8 Biotechnology Quarter 3 – Module 3: Steps in Recombinant DNA Technology DIVISION OF ANGELES CITY Biotechnology– Grade 8 Alternative Delivery Mode Quarter 3 – Module 3: Steps in Recombinant DNA Technology First Edition, 2021 Republic Act 8293, section 176 states that: No copyright shall subsist in any work of the Government of the Philippines. However, prior approval of the government agency or office wherein the work is created shall be necessary for exploitation of such work for profit. Such agency or office may, among other things, impose as a condition the payment of royalties. Borrowed materials (i.e., songs, stories, poems, pictures, photos, brand names, trademarks, etc.) included in this module are owned by their respective copyright holders. Every effort has been exerted to locate and seek permission to use these materials from their respective copyright owners. The publisher and authors do not represent nor claim ownership over them. Published by the Department of Education Regional Director: May B Eclar PhD, CESE III Assistant Regional Director: Rhoda T. Razon EdD, CESO V Development Team of the Module Writers: Larissa Manalili Editors: Sherilyne L. Reyes, Jennifer Praza, Edgardo D. Cortez, Jenny S. Tongol, Edythe Hipolito Reviewers: Gemima A. Estrabillo, Emily F. Sarmiento, Hermes Vargas, Adrian Tamayo, Krislene Ida N. Mercado, Noel S. Reganit Mercedes Bactol, Billy Ray B. Manuel, Marvin R. Leano, Gemmarie G. Rivas Illustrator: Arnold Arceo Layout Artist: Maricon H. Rivera, Noel S. Reganit Management Team: May B. Eclar PhD, CESO V Rhoda T. Razon EdD, CESO V Ma. Irelyn P. Tamayo PhD, CESE Fernandina P. Otchengco, PhD, CESE Librada M. Rubio PhD Ma. Editha R. Caparas EdD Rochella C. David Emily F. Sarmiento PhD Gemima A. Estrabillo EdD Printed in the Philippines by ________________________ Department of Education –Region III – Schools Division of Angeles City Office Address: Jesus St. Pulungbulu, Angeles City Telefax: (045) 322-5722;322-472 888-0582;887-6099 E-mail Address: [email protected] 8 Biotechnology Quarter 3 – Module 3: Steps in Recombinant DNA Technology Introductory Message This Self-Learning Module (SLM) is prepared so that you, our dear learners, can continue your studies and learn while at home. Activities, questions, directions, exercises, and discussions are carefully stated for you to understand each lesson. Each SLM is composed of different parts. Each part shall guide you step-bystep as you discover and understand the lesson prepared for you. Pre-tests are provided to measure your prior knowledge on lessons in each SLM. This will tell you if you need to proceed on completing this module or if you need to ask your facilitator or your teacher’s assistance for better understanding of the lesson. At the end of each module, you need to answer the post-test to self-check your learning. Answer keys are provided for each activity and test. We trust that you will be honest in using these. In addition to the material in the main text, Notes to the Teacher are also provided to our facilitators and parents for strategies and reminders on how they can best help you on your home-based learning. Please use this module with care. Do not put unnecessary marks on any part of this SLM. Use a separate sheet of paper in answering the exercises and tests. And read the instructions carefully before performing each task. If you have any questions in using this SLM or any difficulty in answering the tasks in this module, do not hesitate to consult your teacher or facilitator. Thank you. 2 What I Need to Know This module was designed and written with you in mind. It is here to help you master the steps in Recombinant DNA Technology. The scope of this module permits it to be used in many different learning situations. The language used recognizes the diverse vocabulary level of students. The lessons are arranged to follow the standard sequence of the course. But the order in which you read them can be changed to correspond with the textbook you are now using. The module is about: ● Steps in Recombinant DNA Technology After going through this module, you are expected to: 1. Outline the steps in Recombinant DNA Technology 3 What I Know Directions: Read each question carefully. Choose the letter of the best answer. 1. Which refers to the combination of two DNA strands that are constructed artificially? a. Genetic Material b. Recombinant Cells c. Recombinant DNA d. Restriction Enzymes 2. Which among the following is the Blueprint of Life? a. Cell b. DNA c. Protein d. RNA 3. Which refers to the small accessory ring of the DNA in some bacteria? a. Interferon b. Plasmid c. Restriction enzymes d. Vector 4. What molecule is being used to cut a specific area of the DNA? a. Agarose Gel b. Plasmid c. Recombinant Cells d. Restriction enzymes 5. An organism that contains genetic material from two different organisms is called ___________. a. Clone b. GMO c. Mutant d. Restriction enzymes 6. What technique is being used to make a million copies of a sample DNA? a. DNA Fingerprinting b. Gel Electrophoresis c. Gene Therapy d. Polymerase Chain Reaction 4 7. Which among the following does NOT describe the principle of Gel Electrophoresis? a. DNA fragments are separated on the basis of size. b. The DNA fragments will move towards the negative charge. c. The smallest DNA fragments will move faster than the larger DNA fragments. d. DNA fragments are injected into wells and an electric current is applied along with the gel. 8. What equipment is used to amplify the DNA? a. Centrifuge Machine b. Microscope c. Pipette d. Thermal Cycler 9. Which among the following serves as a starting point for DNA synthesis? a. DNA polymerase b. Primer c. Restriction enzymes d. Taq polymerase 10. What organism is being used to transfer foreign genetic material into a cell? a. DNA b. Plasmid c. Restriction enzymes d. Vector 5 Lesson 1 Steps in Recombinant DNA Technology What’s In Activity 1 Direction: Write True if the statement is correct and False if incorrect. 1. Genetic Engineering is the artificial manipulation of DNA or other nucleic acid molecules in order to modify an organism or population of organisms. 2. Restriction enzyme was discovered by Werner Arber. 3. Genes are small rings of DNA. 4. DNA technology makes it possible to clone genes for basic research and commercial applications. 5. Restriction enzymes cut the DNA into a particular site. In our previous lesson, we learned how genetic materials are manipulated. One of the ways is through Genetic engineering. In Genetic engineering, genes are manipulated for practical purposes. Genetic Engineering DNA technology has launched a revolution in Biotechnology. DNA technology (via gene manipulation) makes it possible to clone genes for basic research and commercial applications. DNA technology is applied to areas ranging from agriculture to criminal law. One example of DNA technology is Genetic engineering. Genetic Engineering is the the artificial manipulation, modification, and recombination of DNA or other nucleic acid molecules in order to modify an organism or population of organisms. In the latter part of the 20th century, however, the term came to refer more specifically to methods of recombinant DNA technology, in which DNA molecules from two or more sources are combined either within cells or in vitro and are then inserted into host organisms in which they are able to propagate. The possibility for recombinant DNA technology emerged with the discovery of restriction enzymes in 1968 by Swiss microbiologist Werner Arber. Most recombinant DNA technology involves the insertion of foreign genes into the plasmids of common laboratory strains of bacteria. Plasmids are small rings of DNA; they are not part of the bacterium’s chromosome. Nonetheless, they are capable of directing protein synthesis, and, like chromosomal DNA, they are reproduced and passed on to the bacterium’s progeny. Thus, by incorporating foreign 6 DNA (for example, a mammalian gene) into a bacterium, researchers can obtain an almost limitless number of copies of the inserted gene. Furthermore, if the inserted gene is operative (if it directs protein synthesis), the modified bacteria will produce the protein specified by the foreign DNA. Editors of Encyclopedia Britannica (2020). What’s New Activity 1. Steps in Recombinant DNA Technology Directions: Arrange the steps in Recombinant DNA Technology in chronological order. Use numbers 1-7. A. Amplification Using PCR B. Isolation of Recombinant Cell C. Isolation of Genetic Material D. Obtaining or culturing the Foreign Gene product. E. Ligation of DNA Molecules. F. Restriction Enzymes Digestion G. Insertion of Recombinant DNA into Host What is It In this lesson, we shall outline the main steps in Recombinant DNA Technology and the importance of Recombinant DNA Technology. Recombinant DNA Technology Recombinant DNA technology refers to the joining together of DNA molecules from two different species that are inserted into a host organism to produce new genetic combinations that are of value to science, medicine, agriculture, and industry. Recombinant DNA (rDNA), on the other hand, is the general name for a piece of DNA that has been created by the combination of at least two different DNA strands. They are DNA molecules formed by laboratory methods of genetic recombination to bring together genetic material from multiple sources, creating sequences that would not otherwise be found in the genome. Aryal (2018). 7 Steps of Genetic Recombination Technology 1. Isolation of Genetic Material - Since DNA exists within the cell membrane along with other macromolecules such as RNA, polysaccharides, proteins, and lipids, it must be separated and purified which involves enzymes such as restriction enzymes. 2. Restriction Enzymes Digestion - The technique ‘Agarose Gel Electrophoresis’ reveals the progress of the restriction enzyme digestion. This technique involves running out the DNA on an agarose gel. On the application of current, the negatively charged DNA travels to the positive electrode and is separated out based on size. This allows separating and cutting out the digested DNA fragments. The vector DNA is also processed using the same procedure. 3. Amplification Using PCR - Polymerase Chain Reaction or PCR is a method of making multiple copies of a DNA sequence using the enzyme – DNA polymerase in vitro. It helps to amplify a single copy or a few copies of DNA into thousands to millions of copies. PCR reactions are run on thermal cyclers using the following components: 1.)Template – DNA to be amplified 2.)Primers – small, chemically synthesized oligonucleotides that are complementary to a region of the DNA. 3.) Enzyme – DNA polymerase 4.)Nucleotides – needed to extend the primers by the enzyme. 5.) The cut fragments of DNA can be amplified using PCR and then ligated with the cut vector. 4. Ligation of DNA Molecules – The purified DNA and the vector of interest are cut with the same restriction enzyme. This gives us the cut fragment of DNA and the cut vector that is now open. The process of joining these two pieces together using the enzyme DNA ligase is ligation. The resulting DNA molecule is a hybrid of two DNA molecules – the interest molecule and the vector. In the terminology of genetics this intermixing of different DNA strands is called recombination. Hence, this new hybrid DNA molecule is also called a recombinant DNA molecule and the technology is referred to as the recombinant DNA technology. 5. Insertion of Recombinant DNA into Host - In this step, the recombinant DNA is introduced into a recipient host cell mostly, a bacterial cell. This process is called transformation. Bacterial cells do not accept foreign DNA easily. Therefore, they are treated to make them competent to accept new DNA. The processes used may be thermal shock, Ca++ ion treatment, and electroporation. 6. Isolation of Recombinant Cells-The transformation process generates a mixed population of transformed and non-transformed host cells. The selection process involves filtering the transformed host cells only. For isolation of recombinant 8 cells from non-recombinant cells, a marker gene of the plasmid vector is employed. 7. Obtaining or culturing the Foreign Gene product - When you insert a piece of alien DNA into a cloning vector and transfer it into a bacterial cell, the alien DNA gets multiplied. The ultimate aim is to produce a desirable protein expression. The cells harboring cloned genes of interest are grown on a small scale in the laboratory. These cell cultures are used for extracting the desired protein using various separation techniques. Recombinant Human Growth Hormone Source: https://simplebiologyy.blogspot.com/2016/02/process-of-recombinantdna-technology-genetic-engineering.html Importance of Recombinant DNA Technology Recombinant DNA technology is playing a vital role in improving health conditions by developing new vaccines and pharmaceuticals. The treatment strategies are also improved by developing diagnostic kits, monitoring devices, and new therapeutic approaches. Synthesis of synthetic human insulin and erythropoietin by genetically modified bacteria and the production of new types of experimental mutant mice for research purposes are some one of the leading examples of genetic engineering in health. Likewise, genetic engineering strategies have been employed to tackle environmental issues such as converting wastes into biofuels and bioethanol, cleaning the oil spills, carbon, and other toxic wastes, and 9 detecting arsenic and other contaminants in drinking water. The genetically modified microbes are also effectively used in biomining and bioremediation. What’s More Activity 1. Enzymes in Recombinant DNA Technology Directions: Draw a Venn Diagram to differentiate DNA Polymerase with DNA Ligase. DNA Polymerase DNA Ligase Guide Questions 1. What is the first step in Recombinant DNA Technology? 2. In what step does the cut fragment of DNA and the cut vector are joined together? 3. What is the final step in Recombinant DNA Technology? 10 Activity 2. Modeling Bacteria Transformation Directions: Using the word choices provided in the boxes, fill in the numbered boxes with the steps of bacteria transformation and the lettered lines with the name of the structure next to them. Source:https://www.teachengineering.org/activities/view/uoh_genetic_lesson01_a ctivity1. Word Choices for Letters foreign DNA with the desired gene plasmid recombinant DNA Word Choices for Numbers bacteria transformed with recombinant plasmid plasmid cut with restriction enzyme DNA ligase joins sticky ends to form recombinant plasmid Guide Questions 1. What is the role of restriction enzymes in Recombinant DNA Technology? 2. What is the function of the DNA Ligase? 3. What is a Plasmid? 11 Activity 3 . Recombinant DNA Technology Directions: Read the choices from each numbered item in Column A and identify which is NOT included from these groups. Then classify it by choosing the correct answer in Column B. Note: To get one (1) point from this activity, two (2) responses must be answered correctly. Column A Column B A. Isolation of Genetic Material 1. a. Primers b. DNA Polymerase c. Agarose Gel d. PCR B. Restriction Enzymes Digestion 2. a. Transformation b. Ligation c. Recombinant DNA d. Ligase C. Amplification Using PCR D. Ligation of DNA Molecules 3. a. Gel Electrophoresis b. Protein expression c. Positive electrode d. Agarose Gel E. Insertion of Recombinant DNA into Host 4. a. Marker gene is employed. b. Filtering of the transformed host cell. c. Negatively charged DNA travels to the positive electrode. d. Mixed population of transformed and non-transformed cells. 5. a. DNA is separated based on size. b. Cutting out of digested DNA fragments. c. Negatively charged DNA travels to the positive electrode. d. Amplify a single copy of DNA into thousands or millions F. Isolation of Recombinant Cells G. Obtaining or culturing the Foreign Gene product Guide Questions 1. What are Primers? 2. What is the function of the PCR in Recombinant DNA Technology? 3. What is the charge of the DNA? 12 Activity 4. Complete the Steps in Recombinant DNA Technology Directions: Complete the figure below by supplying the missing Step in Recombinant DNA Technology. 1. Isolation of Genetic Material. 2. 3. Amplification Using PCR. 4. 5. 6. Isolation of Recombinant Cells 7. Guide Questions 1. What enzyme is used in DNA Ligation? 2. In what step in Recombinant DNA does Transformation occur? 3. What is the function of the Gel Electrophoresis in Recombinant DNA Technology? 13 Activity 5. Describing the steps in Recombinant DNA Technology Directions: Identify which step in Recombinant DNA Technology is involved. Event Steps in Recombinant DNA Technology 1. Running out the DNA on an agarose gel. 2. Harboring cloned genes of interest are grown on small a scale in the laboratory. 3. DNA must be separated and purified. 4. The recombinant DNA is introduced into a recipient host cell 5. Making multiple copies of a DNA sequence Guide Questions 1. What equipment makes multiple copies of the DNA? 2. How do DNA fragments separate in Gel Electrophoresis? 3. What enzyme is being used to separate and purify the DNA from the cell? Activity 6. Application of Recombinant DNA Technology Directions: Read and understand the situation below about the Humulin R. Answer the questions below after reading the article. Eli Lilly began producing insulin from animal pancreas but fell short of the demand, and the potency varied up to 25% per lot. The development of an isoelectric precipitation method led to purer and more potent animal insulin, decreasing the variation between lots to 10%. These discoveries led to the introduction of longeracting animal insulins in the market. Protamine zinc insulin lasted 24–36 hours. Isophane neutral protamine Hagedorn lasted 24 hours and could be mixed with regular insulin. The pharmacokinetics and effects of amorphous lente insulin (semilente, lente, and ultralente) depended on the proportion of zinc. In 1978, the first recombinant DNA human insulin was prepared by David Goeddel and his colleagues (of Genentech) by utilizing and combining the insulin A- and B- chains expressed in Escherichia coli. Thereafter, Genentech and Lilly signed an agreement to commercialize rDNA insulin. In 1982, the first insulin utilizing rDNA technology, Humulin® R (rapid) and N (NPH, intermediate-acting), were marketed. Guide Questions 1. Are you in favor of using the animal pancreas to replace the insulin in the human body? Why? 2. Based on the article, how does Recombinant DNA benefit humans? 3. What is Recombinant DNA Technology? 4. What is the function of a vector? 5. What are the common vectors in Recombinant DNA Technology? 14 What I Have Learned Directions: Fill in the blanks to complete the statements. 1. The general name for a piece of DNA that has been created by the combination of at least two strands of DNA is called _____________. 2. The process of introducing recombinant DNA into a recipient host cell is called _____________ . 3. Agarose Gel Electrophoresis involves running out the DNA on an _____________. 4. The process of joining the cut fragment of DNA and the cut vector together using an enzyme is called_____________ . 5. The _____________ is a method of making multiple copies of a DNA sequence using an enzyme. 6. Recombinant DNA technology refers to the joining together of _____________ from two different _____________ that are inserted into a host organism to produce new genetic combinations. 7. Ligation of DNA molecules uses_____________ to cut the vector. 8. An enzyme called _____________ helps to amplify a single copy or a few copies of DNA into thousands to millions of copies. 9. PCR reactions are run on _____________ to amplify a single copy or a few copies of DNA. 10. The small circular molecules which act as carriers for the DNA fragments are called _____________. What I Can Do Activity 1. Recombinant DNA Technology in our Life Directions: By giving examples, explain the importance of Recombinant DNA Technology in the given fields. 1. Health 2. Food 3. Environment 15 Assessment Directions: Read each statement carefully. Choose the letter of the correct answer. 1. Which enzyme is being used to amplify the DNA? a. Endonuclease b. DNA Helicase c. DNA Ligase d. DNA Polymerase 2. The following statements are all TRUE EXCEPT a. The DNA is located in the nucleus of the cell. b. Recombinant DNA is a combination of two different strands of DNA. c. DNA Ligase is used to amplify a single copy or a few copies of DNA. d. In Agarose Gel Electrophoresis the negatively charged DNA travels to the positive electrode and is separated out based on size. For question nos. 3-9, Arrange in order the steps in Recombinant DNA Technology. Use letters a to g 3. 4. 5. 6. 7. 8. 9. Amplification Using PCR Isolation of Genetic Material Insertion of Recombinant DNA into Host Obtaining or culturing the Foreign Gene product. Ligation of DNA Molecules. Restriction Enzymes Digestion Isolation of Recombinant Cells 10. Which of the following refers to chemically synthesized oligonucleotides that are complementary to a region of the DNA? a. Agarose Gel b. Primers c. Restriction Enzymes d. Vectors 11. Which of the following is TRUE? a. DNA is positively charged. b. Ligase is used to isolate the genetic material. c. Thermal Cycler cuts fragment of DNA and the vector. d. Gel Electrophoresis separates the DNA Molecule according to size. 16 For question nos. 12-15. Identify the steps in Recombinant DNA Technology that are being described in each statement. The choices are as follows: a. b. c. d. e. f. g. 12. 13. 14. 15. Amplification Using PCR Isolation of Genetic Material Insertion of Recombinant DNA into Host Obtaining or culturing the Foreign Gene product. Ligation of DNA Molecules. Restriction Enzymes Digestion Isolation of Recombinant Cells Marker gene of plasmid vectors is employed. The aim is to produce a desirable protein expression. The DNA molecule is separated and purified using enzymes. The recombinant DNA is introduced into a recipient host cell. 17 Additional Activity Activity 1.Implications of Recombinant DNA Technology Explain how recombinant DNA Technology poses a threat to a population or ecosystem. Rubric for the Essay Category Reflective Thinking Analysis Making Connections 4 3 2 The idea explains the student’s own thinking and learning processes, as well as implications for future learning. The idea is an in-depth analysis of the learning experience, the value of the derived learning to self or others, and the enhancement of the student’s appreciation for the discipline. The idea articulates multiple connections between this learning experience and content from past learning, life experiences and/or future goals. The idea explains the student’s thinking about his/her own learning processes. The idea attempts to demonstrate thinking about learning but is vague and/or unclear about the personal learning process. The idea attempts to analyze the learning experience but the value of the learning to the student or others is vague and/or unclear. The idea is an analysis of the learning experience and the value of the derived learning to self or others. The idea articulates connections between this learning experience and content from past learning experiences, and/or future goals. 18 The idea attempts to articulate connections between this learning experience and content from past learning experiences, or personal goals, but the connection is vague and/or unclear. 1 The idea does not address the student’s thinking and/or learning. The idea does not move beyond a description of the learning experience. The idea does not articulate any connection to other learning or experiences. What I 1. C 2.B 3.B 4.D 5.B Know 6. D 7. B 8. D 9. B 10. D 19 What’s In 1. True 2.True 3.False 4.True 5.True What’s More Activity 1 DNA Ligase - joins two pieces of DNA. DNA Polymerase - makes multiple copies of a DNA Similarity: Catalyst for Recombinant DNA Technology Guide Quest ions 1. Isolation of Genetic Material 2. Ligation of DNA Molecules. 3. Obtaining or culturing the Foreign Gene product. Activity 2 A. Plasmid B. Foreign DNA with desired genes C. Recombinant DNA 1. Plasmid cut with restriction Enzymes 2. DNA Ligase joins sticky ends 3. Bacteria transformed Guide Quest ions 1.Restriction Enzymes separate the DNA from the cell. 2.DNA Ligase joins two pieces of DNA. 3.Plasmid is the accessory ring of a bacterial chromosome. Activity 3 1. C, C 2. A, D 3. B, B 4. C, F 5. D, B Guide Quest ions 1. Primers are small, chemically synthesized oligonucleotides. 2. PCR makes a multiple copies of a DNA sequence. 3. Negatively charged. What’s New Activity 1 1. Isolation of Genetic Material 2. Restriction Enzymes Digestion 3. Amplification Using PCR 4. Ligation of DNA Molecules. 5. Insertion of Recombinant DNA into Host. 6. Isolation of Recombinant Cells 7. Obtaining or culturing the Foreign Gene product. What’s More Activity 4 2. Restriction Enzymes Digestion 4. Ligation of DNA Molecules. 5. Insertion of Recombinant DNA into Host . 7. Obtaining or culturing the Foreign Gene product. Guide Quest ions 1. DNA Ligase 2. Insertion of Recombinant DNA into Host Cell 3. Gel Electrophoresis runs out the DNA on an agarose gel. Activity 5 1. Restriction Enzymes Digestion 2. Obtaining or culturing the Foreign Gene product. 3. Isolation of Genetic Material 4. Insertion of Recombinant DNA into Host . 5. Amplification Using PCR Guide Quest ions 1. Thermal Cycler 2. Based on size 3. Restriction Enzymes Activity 6 Answers may vary Possible answer : 1. Yes, it helps to solve the problem on insufficient supply of insulin. 2. Recombinant DNA can produce artificial human insulin. 3.Recombinant DNA technology is the joining together of DNA molecules from two different species that are inserted into a host organism to produce new genetic combinations. 4. Vector is use to transfer foreign genetic material into a cell. 5. Bacteria and Virus Answer Key What I Have Learned 1. Recombinant DNA 2. Transformation 3. Agarose gel 4. Ligation 5. PCR 6. DNA molecules, species DNA ligase DNA polymerase Thermal Cycler Vector 7. 8. 9. 10. 20 What Can I Do Answers may vary Possible answer: Health- Artificial human insulin helps Diabetic people. Food- Golden Rice with a enhanced nutritional value. Environment- Pest resistant crops uses less pesticide, therefore less harm to the environment Assessment 1.D 11. A 2.C 12. G 3.C 13. D 4.A 14. B 5.E 15. C 6.G 7.D 8.B 9.F 10.B Additional Activity Answers may vary Possible answer: Introduction of genetically modified organisms which are product of Recombinant DNA Technology might change the gene frequency of a population that could lead to extinction of some organisms. References Aryal, Sagar, and Rashid Eltayeb. 2020. “Recombinant DNA Technology- Steps, Applications and Limitations: Molecular Biology.” Microbe Notes. https://microbenotes.com/recombinant-dna-technology-steps-applicationsand-limitations/. Bacteria Transformation - Activity.” 2019. TeachEngineering.org. June 20, 2019. https://www.teachengineering.org/activities/view/uoh_genetic_lesson01_ac tivity1. Griffiths AJF, Miller JH, Suzuki DT, et al. 2020. An Introduction to Genetic Analysis. 7th edition. New York: W. H. Freeman; 2000. https://www.ncbi.nlm.nih.gov/books/NBK21881/ Payal. 2016. “7 Main Stages of Recombinant DNA Technology.” Biology Discussion. https://www.biologydiscussion.com/dna/recombinant-dna-technology/7main-stages-of-recombinant-dna-technology/56320. Quianzon, Celeste C., and Issam Cheikh. 2012. "History of insulin." Journal of community hospital internal medicine perspectives 2, no. 2: 18701. Rabago, Lilia M, Joaquin, Crescencia C., and Lagunzad, Catherine Genevieve B. Functional. 2010.Functional biology: modular approach, Vibal Publishing House, Inc. Metro Manila, Philippines. The Editors of Encyclopedia Britannica. 2020. “Genetic Engineering.” Encyclopedia Britannica. Encyclopedia Britannica, Inc. https://www.britannica.com/science/genetic-engineering. 21 For inquiries or feedback, please write or call: Department of Education – Region III – Schools Division of Angeles City Jesus St. Pulungbulu, Angeles City, Pampanga, Philippines 2009 Telefax: (045) 322-5722;322-472; 888-0582;887-6099 Telefax: (632) 8634-1072; 8634-1054; 8631-4985 Email Address: [email protected]
