Gene Cloning with Plasmids- Insulin Production Zachary Sweatman and Haley Eberhart Mrs. Sintich AP Bio DNA Technology Project Due: 4/11/16

H and Z Insulin Production

What is it?: DNA Cloning DNA Cloning is the use of DNA manipulation procedures to produce multiple copies of a single gene or segment of DNA. In 1973, two scientists named Herbert Boyer and Stanley Cohen developed a way to take DNA from one organism and put it in the DNA of bacterium. This process is called recombinant DNA technology. Boyer and Cohen determined to clone DNA correctly, there are four steps to the process:

Step 1: Isolate the Gene The first step is isolation of the plasmid from a bacterium, and also the isolation of a particular gene of interest from another organism. In our case specifically, the insulin producing gene. The gene for producing HUMAN insulin protein is isolated. The gene is part of the DNA in a human chromosome. The gene can be isolated and then copied so that many insulin genes are available to work with.

Step 2: Preparing the Target DNA and insertion of DNA into the plasmid First, a circular piece of DNA called a plasmid is removed from a bacterial cell. Special proteins are used to cut the plasmid ring open. With the plasmid ring open, the gene for insulin is inserted into the plasmid ring and the ring is closed. The human insulin gene is now recombined with the bacterial DNA plasmid. Now the bacteria have the gene for producing insulin and can produce massive amounts of it as the bacteria multiply.

Step 3: Insertion of plasmid back into Dna and cell multiplication The bacterial DNA now contains the human insulin gene and is inserted into a bacteria. Scientists use very small needle syringes to move the recombined plasmid through the bacterial cell membrane. The bacterium is now considered “recombinant.” Many plasmids with the insulin gene are inserted into many bacterial cells. The cells need nutrients in order to grow, divide, and live. While they live, the bacterial cell processes turn on the gene for human insulin and the insulin is produced in the cell. When the bacterial cells reproduce by dividing, the human insulin gene is also reproduced in the newly created cells.

Step 4: Target cells reproduce Once the plasmids reproduce in the bacteria cells, and there are plenty of bacteria cells with plenty of plasmids in them, the target cells are gathered up after reproduction of the entire bacteria. The cells that are made keep on producing the protein for whatever is being coded for, in our case insulin, and can be purified after being harvested for uses in medicine, specifically saving the lives of diabetics. Fun Fact: Insulin used to produced and purified biosynthetically from cows and pigs for humans, but now that great companies like H and Z Insulin are around, that practice is almost completely died out because it is outdated and can have medical complications.

Current uses for DNA Cloning There are hundreds of different uses for DNA cloning: pest resistant plants, bacteria to clean up toxic waste, HGH (human growth hormone) treatments, and a plethora of medical uses INCLUDING what our amazing company, “H and Z Insulin,” does which saves the lives of thousands of people yearly that are fighting the growing battle of diabetes. Millions of people with diabetes now take human insulin produced by bacteria or yeast (biosynthetic insulin) that is genetically compatible with their bodies, just like the perfect insulin produced naturally in your body.

Bioethics, law, and society Insulin production through DNA cloning has a long list of pros including: safer more pure insulin, low cost in comparison to using cow and pig insulin, it can be mass produced in short periods of time, and it helps save thousands of people that are struggling with a disease known as diabetes, where they need insulin. I don’t know of any cons when talking about insulin production through DNA cloning. But if we step back and take a look at DNA cloning as a whole there are problems that occur. For one, since the DNA is exactly the same we would be preserving genetic weaknesses if there are any. Also there would be less introduction into new traits and genes therefore drastically slowing the evolution of a species.

Bioethics, law, and society Cell aging can also be a problem from DNA cloning, because the natural degeneration of cells results in corrupted source material. There is a chance that if the genetic material were corrupted from cell aging that the clone would have a shortened lifespan because nothing can be done about cell degeneration. The only ethical dilemma’s surrounding DNA cloning that I can think of is should humans be cloned to ween out weak traits such as diseases and undesirable traits? DNA cloning of animals is not illegal, but the cloning of a human is illegal in the United States. There is potential for DNA cloning of humans to do a lot of good, but we also have to take into consideration the question of is it ethical to genetically manipulate humans to make them “better.”

Case studies Stephanie Myers, a patient of our company, began showing symptoms of diabetes at age 19. She tried exercising and dieting to treat her symptoms but unfortunately the symptoms got worse and she became completely dependent on insulin injections. The insulin that was administered to Stephanie was created in our lab by recombinant DNA bacteria that was genetically manipulated to produce the insulin needed to save her life.