RNA Extraction from Neuroprecursor Cells Using the Bio-Rad Total RNA Kit

Biology

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Su, J. S., Monuki, E. S. RNA Extraction from Neuroprecursor Cells Using the Bio-Rad Total RNA Kit. J. Vis. Exp. (9), e405, doi:10.3791/405 (2007).

Abstract

Disclosures

The authors have nothing to disclose.

Comments

5 Comments

  1. I see that RNA yield assess by measure absorbance in ²60nm, I want to ask, can we assess DNA double strand from PCR with the same manner with absorbance measurementin ²60nm? because if we can do it, Ithink its important to decide continue to next step to running electroforesis. If we know, our PCR is not success to amplificate target DNA,we should not to continue electroforesis. I hope I get your answer, thanks.   Afiat 

    Reply
    Posted by: Anonymous
    July 6, 2008 - 10:50 AM
  2. Afiat

    Yes, however, spectrometry of PCR products is difficult as you must blank the spec first with your PCR mix including primers, taq, buffer, salt (waste of reagents!) and you will no know if your products are the correct size, specificity, etc... just run a gel, it's cheap and only takes a hour.

    Reply
    Posted by: Anonymous
    July 14, 2009 - 12:46 AM
  3. Measuring PCR product by using the spectrophotometer is not impossible; however, as Sarah pointed out you need to blank for factors that might interfere with OD ²60/²80 reading. Besides, since you will have lots of amplified DNA, you may have to measure the spec several times until you get OD ²60 ranging within 0.1 to 1 of values. Outside of the range (0.1-1 of OD ²60), you can't get accurate measurement due to the optical property of the spectrophotometer itself.
    However, it is easy to estimate the quantity of PCR products by using the mass ladders instead. When you load your samples on the gel to verify the quality of PCR products, simply load known amount of the mass ladder and compare the band intensity to find out rough range of quantities of PCR products.

    Reply
    Posted by: Anonymous
    October 6, 2009 - 6:23 PM
  4. I see that RNA yield assess by measure absorbance in ²60nm, I want to ask, can we assess DNA double strand from PCR with the same manner with absorbance measurementin ²60nm? because if we can do it, Ithink its important to decide continue to next step to running electroforesis. If we know, our PCR is not success to amplificate target DNA,we should not to continue electroforesis. I hope I get your answer, thanks.   Afiat 

    Reply
    Posted by: Anonymous
    July 6, 2008 - 10:51 AM
  5. Thanks for the great movie clip! I clearly see low protein contaminants in the eluted RNA solutions, however, the Spectrophotometer can't explain the integrity (or degradation) of RNAs that are easily monitored by the ratio of ²8S and 18S rRNAs. How is the ratio of 18s and ²8s rRNA after using the Bio-Rad kit? Thanks!

    Reply
    Posted by: Anonymous
    October 6, 2009 - 6:09 PM

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