BIOSYNTHESIZING EVOLUTION FOR THE

ENHANCE PRODUCTION OF CAROTENOIDS IN YEAST

Saniyyah ThomasGilliam Collegiate Academy

Dallas ISD

Dr. Katy Kao and GroupAssistant Professor

Department of Chemical EngineeringTexas A&M University

SUMMER RESEARCH OVERVIEW

1ST STEP: Engineer a generation of fluorescent yeast strains using a plasmid carrying these genes

2nd STEP: Engineer a

generation of carotenoid producing

fluorescent yeast strains again using a

plasmid carrying these genes

3rd STEP: Remove ctt1 genes by

replacing it with an antifungal resistant

gene again by using a plasmid; This is done to

remove a potential competitive response to enhanced

production of carotenoids once

the selective pressure of adding H2O2 is introduced

Miniprep of plasmid DNA

Multiple Chemical reactions take place in this process Three Solutions are prepared

Solution II: NaOH ( this akali condition allows for maximum release of plasmid DNA without release of chromosomal DNA, & without irreversibly denaturing plasmid DNA)

Stop lysis reaction by neutralize

Solution III: acetic acid w/ concentrated HCl (neutralize, forms a salt which precipitates out denatured chromosomal DNA, protein, and SDS, and allows the smaller plasmid DNA to renature and stay in solution)

Associated Chemistry STAAR/EOC Standards 2012

(C.10) Science concepts. The student understands and can apply the factors that influence the behavior of solutions. The student is expected to

(G) define acids and bases and distinguish between Arrhenius and Bronsted-Lowry definitions and predict products in acid-base reactions that form water; Supporting Standard (H) understand and differentiate among acid-base reactions, precipitation reactions, and oxidation-reduction reactions; Readiness Standard

(I) define pH and use the hydrogen or hydroxide ion concentrations to calculate the pH of a solution; and Supporting Standard

(J) distinguish between degrees of dissociation for strong and weak acids and bases. Supporting Standard

NEUTRALIZING THE PROBLEMConceptual Design of the Project

Millions of people around the world suffer from conditions known as acid reflux and acid indigestion.

In this design challenge, students will take the engineering design process and apply it to their knowledge of acid and bases to come up with a short term as well as long term plan to help a sufferer.

For their short term solution, students will ultimately conduct research; design a method and protocols that will prove which brands of antacids that are currently on the market are the most effective.

Students will then establish a long term, sensible, and cost effective diet for a sufferer.

They will have to research various foods and drinks and their pH, design methods and protocols, and then by analyzing their results, they will then come up with a diet they can follow for a month. A “Foods to Avoid” list will be created as well.

Students will have to work thru the engineering design process and established at least four different methods of testing the pH of various antacids, foods, and drinks.

Materials we use in our daily life will be identified as acidic, basic, or neutral. Students will work in groups to perform simple experiments, make observation and prediction, thus enhancing their manipulative, decision making, and problem-solving skills.

DAY 1Project BlueprintDay 1 (block scheduling with an 1 ½ class periods)

Students will be re-introduced to the field of Engineering in early January (pre-test and summer research project will be discussed briefly at the beginning of the school year). A power point presentation will be viewed that combines my summer research and experience overview, definition of engineers and the profession, different industrial engineering opportunities, and the12-step engineering design process. An activity that allows the students to compare and contrast the engineering process with the scientific process and a hands on activity with manipulative (i.e. magnets, Legos) will be completed.

Courtesy of Project Lead The Way

DAY 2Day 2  The mini prep portion of my

summer lab research project will be highlighted, specifically the acid/base chemical reaction process that takes place.

This will lead into the discussion about acids and bases.

Definitions and vocabulary will be discussed, various methods of testing pH will be demonstrated, and real world examples.

Students will do a white board activity identifying acid and bases, predicted products in aqueous solutions.

Daily assessment will be given.

LITMUS PAPER

PHENOLPHTHALEIN

INDICATOR STRIPS

pH METER

VERNIER pH PROBES

DAY 3Day 3 Students will begin and complete the Acid and Base Lab. Due at the end of the class (if on a Friday, it will be due that following week).

DAY 4Day 4 

Scenario: One day, Kisha ate a big hamburger, French fries, and sprite for dinner and went to bed 15 minutes later. She woke up later that night with an upset stomach. She knew what it was because this happens quite often. She took some antacid that night. It took a while but eventually the pain subsided and she fell asleep again. The next morning, Kisha went to the doctor. He told her that for a long term cure she would have to change her diet but for immediate help she would need to find the best antacid on the market that can subdue her pain quickly. Why did Kisha get a stomach ache? Can you describe what happened to Kisha? Why did she take an Antacid?   

Students will be grouped and presented with the project where they will spend this day on the first three steps

of the engineering design process. Step 1: Define the problem; Step 2: Brainstorm; and Step 3: Research

and generate ideas

 

DAY 5Day 5 

This day will be spent working on the design process steps 4-7. Students will complete Step 4: Identify criteria and constraints; Step 5: Explore possibilities; Step 6: Select an approach; and Step 7: Develop a design proposal

DAY 6Day 6 Students will next complete design process steps 8-11. Students will Step 8: Model or prototype; Step 9: Test and evaluate; Step 10: Refine; and Step 11: Create or make

DAY 7, 8, 9Step 12: Communicating results

On the due date, each group must give a 15 minute presentation on their project.

They must explain the entire 12-step design process for both their short and long term solutions and based on their results present the best brand of antacids, the month long diet plan, & “AVOID LIST” for Kisha.

Visuals and models will be required

SHORT-TERM ANTACID RESULTS

FOODS TOAVOIDS/ BUY

LONG-TERM SOLUTION MEAL PLAN

PRE/POST-TEST1. Acid A and acid B are of equal concentration and are tested with a conductivity apparatus. When the electrodes are placed in acid A, the bulb glows dimly. When they are placed in acid B, the bulb glows more brightly. Which of the following is true? A. Acid A is stronger than acid B. B. Acid B is stronger than acid A. C. Acid A and acid B are of equal strength. D. No comparison of strength can be made from the results.

2. The neutralization of a strong acid by a strong base always involves the products A. water and a salt. B. an anion and a salt. C. water and an ion. D. weak acid and a strong base.

ACKNOWLEDGEMENTS TAMU E³ Program National Science

Foundation Nuclear Power Institute

Dr. Katy Kao (Mentor)

Luis H. Reyes (Ph.D. student)

Other lab members:• Priscila Almario• Mian Huang• Yuqi Guo• Jesus • Miguel Ramirez

(partner)

REFERENCES 1. Namitha, K. K. & Negi, P. S. Chemistry and biotechnology of carotenoids. Critical Reviews in Food Science and Nutrition 50, 728–760 (2010).

2. Kao, K. Visualizing evolution in real time (VERT) - application in biocatalyst improvement for carotenoid production 1, 1-3 (2012).

3. http://www.chemguide.co.uk/physical/acidbaseeqia/indicators.html

4. http://forum.grasscity.com/absolute-beginners/163085-soil-ph-testing-methods.html

5. http://grass-roots-science.info/project_isidore

6. http://explow.com/scientific_method