Teacher I LEARNING COMPETENCIES ● Outline the processes involved in genetic engineering (STEM_BIO11/12-IIIa-b-6) ● Discuss the applications of recombinant DNA (STEM_BIO11/12-IIIa-b-7) OBJECTIVES At the end of the lesson, learners shall be able to: ● K: discuss the applications of recombinant DNA; ● S: outline the processes involved in genetic engineering; and ● A: appreciate the importance of recombinant DNA by listing its applications in daily life. Directions: Identify the word(s) described in each item by filling in PRE TEST the correct letters. Write your answers in your notebook. PRE TEST Directions: Identify the word(s) described in each item by filling in the correct letters. Write your answers in your notebook.
1. It produces genetically engineered organisms.
R_CO_B_N_NT DNA 2. Genetically modified crops A_R_C_LT_R_L APPLICATIONS PRE TEST 3. This is an extrachromosomal, small and circular DNA molecule, which replicate independently. P_AS_I_ 4. It is a molecule which contains all information necessary to build and maintain an organism. D_O_YR_B_N_C_IC AC_D PRE TEST 5. Production of chemical compounds of commercial importance _ND_STRI_L A_PL_C_TI_N 6. It is a process using recombinant DNA technology to alter the genetic makeup of an organism. G_N_T_C ENG_N_ER_NG PRE TEST 7. It is the basic unit of heredity. G_N_S 8. It produces copies of genes or segments of DNA. G_N_ C_O_I_G PRE TEST 9. An individual that supplies living tissue to be used in another body. D_N_R _RG_N_S_ 10. A DNA nucleotide used as a vehicle to artificially carry foreign gene/materials. V_CT_R G_N_M_ What are the roles of the DNA, genes and proteins in a given trait?
Would the manifestation of a trait
be affected once the DNA nor the genes are altered? Why? What do you think are the objectives of scientists in genetic engineering? TERMINOLOGIES
DONOR ORGANISM -the source of the gene that will be used in genetic engineering process.
GENETIC VECTOR - often a virus or a plasmid that is used to carry a desired DNA sequence into a host cell as part of a molecular cloning procedure. VECTOR GENE OF INTEREST - the desired gene which will be introduced into the vector to be amplified. RESTRICTION ENZYMES -a protein produced by bacteria that cleaves/cut DNA at specific sites along the molecule. It cleaves DNA into fragments at or near specific recognition sites within the molecule known as restriction sites.
STICKY ENDS - an end of a DNA double helix at which a few nucleotides of one strand extend beyond the other. . DNA LIGASE - type of enzyme that facilitates the joining of DNA strands together by catalyzing the formation of a phosphodiester bond.
AMPLIFICATION - the action of making multiple copies of a gene or DNA sequence Recombinant DNA Recombinant DNA is made by mixing DNA from two different sources. A Brief History of Recombinant DNA The use of recombinant (r)DNA technology to produce genetically engineered organisms started in the early 1970s with the pioneering transfer of genes between bacteria of the same Escherichia coli (E. coli) species. Following these successful pilot experiments, in 1978 Cohen and colleagues progressed to transfer an insulin synthesis gene into a plasmid of E. coli, with that producing the first genetically modified organism (GMO). A Brief History of Recombinant DNA By 1982, this protocol received full approval from national drug regulatory authorities, notably the US Food and Drug Administration, thereby enabling the economically viable mass production of human insulin, a hormone that regulates blood sugar levels and is made naturally by beta cells in the pancreas. This facilitated the widespread commercial availability of insulin at a price affordable to patients with the metabolic disorders’ types 1 and 2 diabetes mellitus, A Brief History of Recombinant DNA who either fail to produce or to metabolize sufficient insulin. This proof of principle demonstration of the translational medical benefits of genetic modification pioneered a trend in biotechnology for molecular cloning methods to transfer genes expressing desirable traits into another host organism thereby producing favorable characteristics. Example: Building a recombinant plasmid Let's see how restriction digestion and ligation can be used to insert a gene into a plasmid. Suppose we have a target gene, flanked with EcoRI recognition sites, and a plasmid, containing a single EcoRI site: Cutting and pasting DNA Our goal is to use the enzyme EcoRI to insert the gene into the plasmid. First, we separately digest (cut) the gene fragment and the plasmid with EcoRI. This step produces fragments with sticky ends: Next, we take the gene fragment and the linearized (opened-up) plasmid and combine them along with DNA ligase. The sticky ends of the two fragments stick together by complementary base pairing: Once they are joined by ligase, the fragments become a single piece of unbroken DNA. The target gene has now been inserted into the plasmid, making a recombinant plasmid. REMEMBER:
If the restriction enzymes cuts within the sequence of the gene of
interest or within essential plasmid vector components, such as the origin of replication, selectable marker or promoter region, those interrupted sequences will no longer be functional or code for functional products.
It is also important to ensure that your restriction enzymes do not
cut in multiple places on the plasmid, as this could result in your gene of interest being inserted in the wrong location or generation of an incomplete plasmid. The same restriction enzyme should be used when cutting out a gene of interest and cutting the plasmid vector where the DNA will be inserted so that the sticky ends of both will match up. Bacterial transformation and selection Plasmids and other DNA can be introduced into bacteria, such as the harmless E. coli used in labs, in a process called transformation. During transformation, specially prepared bacterial cells are given a shock (such as high temperature) that encourages them to take up foreign DNA. Bacterial transformation and selection A plasmid typically contains an antibiotic resistance gene, which allows bacteria to survive in the presence of a specific antibiotic. Thus, bacteria that took up the plasmid can be selected on nutrient plates containing the antibiotic. Bacteria without a plasmid will die, while bacteria carrying a plasmid can live and reproduce. Each surviving bacterium will give rise to a small, dot-like group, or colony, of identical bacteria that all carry the same plasmid. Protein production
Once we have found a bacterial colony with the right plasmid, we can grow a large culture of plasmid-bearing bacteria. Then, we give the bacteria a chemical signal that instructs them to make the target protein. What are the important applications of recombinant DNA? Medicinal
Agricultural Industrial
Important Applications I. Agricultural applications ⚫ (applications in crop improvement) 1. Distant hybridization -also known as wide hybridization, is the process of breeding two different species, genera, or higher- ranking taxa. It can result in the creation of new species, the development of fertile strains, and the improvement of characteristics in new populations. I. Agricultural applications 2. Development of transgenic plants - Genetically transformed plants (resistance to pests, disease, etc.) that contain foreign genes are called transgenic plants. BT-cotton resistant to bollworms is a glaring example. GOLDEN RICE Shoot Borer II. Medicinal applications ⚫ (applications in medicines) 1. Production of antibiotics -Penicillium and Streptomyces are used for the mass production of famous antibiotics penicillin and streptomycin. 2. Production of the hormone insulin - Insulin, a hormone used by diabetics, is usually extracted from the pancreas of cows and pigs. II. Medicinal applications
3. Production of vaccines -Vaccines are now produced by the transfer of antigen- coding genes to disease-causing bacteria. 4. Production of interferon - Interferons are virus-induced proteins produced by virus-infected cells. Natural interferon is produced in very small quality from human blood cells. II. Medicinal applications 5. Production of enzymes - the enzyme urokinase, which is used to dissolve blood clots, has been produced by genetically engineered microorganisms. 6. Gene therapy Genetic engineering may one day enable medical scientists to replace the defective genes responsible for *hereditary diseases with normal genes. II. Medicinal applications 7. Diagnosis of disease - Most of these involve the construction of probes. By testing the DNA of prospective genetic disorder carrier parents, their genotype can be determined and their chances of producing an afflicted child can be predicted. II. Medicinal applications 8. Production of transgenic animals -Animals that carry foreign genes are called transgenic animals. Ex: Cow, sheep, and goat – therapeutic human proteins in their milk. A. Spider goat: gene from spider inserted into goat. Goats makes silk of the spider web in their milk. Flexible, stronger than steel. Used in bullet proof jackets. B. Glow-in-the-dark cats Scientist used a virus to insert DNA from jellyfish The gene made the cat produce a fluorescent protein in its fur. III. Industrial applications -In industries, recombinant DNA technology will help in the production of chemical compounds of commercial importance, improvement of existing fermentation processes, and production of proteins from wastes. Specially developed microorganisms may be used even to clean up the pollutants. III. Industrial applications Bioremediation It is a branch of biotechnology that uses living organisms, like microbes and bacteria, to clean up contaminated areas. rDNA can be used to genetically modify organisms to break down pollutants in the environment. Recombinant microorganisms: These organisms can be used to digest xenobiotics through degradative genes. Genetically modified microbes: These microbes can be used to break down pollution. III. Industrial applications Activity 2: Directions: Classify the different applications of recombinant DNA technology. Write A- agricultural, M –medicinal & I-industry 1. Golden rice 2. Bt corn 3. Human Growth Hormone 4. Human Insulin 5. Fermentation of food products What is genetic engineering? Genetic engineering
● It is the process of using rDNA technology to alter the
genetic makeup of an organism. Traditionally, humans have manipulated genomes indirectly by controlling breeding and selecting offspring with desired traits. ● Since ancient times, the practice of genetic engineering had begun Five Basic Processes in Genetic Engineering DNA Extraction
Gene Cloning
Gene Design
Transformation or Gene Insertion
Backcross Breeding Five Basic Processes in Genetic Engineering
DNA Extraction
Gene Cloning
Gene Design
Transformation or Gene Insertion
Backcross Breeding Five Basic Processes in Genetic Engineering DNA Extraction
Gene Cloning
Gene Design
Transformation or Gene Insertion
Backcross Breeding Five Basic Processes in Genetic Engineering DNA Extraction
Gene Cloning
Gene Design
Transformation or Gene Insertion
Backcross Breeding Five Basic Processes in Genetic Engineering DNA Extraction
Gene Cloning
Gene Design
Transformation or Gene Insertion
Backcross Breeding Five Basic Processes in Genetic Engineering\ DNA Extraction
Gene Cloning
Gene Design
Transformation or Gene Insertion
Backcross Breeding Five Basic Processes in Genetic Engineering\ Five basic steps in crop genetic engineering: DNA Extraction
1. DNA extraction – DNA is extracted from an organism known to
have the desired trait. Gene Cloning 2. Gene cloning – The gene of interest is located and copied. 3. Gene modification – The gene is modified to express in a desired way by altering and replacing gene regions. Gene Design 4. Transformation – The gene(s) are delivered into tissue culture cells, using one of several methods, where hopefully they will land in the nucleus and insert into a chromosome. Transformation or Gene Insertion 5. Backcross breeding – Transgenic lines are crossed with elite lines to make highyielding transgenic lines. Backcross Breeding Genetic Engineering Techniques 1. Artificial selection A. selective breeding B. hybridization C. inbreeding 2. Cloning 3. Gene splicing 4. Gel electrophoresis 1. artificial selection: breeders choose which organism to mate to produce offspring with desired traits. A. Selective breeding - when animals with desired characteristics are mated to produce offspring with those desired traits. ● Passing important genes to the next generation. ● Example: Champion racehorses, cows with tender meat, large juicy oranges on a tree. ⚫Selective breeding occurs when you choose the best male and female to breed. ⚫This allows you to fine-tune and control the traits ⚫The offspring or babies will then have the best traits. ⚫Then you continue to breed those organisms with the best traits, and those traits will be maintained. B. Hybridizations - two individuals with unlike characteristics are crossed to produce the best in both organisms. ● Example: Luther Burbank created a disease- resistant potato called the Burbank potato. Examples of hybridization: Lion + Tiger = Liger C. Inbreeding - breeding of organism that genetically similar to maintain desired traits. - It keeps each breed unique from others. Risk: since both have the same genes, the chance that a baby will get a recessive genetic disorder is high. 2. Cloning - creating an organism that is an exact genetic copy of another. Clone: group of cells or organisms that are genetically identical as a result of asexual reproduction - They will have the same exact DNA as the parent. How is cloning done? ● A single cell is removed from a parent organism. ● An entire individual is grown from that cell. ● Remember one cell has all the DNA needed to make an entire organism. ● Each cell in the body has the same DNA, but cells vary because different genes are turned on in each cell. Benefits of cloning: 1. you can make exact copies of organisms with strong traits. 2. Increase food supply 3. Medical purposes: clone organs for transplants. 4. Bring back or stop species from going extinct.
Saber Tooth Tiger extinct
Risks of cloning: 1. Decreases genetic diversity 2. If one of your clones gets a disease, they all get it: same immune system. 3. Inefficient: high failure rate: 90%+ 4. Expensive 3. Gene splicing - DNA is cut out of one organism and put into another organism Example: the human insulin gene can be removed from a human cell. Gene therapy: when disease causing genes are cut out and good gene are inserted. ⚫Restriction enzymes are used to cut out bad genes. ⚫Viruses are used to insert good genes. ⚫Not approved for human use yet. ⚫Some possible side effects. 4. Gel electrophoresis - a technique used to compare DNA from two or more organisms. Why compare DNA: 1. Find your baby’s daddy 2. Who committed a crime. 3. How closely species are related. How is electrophoresis done? A. The DNA is cut into fragments with a restriction enzyme. B. The cut DNA is then put into the wells of a machine filled with gel. ⚫The gel is spongy and the DNA squeezes through the pores. C. The machine is plugged in and the fragments get separated based on their size. ⚫The smaller fragments move further than the large. DNA collected from the crime scene would be an exact match to DNA from the culprit. DNA fingerprinting can also be used to determine paternity. In the case of paternity, the DNA of a child would exactly match the DNA of either of the two parents. Genetic engineering creates organisms with recombinant DNA. Recombinant DNA - when DNA is combined from at least two organisms. Which techniques create recombinant DNA? 1. Sexual reproduction: natural 2. selective breeding 3. Hybridization 4. Gene splicing Exercise 2: Genetic Engineering Techniques Directions. Distinguish the techniques in genetic engineering as based on the situation and examples given. Write the letter of the choices. A. Artificial selection B. Selective breeding C. Hybridization D. Inbreeding E. Cloning F. Gene splicing G. Gel electrophoresis: analyzing DNA A.Artificial selection E. Cloning B.Selective breeding F. Gene splicing C.Hybridization G. Gel electrophoresis: analyzing DNA D.Inbreeding
____1. When animals with desired
characteristics are mated to produce offspring with those desired traits. Dachshund were once bred to hunt badgers and other burrowing animals. A.Artificial selection E. Cloning B.Selective breeding F. Gene splicing C.Hybridization G. Gel electrophoresis: analyzing DNA D.Inbreeding
____2. Creating an organism that is an
exact genetic copy of another. They will have the same exact DNA as the parent. A.Artificial selection E. Cloning B.Selective breeding F. Gene splicing C.Hybridization G. Gel electrophoresis: analyzing DNA D.Inbreeding
____3. DNA is cut out of one organism and
put into another organism. A trait will be transferred from one organism to another. A.Artificial selection E. Cloning B.Selective breeding F. Gene splicing C.Hybridization G. Gel electrophoresis: analyzing DNA D.Inbreeding
____4. Luther Burbank created a disease
resistant potato called the Burbank potato. He crossed a disease resistant plant with one that had a large food producing capacity. A.Artificial selection E. Cloning B.Selective breeding F. Gene splicing C.Hybridization G. Gel electrophoresis: analyzing DNA D.Inbreeding
____5. A technique used to compare DNA
from two or more organisms. A.Artificial selection E. Cloning B.Selective breeding F. Gene splicing C.Hybridization G. Gel electrophoresis: analyzing DNA D.Inbreeding
____6. Breeding of organism that genetically
similar to maintain desired traits. It keeps each breed unique from others. Activity: Forensics Forensics is simply defined as the application of science in law. It uses scientific techniques such as fingerprint identification, autopsy, chemical analysis, and DNA technology that includes DNA fingerprinting or profiling, to get reliable data. This valuable information helps investigators arrive at a solution to a complicated criminal case. Crime Scene Analysis: Case #1: A stolen car was found on the side of the road with an empty bottle of Sprite. Detectives were able to collect enough DNA from the saliva left on the top of the bottle to create a DNA fingerprint. Below are the DNA fingerprints from the crime scene and two suspects who were seen near the abandoned car. Case #2: A mother files a lawsuit for child support against a man she claims is the father of her child. The man claims that he has no children and does not even know the woman and so shouldn’t have to pay child support. Below are the DNA fingerprints of the child, the mother and the man. Remember, children receive half of their DNA from their mother and the other half from their father. Analysis:
Who stole the car?
How do you now? Analysis: 1. Could the man be the father of this child?
