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bacterial rna isolation


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#1 gorkem

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Posted 11 May 2016 - 05:09 AM

Hi,

 

i have some questions about rna isolation from gram negative bacteria. 

 

1- do i have to prepare the medium with depc threated water fro bacteria growth?

2- I am using thermo rneasy kit and i do all the santrifuges at room temp. instead of pelleting the culture. but I have no rna . shall i do the elution step at 4 C?

Does anyone who used thermo rneasy kit can explain the growing conditions and dilutions of the bactreial culture?

can i reuse my falcons for bacteria growing after autoclave?

 

thanx, have a nice day 

 

 



#2 merlav

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Posted 12 May 2016 - 12:44 PM

1. No, prepare as usual.

2. Do you mean Qiagen Rneasy? because from thermo I don't recall any kit with the name rneasy from thermo are pure link, trizol, magmax, ribopure. For Qiagen rneasy you have to pellet the cells, remove media and add the lysis buffer. Because you are using bacterias check if there is a pre treatment step. The elution is at room temperature. For any column base extraction you have to make sure you aren't using more cells than the protocol recall or you will clog the column and get 0 nucleic acids. 

3.  Check if the tubes are one use or multiple use.  If is the type of single use you CAN'T recycle. Tubes are cheaper than re-do an experiment because of a contamination. 


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