11 replies to this topic

[SF_HK]

I'm preparing to make competent cells. Is there a reason why some people pefer to fliter sterilise CaCl2 over autoclave? Can CaCl2 be autoclaved?

Thank you

[Inbox]

I do filter sterilization.
You might want to look @
http://www.protocol-...osts/18679.html

[SF_HK]

Thank you. Also, to make 0.1M CaCl2, is it necessary to make the dilution from 1m CaCl2 or can I directly make 0.1m CaCl2? I read somewhere that CaCle concentartions must be prepafed from 1M stock solution.

I just simply prepared 0.1M CaCl2 by dissolving 3.67g CaCl2 (dihydrate, FW 147.02) in 250ml water.

[Inbox]

I just simply prepared 0.1M CaCl2 by dissolving 3.67g CaCl2 (dihydrate, FW 147.02) in 250ml water.

Thats fine.

But how you will make last 0.1M Cacl2 with 15% Glycerol for storing @ - 80^oC?

Its quiety handy to make 1M cacl2 , filter it and store @ 4^oC. For making 0.1 M Cacl2 just dilute it 10 times by adding to autoclaved distill water.

[badguy]

i did both types of sterilization and both also works good.
but usually i did filter sterilization because it saves time

[SF_HK]

to make 50ml 0.1M Cacl2 with 15% glycerol, I would add 42.5 ml (0.1M Cacl2), 7.5ml 100% glycerol. This is OK?

[Inbox]

to make 50ml 0.1M Cacl2 with 15% glycerol, I would add 42.5 ml (0.1M Cacl2), 7.5ml 100% glycerol. This is OK?

Not ok.

If you add any solution to 0.1 M it will decrease its concentration. You can consider adding cacl2 to 7.5ml 100% glycerol to make it 0.1M cacl2 with 100% glycerol and add 42.5 ml (0.1M Cacl2). But filtering 100% glycerol then for sterilization will be difficult task for u.

Better make 1M cacl2. then dilute accordingly.

[SF_HK]

Ok,

Lets say I make 1M Cacl2 (filter sterilise).

to make 200ml 0.1M Cacl2 (+15 glycerol)
- 20ml 1M CaCl2 (filter sterilised)
-30ml autoclaved 100% glycerol
-170ml autoclaved water

[Inbox]

Right.

[SF_HK]

Thank you. I'm just horrible at mathematics.

[mdfenko]

Ok,

Lets say I make 1M Cacl2 (filter sterilise).

to make 200ml 0.1M Cacl2 (+15 glycerol)
- 20ml 1M CaCl2 (filter sterilised)
-30ml autoclaved 100% glycerol
-170ml autoclaved water

150ml water

[SF_HK]

Thank you. Yes I realised the calculation error when I was preparing the solutions.