Discussion in 'Supporting Biology' started by Jade, Nov 13, 2017.
is it a spread plate?
Always autoclave nutrient agar (unless this is a special type). Some agars can be sterilised by just boiling but nutrient agar has to be autoclaved or you run the risk of growing all sorts of bacteria/fungi alongside your true culture.
Well we've done it, I'll let you know the results. The e-coli growth over night was a little disappointing so we will see.
...so most of the plates appear to have formed bubbles in and under between the bottom of the plates and the agar). They werent there this morning and have only appeared after some time in the incubator. Any ideas?
From what I've seen through googling apparently agitating the agar too much before pouring can do this? Never seen it before...
Yes Jade the bubbles are an artefact of agitating the agar. It will make viewing the colonies a little more difficult however you should have no trouble seeing the transformants. Was it E. coli B that you used as the host for the plasmid containing the FGP? Look forward to seeing the pics.
No, we used a K-12 strain: E. coli strain HB101 K-12, lyophilized to be precise.
I have 2 more to do soon, hope they work well!
I use a microwave and have not encountered any further problems with my plates, all works very well, and the unused plates keep for ages in the fridge.
We use this kit all of the time - works like a dream
I decided to ask bio-rad, this is their response.
This practical is my absolute favourite so far in learning to be a bio and science tech. I followed all instructions to a T. For the first kit I used the microwave method and for the second kit I used the autoclave method. the microwave method gave bigger colonies. but all plasmid colonies glowed under UV light. I made sure to use transformation solution where possible and kept a constant read of the water bath at 42 degrees during class.
Well done you. I think that you deserve a round of applause for inspiring and engaging your Students.
We use the biorad kits all the time, and just microwave it no matter powder or prepared agar in a bottle. We've never had any problems by not using an autoclave.
I contacted them, they say microwaving is better than autoclaving as less nutrients are broken down and the bacteria/fungi are killed by the microwaves.
Well some of them are.
apparently it's current practice..
there is a paper here.
It's pretty easy to sterility test your media.
Sciencebuddies puts it quite nicely: LB agar is "a subtype of nutrient agar, this is the general medium for microbiology studies and may be used for routine cultivation of not particularly fastidious microorganisms. Also, does not preferentially grow one kind of bacteria over another." That means it should be fine to just put through autoclaving or however you normally make up agars.
Pointed out the kit several times but although people have said they will consider doing it, as yet noone has requested.
Separate names with a comma.