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why my solution II turns cloudy during plasmid extraction?


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6 replies to this topic

#1 Curtis

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Posted 04 February 2012 - 10:11 AM

I use conventional Sambrook's method for plasmid extraction, but recently when I make solution II with SDS and NaOH the solution turns cloudy. I also noticed the my DNA pellet is getting smaller. has anybody ever have the same issue ?

#2 scolix

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Posted 04 February 2012 - 10:45 AM

If SDS has precipitated, the solution is not clear. Warm it a bit and it should be clear.

#3 bob1

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Posted 04 February 2012 - 01:09 PM

To add to Scolix's comment, the reason you pellet is smaller when you get a precipitate of the SDS, is that the cells are not lysed as efficiently as you would with the SDS in solution fully.

You should either make the solution II fresh each time (best), or store it with the lid tightly closed to prevent the NaOH from absorbing CO2 from the atmosphere and altering the pH.

#4 Curtis

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Posted 06 February 2012 - 10:38 AM

yeah, I prepared fresh and it worked fine this time. It must have been NaOH. though I have heard SDS solution also expires after a while. I have also noticed when I put solution 2 on ice it freezes fast.

#5 phage434

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Posted 06 February 2012 - 11:06 AM

I declare today to be "Throw away your NaOH solution day." In honor, I'm remaking all of my NaOH solutions, and I recommend everyone else do the same. Over time, NaOH solutions absorb CO2 from the air and form sodium bicarbonate. If you are like me, the NaOH bottle on your bench has been there too long. The drains can use a cleaning anyway.

#6 bob1

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Posted 06 February 2012 - 12:58 PM

yeah, I prepared fresh and it worked fine this time. It must have been NaOH. though I have heard SDS solution also expires after a while. I have also noticed when I put solution 2 on ice it freezes fast.

The freezing is the SDS freezing, it will also do it if you cool it below about 15 deg C.

#7 Curtis

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Posted 08 February 2012 - 08:31 AM

it was from 2009. thanks guys




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