isolation of DNA from bracken

Timothy Bushnell bushnt at rpi.edu
Tue May 3 01:12:21 EST 1994


  AJW Hall writes:

>   I am isolating DNA from bracken (pterdium aquilinum) using a CTAB method...

    In our lab we have also used the CTAB method for isolating DNA from
plant tissue and seen similar problems in dissolving and restriction
digestions.  One possible problem that we noted was that the CTAB had not
been extracted from the solution before precipitation.  Typically we now do
2 or 3 chloroform:isoamyl alcohol (24:1) extractions before we precipitate.  

    Additionally, if you still can not get the DNA to dissolve, and it is
due to the CTAB, readjust the concentration of NaCl in the solution to at
least 0.7M and allow the tube to incubate at 60C for 30 to 60 minutes.  Then
reextract the solution with chloroform:isoamyl alcohol and reprecipitate.

    As for the digestions, what enzymes are you using?  Some may be
inhibited by the residual salt that is present in the sample, or if you have
done a phenol extraction, residual phenol, even in very low amounts, will
tend to inhibit reactions.  
    These in days when heavens mouring/ the days when earths foundation shook/
followed their mercenary calling and took their wages and are dead.
    Their shoulder held the sky suspended/ they stood and earths foundation
stayed/  What god abanded, these defended/  And saved the sum of things for pay



More information about the Methods mailing list