Science Buddies: "Ask an Expert"

Hi Arnlan,

I'm wondering how you described your experiment with her. The tube without plasmid is your negative control, i.e. no DNA, but you ALSO have a condition with 1X plasmid, remember. That will act as a "control" condition while your 10X plasmid is your experimental condition. You definitely have all of the conditions you need to test whether increased plasmid affects transformation efficiency. Have you discussed this fully with her or did you just answer a question about the control with your negative control? I understand controls can get confusing because we use the word to mean multiple things sometimes. In this case, I believe she wants to hear about your control condition, or the 1X plasmid, as I said above.

Of course, you also could decide to do a different experiment testing the effects of pH, salt concentration, temperature, etc., but that is a separate experiment.

I hope this helps,
JMP

Yeah thanks, i appreciate it. I will confer with her about it.

Yeah, that was the constant she was looking for.
I will be doing my project in less than 2 weeks.

-Thanks

I'm still having trouble with the problem and purpose.

Hi Arnlan,

Can you please clarify exactly what you are having a problem with? Last time you wrote a problem and a purpose and I gave you some suggestions to change them slightly. Have you come up with a new draft? What exactly are you having trouble with?

JMP

Hey,

The problem i'm having is trying to tie my purpose and problem to the project.

-Arnlan

Hi Arnlan,

Previously, you suggested writing your problem as follows:

Will it be fine if i put this as my problem ? : It can be very expensive to chemically synthesize very short peptides, never mind complex polypeptides and whole proteins which may have post-translational modifications. As the biology of bacteria becomes clearer, coupled with the abundance of bacterial species and strains available and the exciting advances made in molecular biology research and biotechnology, the possibilities and applicability of transformation becomes phenomenal.

and I said that you should tie it to your actual experiment more. What I mean is this that while it is a problem that it can be expensive to synthesize peptides and proteins, you haven't told me how YOUR experiment helps with that problem. In what way does testing the effect of plasmid concentration on bacterial transformation efficiency relate to the problem you have written? Your problem is to make synthesizing of proteins cheaper by doing it in the MOST efficient and cost-effective way, so it is important to know how the amount of plasmid effects transformation efficiency. Does that make sense?

As for your purpose, I think the purpose should be a very simple and direct statement of what you are planning to test. I would look at the Objective statement from this project as a good example. https://www.sciencebuddies.org/science- ... p013.shtml

I hope this helps,
JMP

Yeah! That makes alot of sense. So for my purpose will it be fine if i put this"

The purpose of bacterial transformation is to introduce a foreign plasmid into a bacteria and use that bacteria to amplify the plasmid in order to make large quantities of it. This is based on the natural function of a plasmid : to transfer genetic information vital to the survival of the bacteria.

Thanks for your help!!!
-Arnlan

Will it be fine if i put this for my purpose?

The main purpose of this project is to synthesize proteins in a cheaper way, by doing it in the most efficient and cost-effective way, so it is important to know how the amount of plasmid effects transformation efficiency. Bacteria are transformed for numerous different reasons. Some of these reasons may include expression of medically useful recombinant proteins such as insulin for treating a disease or vaccines for prevention of disease. Other reasons could be expression of proteins that confer on bacteria the ability to survive in particular environments such as to "clean up" contaminated environments in bioremediation.

It's looking much better, arnlan, but I'd consider reversing the order of your paragraph slightly. Start with the end of your paragraph

Bacteria are transformed for numerous different reasons. Some of these reasons may include expression of medically useful recombinant proteins such as insulin for treating a disease or vaccines for prevention of disease. Other reasons could be expression of proteins that confer on bacteria the ability to survive in particular environments such as to "clean up" contaminated environments in bioremediation.

and then continue with the specifics of your experiment

The main purpose of this project is to synthesize proteins in a cheaper way, by doing it in the most efficient and cost-effective way, so it is important to know how the amount of plasmid effects transformation efficiency.

You generally want to move from the general to the specific, and this order will do that much better.

Alright, Thanks

I have 1 last question. Currently my title is bacterial transformation efficiency. But i think i should change it to pertain more to my experiment. So far this is what i have

How different concentrations of pGLO plasmid effect the growth of e coli plated on LB/ARA/AMP plates.

I need help on shortening it.

I think you have a very good start on the title. If you are looking to shorten it, I would take out some of the details which are a bit too specific for a title. For example, you can just say "effect of growth of E. coli" without mentioning the kind of plates you are using. Those kinds of details belong in your full report, but not the title. The same can be said for the name of the plasmid (i.e. pGLO).

The only other point I would make about your title, is that you don't strictly care about growth. You are counting growth of E. coli because in this case it is a measure of transformation efficiency, so you may want to include that in your title.

Will it be fine if i just put it as Effect of pGLO plasmid on the growth of E Coli?

Will it be fine if i just put it as Effect of pGLO plasmid on the growth of E Coli?

Hello,

My final title i will use is

-I have removed it due to privacy, -thanks


How does that sound?

I think that sounds very good arnlan. Way to go!

Alright cool,

If i have any more problems, i'll make sure to let you know.

Hello,

I hope you are doing fine.

I actually won 1st place at the district level of the science fair. And Regionals science fair is later this week. I'll let you know how I do.

I'm also starting to think of how I could improve this experiment for next year. I've just been thinking and coming up with projects that I could do. This is what I have so far : Determining the effect of bile salt on E Coli bacteria along with heat shocking. << Does this lead anywhere? Could I get any results from that? If so, I'll try to test it on some cancer cells in the institution that i'm working with.

I'm just thinking and I just came up with that project. Could that work? Can bile salt kill bacteria? Or do I have to test to see? I will start by doing a home investigation to test it on E Coli, if it does work, then I will take it to the lab to test on cancer cells. I could also transform the bile into the bacteria and see how it effects it. This project could be a transformation efficiency project. The title could be- Transformation Efficiency : Determining the effect of Transforming bile into a cancer bacteria cell along with the effect with heat shocking.

I could also add bile into the agar that the bacteria is grown in, and I could test the effects of it.

Does this lead anywhere?

Hi arnlan,

I'm glad to hear that your science fair has been going well, but I have a few concerns about your new project.
1) I'm not entirely sure what you mean by "transform bile into the bacteria." If you want to see how the presence of bile salts affects bacterial transformation efficiency, then I think you can absolutely test this. Using the same kit you have now, you can try the transformation reaction with and without the presence of bile salts. However, when you say "transform bile into the bacteria" it makes me think you don't exactly understand what transformation is. By definition, transformation is the process of introducing foreign DNA (new DNA) into bacteria, and since bile salts aren't DNA, what you are suggesting would not be transformation. You could certainly just test the affect of bile salts on bacteria (i.e. do the bile salts kill the bacteria as you suggested), but this would not be a transformation reaction. Do you understand?

2) I'm not sure how the project you are proposing relates to cancer cells. Bacteria are not cancer cells. Cancer is caused by mutations in the cells in your body, so you would need mammalian cells to test the effect of bile salts on cancer. It is possible to introduce new DNA into mammalian cells (called transfection), but this would not be readily possible at home. Growing mammalian cells and transfecting them really requires a lab environment. If you have access to that, you could consider a project like that, but again, you can't transfect cancer cells with bile salts because bile salts are not DNA. Again though, you could certainly propose a project looking at the effect of bile salts on cancer cell growth or survival, etc.

If you are interested in the effect of bile on bacteria, then I think your best bet may be to test the effect of it on bacterial growth. You even suggested a method to suggest that in your last post:

I could also add bile into the agar that the bacteria is grown in

, or as I said above, you could test how different bile concentrations affect transformation efficiency.

I hope this helps!
JMP

From what you know, could this project be successful? I would also like to know where I could purchase the mammalian cells.

2 years ago, my sister did a project on bile salt as a stain remover, and it worked better than detergent. So I guess I'll find out if it can kill bacteria.

And yes, I understand the process of transformation.

-The results for the regionals science fair will come out tonight, I will post on here how I do.

I'm not sure whether the project can be successful, because I don't know exactly what you are proposing. Certainly I don't see why you couldn't test the effect of bile salts on bacterial growth or transformation very readily.

As for where you can purchase mammalian cells, as I said, you'll need access to a university level lab to be able to grow mammalian cells. If you find a lab that is willing to help you with your experiment, they'll probably have at least some kind of cells you can use. Otherwise, we often buy them from the ATCC (atcc.org), but I don't believe they'll sell to an individual. If you do have to buy your own mammalian cells, be aware that they are quite expensive (usually several hundred dollars), so your best bet is definitely to find a university lab that will let you use some.

JMP

I did well at the regionals science fair, and I will be advancing to the State Science Fair.

JMP wrote:I'm not sure whether the project can be successful, because I don't know exactly what you are proposing. Certainly I don't see why you couldn't test the effect of bile salts on bacterial growth or transformation very readily.

What i'm trying to do is use bovine bile and see it's effects on bacteria, and even cancer cells. As I said before, I might have different types of plates, and one plate has bovine bile. I might consider adding other substances to see which one effectively controls the cancer cell. I'm not sure if that makes sense.

I just watched a video, and learned that bile disinfects your food.. So could that be applied to bacteria?

JMP wrote: As for where you can purchase mammalian cells, as I said, you'll need access to a university level lab to be able to grow mammalian cells. If you find a lab that is willing to help you with your experiment, they'll probably have at least some kind of cells you can use. Otherwise, we often buy them from the ATCC (atcc.org), but I don't believe they'll sell to an individual. If you do have to buy your own mammalian cells, be aware that they are quite expensive (usually several hundred dollars), so your best bet is definitely to find a university lab that will let you use some.

JMP

I will be working in a university lab, the same one that I did the bacterial transformation efficiency project. I'll ask her if they have those cells, if not, ill be willing to purchase it.


I would also like to incorporate the pGLO Plasmid into it. Could that effect anything?

I'm glad that you've been doing well in your science fairs. As for your next project, the problem you have right now is that you really need to narrow down your topic. So far, you've proposed to look at the effect of bile on cancer cells, bacterial growth, and bacterial transformation efficiency. Those are all fine things to test, but they are three separate projects, not one. Think about what you are really interested in, and pick one of them to follow up with, and then we can talk about the details. I think all three of these projects are doable and potentially interesting, but you really do need to narrow down your area of interest and just pick one so that you can have a coherent hypothesis. Remember, a good experiment tests only ONE variable. It should answer one specific question.

JMP

I would like to work on the effect of bovine bile on cancer cells.

Thanks for clarifying.

Hello,

I have a few questions about this years project.

My purpose in this experiment was : Bacteria are transformed for numerous different reasons. Some of these reasons may include expression of medically useful recombinant proteins such as insulin for treating a disease or vaccines for prevention of disease. Other reasons could be expression of proteins that confer on bacteria the ability to survive in particular environments such as to "clean up" contaminated environments in bioremediation. The main purpose of this project is to synthesize proteins in a cheaper way, by doing it in the most efficient and cost-effective way, so it is important to know how the amount of plasmid effects transformation efficiency.

And my problem in this experiment was : It can be very expensive to chemically synthesize very short peptides, never mind complex polypeptides and whole proteins which may have post-translational modifications. As the biology of bacteria becomes clearer, coupled with the abundance of bacterial species and strains available and the exciting advances made in molecular biology research and biotechnology, the possibilities and applicability of transformation become phenomenal.

My question is, when I present to the judge and they ask me how I tackled the problem what do I say? My results are as follows : On the 1x plate I had 3 glowing cells and the 10x plate had 0 glowing cells. The 1x had a transformation efficiency was 18.75 and the 10x plate had a transformation efficiency of 0. So due to my results how did I fix the problem? And how does it tie to my purpose? How did I synthesize proteins in a cheaper way?

I'm sorry this is late.

-Arnlan

Hi Arnlan,

I think that the question you proposed is too broad based on the results of your experiment. You have a great big picture idea, but on a smaller scale what was your question that guided your experiment? Then I think it will be much easier to tackle the question from the judges. I am happy you had successful bacterial transformations, but were you able to test if any protein was being produced?

Michelle

masallin wrote:Hi Arnlan,

You have a great big picture idea, but on a smaller scale what was your question that guided your experiment?


Michelle

The only thing that I had in mind when doing the experiment was getting the greatest transformation efficiency as my results. On my purpose it says that to synthesize proteins in a cheaper way so it is important to know how the plasmid concentration effects transformation efficiency. So due to my results on having a better transformation efficiency with a smaller concentration can I conclude that using a small concentration of plasmid is cheaper and moreover gives a better results and a better transformation efficiency?

The judge actually confused me and told me that she works in the lab with a high concentration of plasmid and it doesn't effect transformation efficiency. So does that mean that I did my project incorrectly??? I did this experiment with a microbiologist in a near institution, and she didn't see any problem in my results. She actually said and concluded that maybe the more plasmid you insert into a cell, it overwhelms the cell or even stops the growth.

I'm just gathering my thoughts before I head to the state science fair, because I know I have to answer with a good justified answer that shows the judge that I know what I'm saying.

masallin wrote: were you able to test if any protein was being produced?

I know that It's not the plasmid that glows but the gene it encodes. I saw a glowing colony on the 1x plate after I shined it with a uv pen light, so does that test if any protein was produced? Becuase I had a control group which showed that when you had arabinose in the agar, then the protein would be produced in the bacteria.


Thanks for your quick response.
I really appreciate it. I have about 3 weeks left until the state science fair.

THANKS!!!!!