Genetic Engineering Challenge How can scientists develop a type of rice that could prevent vitamin A deficiency? 1 Vitamin A deficiency can result in blindness, severe infectious diseases, and even death, especially for children in poor countries. Vitamin A deficiency results when people do not have enough vitamin A or pro-vitamin A in their diets. Pro-vitamin A is a molecule that our bodies can easily convert to vitamin A. Some plant foods (e.g. carrots and sweet potatoes) are good sources of pro-vitamin A. Some animal foods (e.g. liver and eggs) are good sources of vitamin A. White rice is part of the rice seed. Brown rice includes white rice, the germ (which is the embryo that can grow into a new rice plant), and the surrounding bran layers. Rice is a good source of starch for energy and also provides some protein. However, neither white rice nor brown rice has pro-vitamin A. Therefore, many poor people in developing countries who eat mainly rice do not get enough pro-vitamin A or vitamin A in their diet. This results in many cases of blindness, severe infectious disease and/or death. 1. Why do rice plants include starch and protein in their seeds? What is the benefit for the rice plants? The germ is the embryo inside the seed. The bulk of the rice seed is the white rice which contains starch and storage proteins. For the rest of this activity, we will use the term rice grains to refer to white rice. No type of rice plant has been discovered that has pro-vitamin A in the rice grains; therefore, conventional breeding techniques cannot be used to develop a type of rice plant that has pro-vitamin A in the rice grains. So, some scientists who want to prevent vitamin A deficiency have used genetic engineering to develop rice plants that have rice grains with substantial pro-vitamin A. Genetic engineering is the process of manipulating genes in order to produce desired characteristics. In this activity we will discuss the type of genetic engineering that produces recombinant DNA (DNA which contains genes from two different organisms). We will learn how genetic engineering has been used to produce rice plants that have two additional genes inserted in their DNA, so these rice plants make substantial quantities of pro-vitamin A in their rice grains. 1 By Dr. Ingrid Waldron, Department of Biology, University of Pennsylvania, 2014. Teachers are encouraged to copy this Student Handout for classroom use. A Word file (which can be used to prepare a modified version if desired), Teacher Preparation Notes, comments, and related activities are available at http://serendip.brynmawr.edu/exchange/bioactivities/geneticengineer 1
2. Vitamin A and pro-vitamin A are molecules that contain only carbon and hydrogen atoms and do not have any amino acids. If scientists want to genetically engineer a plant to make provitamin A, what type of gene or genes would the scientists need to insert in the plant cells? Would they insert a gene that codes for pro-vitamin A? If not, what type of molecule would the gene or genes have to code for? 3. Scientists have identified genes for two enzymes needed to make pro-vitamin A. One of these genes comes from corn. If this gene from a corn plant is inserted in the DNA of a rice plant, will the sequence of amino acids in the protein produced by the rice plant be the same as the sequence of amino acids in the protein produced by the corn plant? In other words, will rice plants that have this gene produce the same enzyme as corn plants produce? Explain why or why not. You will learn about how scientists have genetically engineered rice plants to make pro-vitamin A in rice grains, but first answer question 4 to develop your own ideas about how this could be accomplished. 4a. Once scientists have identified the genes for enzymes to produce provitamin A, how could they insert these genes in the DNA of rice plant cells? Suggest one possibility. Be inventive! 4b. Would you recommend that scientists try to insert the genes for enzymes to produce provitamin A into: all the cells in a rice plant the thousands of cells in each rice grain or a small group of embryonic rice plant cells that can divide and develop into a rice plant? 2
Inserting the Desired Genes in the DNA of Rice Plants To insert genes from one organism into a different organism, scientists often take advantage of the natural genetic engineering capabilities of bacteria or viruses. One type of bacteria genetically engineers plant cells by inserting part of its bacterial DNA into the plant cell DNA, thus producing recombinant DNA. The inserted bacterial genes code for proteins that: stimulate the genetically engineered plant cells to produce food molecules that only the bacteria can use stimulate these genetically engineered plant cells to divide and form a growth that bulges out from the stem or root. 5. Explain how this type of genetic engineering is useful for the bacterium. This figure shows how the bacterium inserts some of its genes into the plant cell DNA. The plasmid is a small circle of DNA in the bacterium, separate from the chromosome. Only the T-DNA from the plasmid is inserted in the DNA in the nucleus of the plant cell. The genes in the T-DNA code for the proteins that result in the production of the food molecules for the bacteria and the proteins that result in increased cell division. The bacterial chromosome is shown at the bottom of the bacterium (A. tumifaciens). As in many diagrams, the relative dimensions are distorted; e.g., the bacterium appears much larger than it actually is relative to the plant cell and the plasmid appears much larger than it is relative to the bacterium. The plasmid is called the Ti or tumor inducing plasmid because the infected plant cells multiply and produce a bulging growth. The vir genes in the plasmid code for the proteins that carry out the transfer of the T-DNA from the bacteria into the plant cell DNA. (Figure from http://www.open.edu/openlearn/science-maths-technology/science/biology/gene-manipulation-plants/content-section-2.2) 6. In this example of genetic engineering in nature, where is the recombinant DNA found? This recombinant DNA contains DNA from the and the. 3
7. If a scientist wants to use the genetic engineering capabilities of these bacteria to carry the genes for the enzymes to make pro-vitamin A into a plant cell nucleus, where should she insert the genes for these enzymes? Use an arrow to indicate specifically where these genes should be inserted in the bacterium. The basic sequence of steps to use genetic engineering to produce rice plants that make provitamin A in rice grains is as follows: Scientists insert the genes for the enzymes to make pro-vitamin A in the bacterial plasmid. Bacteria with this modified plasmid are grown together with tissue culture rice plant cells, so the bacteria insert the genes for the enzymes to make pro-vitamin A in the DNA of the tissue culture rice plant cells. After the rice plant tissue culture cells have the genes for the enzymes to make pro-vitamin in their DNA, these genes are replicated together with the rest of the DNA before each cell division. These cells multiply and develop into a rice plant with the genes to make provitamin A in the DNA of each cell in the rice plant, including the thousands of cells in each rice grain. Ensuring that the Genes for the Enzymes to Make Pro-Vitamin A are Active in Rice Grain Cells 8. Almost all the cells in an animal or plant have exactly the same genes, but different types of cells make different types of proteins. This allows each type of cell to carry out its particular specialized function. Example 1: In humans, red blood cells have lots of, while muscle cells (contractile proteins or hemoglobin = O 2-carrying protein) have lots of. (contractile proteins or hemoglobin) Example 2: Almost all the cells in a rice plant have exactly the same, (genes or proteins) but cells in the rice grains make storage proteins which provide nutrition for the developing embryo, whereas cells in the leaves make enzymes that make chlorophyll. 9. What is transcription? Why do cells need to carry out transcription of genes in order to make proteins? 4
10. Match each type of gene in the top list with the best match from the bottom list. Genes for enzymes to make chlorophyll Genes for storage proteins a. rate of transcription is higher in cells in rice grains b. rate of transcription is the same in cells in rice grains and cells in rice plant leaves c. rate of transcription is higher in cells in rice plant leaves For genetic engineering to be successful, scientists need to ensure that the genes they have inserted are transcribed, so the desired proteins are produced. Specifically, the scientists need to ensure that the genes for the enzymes to make pro-vitamin A are transcribed in the rice grain cells. Plant cells have a rather complex molecular mechanism to ensure the appropriate rate of transcription for each gene in each type of cell. A crucial part of this molecular mechanism is the promoter segment of DNA (located at the beginning of the gene). For example, the promoters for the genes for storage proteins promote high rates of transcription in the cells in rice grains. 11. Which promoter do you think would be a better promoter to insert at the beginning of the genes that code for the enzymes to produce pro-vitamin A? the promoter for a gene for an enzyme to make chlorophyll the promoter for a gene for a storage protein 12. The promoter is inserted in the bacterial plasmid at the beginning of the genes for the enzymes to produce provitamin A. Explain how the promoter and genes get from the plasmid in a bacterium into the DNA of a rice plant cell and then into the DNA of every cell in a rice plant. Plasmid Promoter + genes for enzymes 5