[PStepper]

Hi everyone,

this is the main discussion space for BOMB protocol #6.5 TNA extraction from yeast.
Here you can post all your questions and feedback!

[PStepper]

reserved space

[Iason K.]

Hi ,

I have been using the bomb bio 6.5 protocol for yeast and works like a charm. It provide us with  long DNA fragments that we require but I want mainly the gDNA. Can I use RNase A and at which step would you recommend and at what concentration?

[admin]

Ahoy,

I am stocked to hear that you like our protocol!
I presume that you have thought through whether RNA contamination actually poses an issue.
Because unless you are doing super sensitive quantification, transfections or steps involving reverse transcriptase, I never really encounter any issues whatsoever with residual RNA. But, I am mostly  running PCRs or similarly robust techniques.

I am not sure whether RNAse A has a tolerance for Lyticase and GITC but I would not risk it.

Unfortunately the save option would be to finish the protocol transfer your elution to a tube loaded with RNAse A + Buffer –> Incubate  (+ maybe deactivate) and then perform something like a PEG (SPRI \ Ampure) purification or if you want to go DIY all the way BOMB protocol 4.1., obviously you can also ethanol precipitate etc at this point.

Cheers

Tim M

 

[Author]

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