PGLO Question - Help Please! Very urgent!

[Nerdygirl]

I doing my first BIG science fair for my Science bound program. I have chosen to do Bactria Transformation using pGLO as my project. The problem here is I cant seem to find an attention grabber and good question to use for this project/experiment. Can you please help me by giving me some suggestions on what I could use? It that would me SOO much so i can finally begin doing it. Also do you think that is a good science fair project to do?

Thank you,

[donnahardy2]

Hi,

I apologize for the delay in responding to your question.

It sounds like you are interested in doing this very excellent project.

https://www.sciencebuddies.org/science- ... p013.shtml

This is a very challenging project, but you will learn a lot doing it. However, you are asking a good question, because you probably do need to make sure you are doing a controlled experiment. For a science project, you should be doing an experiment to answer a question. For this project, you might want to optimize the growth media, the concentration of antibiotics, the temperature, or some other parameter to determine if there is an effect on the yield of the product. If you just follow the protocol and purify the protein, then the project will be more of a demonstration, rather than an experimental project.

Do you know if there is a step in the basic protocol that causes a problem that needs to be optimized? What have you learned from your background reading?

Please let me know exactly what you have done so far, and I'll make some specific suggestions.

Donna Hardy

[Nerdygirl]

Well I haven't exactly started, but I'm trying to as soon as I have a question. Right now I have just been watching lectures and reading online to learn more about DNA, pGLo, GFP ..etc. I understand it very well since, I've done this experiment before, but hadn't really paid much attention on optimizing the growth media. So you are asking me to setup or change the procedure to determine an effective way to grow the media? Please explain more.

Thank you,

[donnahardy2]

Hi,

Yes, ideally you should design an experiment that will help you discover something new about pGlo. I recommend that you read through the steps for doing a science fair project from the Science Buddies Website:

https://www.sciencebuddies.org/science- ... ndex.shtml

You are at the stage of asking a question that will be the basis for your entire project, and this is a very critical step. Your question should be worthwhile and interesting for you to do:

https://www.sciencebuddies.org/science- ... tion.shtml

It is helpful that you have done the procedure before and have been doing background reading. Based on your experience, was there anything difficult about the procedure that you think could be improved? Based on your knowledge of the subject, is there anything that you don’t understand that you would like to discover? Think about your subject, and I’m sure you will think of a question. You will answer the question by designing a carefully controlled experiment.

I hope this is helpful. Let me know if you have thought of a question, or if you need more ideas.

Donna Hardy

[Nerdygirl]

Okay, I understand what you mean by that. Well I had an idea maybe I could do 5 different procedures, the first following the correct procedure, in the second one I could change the temperature 5 degrees higher then needed, the third 5 degrees lower than needed, the fourth 10 degrees more then needed, and the fifth 10 degrees less then needed. I want to do this because when I did this lab experiment last time i noticed some groups had a ton of colonies on their plates, some had very few, and some had non at all and I want to know why is that? I haven't decided if i will change the temperature or another control but its a start.

Thank you,

[donnahardy2]

Hi,

Excellent! Your observation about the variation in numbers of colonies between the various groups of students gives you a good reason to investigate experimental parameters that could affect results. And, your idea to investigate the effect of temperature is a good one as you will be able to design a well controlled experiment and obtain measurable results.

I believe that the E. coli used in the pGlo experiment is incubated at 37 degrees Centigrade. Do you have access to 5 different incubators that can maintain a constant temperature for 24-48 hours? If not, how many incubators do you have access to? You will want to set up all of the plates for the experiment at the same time so everything is identical except the temperature. Using 5 different temperatures would be good, but very ambitious. It would be OK if you could 3 temperatures, or even just 2. For example, if you have two incubators, you could use one 42 degrees Centigrade, one at 37 and incubating one set of plates at ambient temperature (about 22 degrees Centigrade).

But first, before you decide what temperatures to use do some background reading and find out what the range of temperatures is compatible with E. coli. What is the effect of temperature on the growth of bacteria? Has anyone ever noticed an effect of temperature on the production of green fluorescent protein? The idea behind the background reading is to find out what other researchers have discovered so you can design the best possible experiment.

You should also think about how many plates you will use. Ideally you will want to do your entire experiment 2 times to find out if results are reproducible. And, ideally, you will want to include enough replicates in each experiment to ensure that your results are statistically significant. The minimum number of plates for each temperature should be two (duplicate), and including a higher number (3-5) would be better. Let me know if you need more information about the statistical analysis of results.

You are making great progress. Let me know if you need additional explanation.

Donna Hardy

[Nerdygirl]

Hi,

Yes, I'm going to start on my background information. I understand how to do the procedure and I'll probably do 3 duplicate plates at a time. About the incubators, I currently have none. In order for me to do this project I might have to drive up to a near by college and use their equipment. I was wondering where I could get or borrow used ones. I know they are pretty expensive but sometimes companies give out used ones to schools for free. Do you now of anyplace that would probably give me one? That would be really big help.

Thank you!

[donnahardy2]

Hi,

You do need to use the best possible equipment that you can use to control the temperature. Incubators are too expensive to purchase, so I recommend calling the local college and ask if you can use their lab space and incubators for your experiment. There should not be a problem if you schedule the experiment when classes are not using the incubators, but you do need permission from the lab manager.

If there is a local used scientific equipment company, you could contact the company and ask to borrow an incubator or two for a couple of weeks.

Is your project actually due on August 15? I don’t think you have time to make your own incubators or to find a company who is willing to donate, so if you can’t locate at least one incubator, then you will need to find a different parameter to investigate.

Running plates in triplicate is an excellent plan and you will be able to do the analysis to show that your results are statistically significant.

Donna

[Nerdygirl]

Okay thank you so much.
My science Fair is not for a long while but I have to present and show what I have done so far to my instructors, I don't want it to seem like I haven't done anything. I have plenty of time before the big one though. Any more suggestion on how I can do it my best would defiantly help me work further. If I have any further questions when I begin my experiment where can I find you? Should I just re-post a new question or continue on with this one? Thank you so much you have helped me alot!

[donnahardy2]

Hi,

You've done plenty if you've selected a topic and have a good question that you can answer with an experiment. And if you have more time, you can do more background reading and look for incubators that you can borrow. Be sure a plan ahead so you will have time to do the complete experiment twice, and allow at least a week before deadline for data analysis and final write up.

One good thing to do at this point is to read the rules for the science fair that you will be entering, and make sure that your project will comply. Your instructors will also probably have special requirements that they will want to have included.

When you are ready to continue, just post a new question on this topic, and I'll automatically be notified.

Donna

[Nerdygirl]

I've decided to change my experiment a little bit. Instead of answering the question, "how does temperature affect bacteria transformation?" I'm answering "Do all bacteria cells transform during a transformation?" I will do this by transforming my bacteria as before and then calculate the transformation efficiency and compare it to the amount I spread onto the plates. Do you have any suggestions?

[donnahardy2]

Hi,

Your new question is a good question to answer, however, I'm not sure how you are going to measure your results. A science fair project should include a controlled experiment comparing results of independent variables. However, what type of experiment did you have in mind?


Donna Hardy