Kate06
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Bacteria lawn from single colony?

Postby Kate06 » Sun Jan 03, 2021 10:28 pm

Hello, I'm a seventh grader doing a project on the antibacterial properties of natural Aloe Vera and how it would compare to hand sanitizer and antibacterial soap. I have successfully grown some individual colonies of bacteria from swapping door handle, phone, etc. and would now like to use an isolated colony to create a bacterial lawn on a separate Petri dish so that I can test zones of inhibition. I have read there are two methods to do this, 1. broth and swap or 2. colony and wire loop. The broth and swap method looks like the easier of the two and a more successful way to achieve an even lawn. Is there an easy way to to do this successfully with the isolated colonies on my current streak plates? Or should I stick to wire loop method. Thank you for your advice.

lmp1341
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Re: Bacteria lawn from single colony?

Postby lmp1341 » Mon Jan 04, 2021 9:47 am

Hello and Welcome to Science Buddies!!
Bacteria absolutely RULES and I love your science fair project idea!!

So I can help you a little better, do you have access to a lab at school where you have materials to choose from for this experiment or are you doing this from home? The reason I am asking is because both methods involve sterilization of your materials, and you need a butane burner for this. If you're doing this project at home, no worries, we can improvise :D

Broth and Swap
The broth and swap method works if you have LB Broth Media available. Do you have broth available to use? To do this, you would have to use a sterile toothpick and pick up one or two colonies (dots) from your starter plate, then swirl it in the broth. Once you've done this, you'll want to leave the broth in an incubator for a little while to allow the bacteria to replicate. Then, you will use a sterile pipette or micropipette to place a drop of the broth onto a new Agar dish, and swirl it around. If you have sterile hockey sticks to use, you can lawn and spread it around that way. You are right, I think this method would produce a more even lawn than the wire loop method, especially if you're looking into zones of inhibition. I am going to link a few articles here about procedures for this. One will be with lab equipment, and one is an at home version in case you don't have lab materials readily available.

Here's what I'd advise:
1. Use a sterile toothpick to pick up 1-3 colonies (depending on size) and swirl into the broth culture.
2. Place a cap on your broth culture and gently invert to spread the colonies throughout the liquid media.
3. Place your broth culture into an incubator at 37C
4. Leave the culture there for 24h before proceeding to lawning, and make sure you clean your work station with 70% Ethanol to prevent contamination or hazard. You should handle bacteria with a parent or teacher around to ensure this.
5. Prepare your next step by labeling a new agar dish with the contents and date.
6. After 24h has passed, remove the broth from the incubator and use a STERILE cotton wool swab to dip into your solution once, and spread this evenly over the brand new agar dish.
7. Incubate the dish at 37 C for 24 h and check for growth!

https://practicalbiology.org/standard-t ... lawn-plate
https://www.madaboutscience.com.au/shop ... gar-plates

The first link has helpful protocol info and the second is a more at-home friendly version.

Let me know if you have any other questions as we are happy to help!!

Best of luck with your project and happy streaking!!!

Stay nerdy,
lmp1341

Kate06
Posts: 2
Joined: Sun Jan 03, 2021 10:05 pm
Occupation: Student

Re: Bacteria lawn from single colony?

Postby Kate06 » Mon Jan 04, 2021 12:31 pm

Hello lmp1341,
Thank you for replying back and providing some helpful suggestions. I am doing this project from home, without the use of a lab, but I do have a Bunsen burner and inoculating loops. Is there a recipe for LB broth that is easy to prepare at home? If not, would this LB broth from Carolina Supply be what I need?
https://www.carolina.com/biotechnology- ... /216655.pr

I also do not have access to an incubator. Would that make the broth technique impossible? Or could I improvise with a homemade incubator that would keep the nutrient broth at the correct temperature?

Thank you again,
Kate

lmp1341
Student Expert
Posts: 67
Joined: Sat Aug 31, 2019 1:54 pm
Occupation: Student

Re: Bacteria lawn from single colony?

Postby lmp1341 » Tue Jan 05, 2021 8:51 am

Hi Again!
That's great to hear. If you want an even lawn as opposed to a Z-streak, then I would go with a sterile q-tip if possible.
If you don't have access to an incubator, it's okay! You can make your own:
https://sciencing.com/ways-grow-bacteri ... 13461.html
You can use an aquarium or other box with a lamp on top to maintain an optimal temperature. Otherwise, you can use the same method that you used for your plates to grow. Just note that your broth should be a little cloudy by the time you're ready to use it for streaking the next plate.

For the broth, you'd need access to an autoclave for sterilization, so I would recommend buying the broth pre-made. Here's some ready to use broth I found online, https://www.thermofisher.com/order/cata ... #/10855001
but the link you sent looks like it would work just as well!

Any other questions? It sounds like you're going to do great!!!
I'm excited for you!

Let us know if we can help any further!

Stay nerdy,
lmp1341


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