segments together. It should be obvious that enzymes such as these can be used by scientists as submicroscopic scissors and glue with which one or more DNA molecules can be cut apart, rearranged, and the put back together again. Genetic engineering procedures Genetic engineering requires three elements: the gene to be transferred, a host cell into which the gene is inserted, and a vector to bring about the transfer.

Suppose, for example, that one wishes to insert the gene for making insulin into a bacterial cell. Insulin is a naturally occurring protein made by cells in the pancreas in humans and other mammals. It controls the breakdown of complex carbohydrates in the blood to glucose. People whose bodies have lost the ability to make insulin become diabetic. The first step in the genetic engineering procedure is to obtain a copy of the insulin gene. This copy can be obtained from a natural source (from the DNA in a pancreas, for example), or it can be manufactured in a laboratory.

The second step in the process is to insert the insulin gene into the vector. The term vector means any organism that will carry the gene from one place to another. The most common vector used in genetic engineering is a circular form of DNA known as a plasmid. Endonucleases are used to cut the plasmid molecule open at almost any point chosen by the scientist. Once the plasmid has been cut open, it is mixed with the insulin gene and a ligase enzyme. The goal is to make sure that the insulin gene attaches itself to the plasmid before the plasmid is reclosed.

The hybrid plasmid now contains the gene whose product (insulin) is desired. It can be inserted into the host cell, where it begins to function just like all the other genes that make up the cell. In this case, however, in addition to normal bacterial functions, the host cell also is producing insulin, as directed by the inserted gene. Notice that the process described here involves nothing more in concept than taking DNA molecules apart and recombining them in a different arrangement. For that reason, the process also is referred to as recombinant DNA (rDNA) research.

The early 1990s saw the creation of formalized working relations between universities, individual researchers, and the corporations founded by these individuals. Despite these arrangements, however, many ethical issues remain unresolved. Sci/Tech GM Christmas tree would glow No more ceremonial switch-ons? Frustrated fiddling with Christmas tree fairy lights could become a thing of the past as genetic engineers have proposed a tree which grows its own lights. The idea for glowing pine needles was dreamed up by five postgraduate students at the University of Hertfordshire, UK, as their entry in a biotechnology competition.

It is a perfectly possible proposition, as genetic engineers elsewhere have already created glowing mice, silk and potatoes. ‘Only problem cost’ Neurophysiology student Katy Presland, 29, said: “We’re talking about a green luminescent Christmas tree that glows in the dark and produces a noticeable light during the day. “It is quite feasible. The only problem in reality is the cost,” she added. “We calculate that the initial trees would cost about ? 200, which means going for the upper end of the market. But I’m sure a lot of people would love them, especially the Americans. Jellyfish and fireflies The team detail a plan to modify a Douglas spruce with two genes to make it illuminate. These would taken from fluorescent jellyfish and fireflies. The first gene produces a substance called green fluorescent protein (GFP), while the second results in an enzyme called luciferase. The trees would be modified by infecting seedlings with a harmless bacterium carrying the genes. A chemical compound called luciferin is needed to activate luciferase, which in turn “switches on” GFP and makes it glow.

In the case of the luminous Douglas spruce, the luciferin would be mixed into a special fertilizer sold with the tree. The genes for green fluorescence have been widely used by genetic engineers because they allows scientists to see at a glance whether an attempt to introduce a gene into an organism has been successful. Blue fluorescent proteins have also been discovered and, last month, a red fluorescent protein was found in a coral. This means that, in theory, the GM Christmas tree could grow its own multicoloured lights. een a

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