BIONET DNA Sequencing Survey

Andre Hamel hamel at ccu.umanitoba.ca
Mon Jun 21 12:54:51 EST 1993


In article <20JUN199309042751 at aardvark.ucs.uoknor.edu> broe at aardvark.ucs.uoknor.edu (Bruce Roe) writes:
>In article <C8vqIC.KF8 at mentor.cc.purdue.edu>, pmiguel at bilbo.bio.purdue.edu writes...
>>In article <19JUN199308260889 at aardvark.ucs.uoknor.edu>, broe at aardvark.ucs.uoknor.edu (Bruce Roe) writes:
>>>


>"I call this "fall-off" because it is so refered to in the Sequenase manual"
>
>although I believe Tabor and Richardson might have implied "fall off" in
>their PNAS paper.
>
>I'd also really like to know what's on the 3' end of those "fall off" 
>(as you put it) or "template associated compression" (as I put it) bands.
>If these bands truly are due to "fall off" (which is caused by the enzyme
>pausing at regions of template secondary structure), then the vast majority
>of the products would have a 3' deoxynucleotide.  If however, these "false
>terminations" are due to "mis-incorporation" as the enzyme trys to continue
>through regions of strong secondary structure, then it might be that 
>the observed "false terminations" are due to mis-incorporation of a
>dideoxynucleotide and the subsequent dissociation of the enzyme from 
>the template.
>
>	Anyone got any clues from experiments they've done or from the
>primary literature?

Yes, I suspect I've (in some way/shape or form) provided at least
some info in responce to above posed questions (perhaps some answers
too?). TdT tailing, when one has conditions right, usually works fine. I
recall there being several publications on this matter ... as far back as
late 1980's, as recent as few weeks ago ... we've used TdT tailing since
about 3 yrs now, when we came across a Nucl. Acids Res. article found in
an issue from around 1985 ... if any specific inquiries on this, I'll
simply do a Medline search then post those references ... hopefully
somebody reading this will have them handy and be so kind as to relay the
specifics.

regards, Andre
                            ********************
Andre Hamel                                 email: hamel at ccu.umanitoba.ca
Manitoba Veterinary Services                lab tel.: (204) 945-7630
Infectious & Genetic Disease                     FAX: (204) 945-8062     
Mol.Biol.Lab., Winnipeg, Manitoba, CANADA
                            ********************



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