1. Irene is 10 years old and in the last few weeks, she suddenly experienced extreme tiredness, weight loss, and increased thirst. Her parents were concerned, so they took her to the doctor. Dr. Ross took Irene’s blood to test for her blood sugar. The results of the test indicated that she was a diabetic. When the doctor shared the results, Irene broke into tears. The combination of her sudden poor health and the news of her diabetes was too much for her to handle. “I don’t understand. What is diabetes and how can I get better?” The doctor felt bad for Irene. She said, “Irene, I can explain how this happened, and how we can fix it.” What Is the Problem? Irene is diabetic

2. What Is Diabetes? People with diabetes may not have enough insulin or may not be able to use it properly. Insulin is a hormone that controls the level of blood sugar (also called glucose) in your body. The sugar then builds up in the blood and overflows into the urine, passing out of your body unused. This deprives you of an important source of energy.

3. How Can Irene Get Better? All people with type 1 diabetes, and some people with type 2, need to take insulin to help control their blood sugar levels. Type 1 diabetes means your body doesn’t make any insulin. Type 2 diabetes means your body either doesn’t make enough insulin or doesn’t use it properly. The goal of taking insulin is to keep your blood sugar level in a normal range as much as possible so you’ll stay healthy. Over time, high blood sugar levels can cause serious health problems such as blindness and kidney failure.

4. How Can We Make Insulin? The first successful insulin preparations came from cows (and later pigs). In the 1980's technology had advanced to the point where we could make human insulin. The technology which made this approach possible was the development of recombinant DNA techniques. In simple terms, the human gene which codes for the insulin protein was cloned (copied) and then put inside of bacteria. A number of tricks were performed on this gene to make the bacteria want to use it to constantly make insulin. Big vats of bacteria now make tons of human insulin. From this, pharmaceutical companies can isolate pure human insulin

5. Describe how to make insulin for Irene.

6. Aim: How to Make Human Insulin? Step #1: Extract the DNA from a human cell.

7. Aim: How to Make Human Insulin? Step #2: Remove the plasmid from a bacterium. Small, circular DNA molecule presents in bacteria. What is a plasmid?

8. Aim: How to Make Human Insulin? Step #3:Use restriction enzymes to isolate a segment of (DNA) that contains the insulin gene

9. Aim: How to Make Human Insulin? Step #4: Treat the bacterial plasmid with the same restriction enzyme.

10. Aim: How to Make Human Insulin? Step #5: Binds the insulin gene with the opened plasmid to form a recombinant plasmid.

11. Aim: How to Make Human Insulin? Step #5: Binds the insulin gene with the opened plasmid to form a recombinant plasmid.

12. Aim: How to Make Human Insulin? Step #6: The recombinant plasmid is re-inserted back into the bacterium.

13. Aim: How to Make Human Insulin? Step #7: Bacteria clone into a large number of identical daughter cells.

14. Aim: How to Make Human Insulin? Step #8:The recombinant plasmid replicates as part of the bacteria’s DNA.

15. Aim: How to Make Human Insulin? Step #9. Insulin is produced by bacteria and extracted for human use.

16. Human Cell Gene for human insulin Recombinant DNA Gene for human insulin Sticky ends DNA recombination DNA insertion Bacterial Cell Plasmid Bacterial chromosome Bacterial cell containing gene for human insulin Figure 13-9 Making Recombinant DNA Gene for human insulin Recombinant DNA Gene for human insulinSticky ends Bacterial cell containing gene for human insulinPlasmid DNA recombinationDNA insertion

17. _____ Bacteria clone into a large number of identical daughter cells _____ Binds the insulin gene with the opened plasmid to form a recombinant plasmid. _____ Extract the DNA from a human cell. _____ Insulin is produced by bacteria and extracted for human use. _____ Remove the plasmid from a bacterium. _____ The recombinant plasmid is re-inserted back into the bacterium. _____ The recombinant plasmid replicates as part of the bacteria’s DNA. _____ Treat the bacterial plasmid with the same restriction enzyme. _____ Use restriction enzymes to isolate a segment of (DNA) that contains the insulin gene.

18. 1. Extract the DNA from a human cell. 2. Remove the plasmid from a bacterium. 3. Use restriction enzymes to isolate a segment of (DNA) that contains the insulin gene. 4. Treat the bacterial plasmid with the same restriction enzyme. 5. Binds the insulin gene with the opened plasmid to form a recombinant plasmid. The recombinant plasmid is re-inserted back into the bacterium. 6. Bacteria clone into a large number of identical daughter cells 7. The recombinant plasmid replicates as part of the bacteria’s DNA. 8. Insulin is produced by bacteria and extracted for human use.

19. Genetic engineering is a technique used by scientists to combine or splice genetic material from different organisms. Gene splicing involve changing the normal base sequences of DNA by removing a section of DNA and introducing another gene. The technique may involve the use of the bacterium E. coli. The bacterium has one large chromosome and several small plasmids, which are ring-shaped pieces of DNA found in the cytoplasm. Genetic engineers have been able to extract plasmids from E.coli. Restriction enzymes are used to cut the DNA of the plasmid at designated places in the base sequence. The same enzymes are used to cut a section of human DNA. This section of human DNA is then placed into the space in the cut DNA of the bacterial plasmid. The human DNA codes for the synthesis of a product such as human growth hormone. The spliced bacterial DNA, which now contains a piece of human DNA, is referred to as a hybrid. This hybridized plasmid is then transplanted into E.coli. When the bacterium reproduces, the hybrid DNA will replicate. The offspring will possess the ability to synthesize the human growth hormone

20. 1. What is a bacterial plasmid? 2. What is a hybrid plasmid? 3. Explain how genetic engineers remove sections from human DNA for splicing Into bacterial DNA 4. State one benefit of gene splicing 5. Explain why it is not necessary to continue splicing the gene for human growth hormone into E.coli once cultures of the bacteria with the spliced gene are established Small, ring-shaped pieces of DNA found in the bacterial cytoplasm. It is a recombinant plasmid. A bacterial plasmid that contains a piece of human DNA Genetic engineers use restriction enzymes to cut the human DNA and the bacterial plasmid, and insert the human DNA in the bacterial plasmid One benefit of splicing is to produce human hormones, like insulin, growth hormone, etc…, for sick people Because the transformed bacteria can be cloned and new bacteria with the recombinant plasmid

21. Describe the picture above following the numbered sequence. Assure to include these terms: restriction enzymes, DNA, plasmid, gene, recombinant plasmid, replication, cloning, insulin.

22. 4. The recombinant plasmid replicates and a large number of identical bacteria are cloned. They produce human insulin. Describe the picture above following the numbered sequence. Assure to include these terms: restriction enzymes, DNA, plasmid, gene, recombinant plasmid, replication, cloning, insulin 1.First, remove the DNA from a human body cell, then isolate the human gene of insulin using restriction enzymes. 2. Second, remove a plasmid from a bacterium and treated with the same restriction enzymes. 3. Third, bind the plasmid with the human gene to form a recombinant plasmid. Then the recombinant plasmid is re-inserted back into the bacterium