how to avoid poly(A)tails in a cDNA library

Michael L. Sullivan mlsulliv at facstaff.wisc.edu
Fri May 5 13:11:47 EST 2000


Clontech's cDNA synthesis uses an oligo(dT) primer isn't simple T's, it's
actually (dT)16(A,G, or C)N.  Thus, the primer will (mostly) anneal to the
5' end of the poly(A) tail and the clones will only have a stretch of 16
A/T at the 3' end.  If you didn't want to use that cDNA synthesis kit, you
could probably make that primer yourself and use it with another kit.  And
yes, I do believe several vendors kits for making random primed cDNA.

Mike



>Dear netters,
>
>Could anyone please provide some information for avoiding the long
>tails of poly(A/)or(T) in cDNA clones of a cDNA library? We are
>constructing cDNA libraries with Stratage cDNA synthesis kit for
>EST sequencing. With these cDNA libraries we will do the normalization
>and/or subtraction. Somebody argues that the possible long tails
>of poly(dA/dT)(>100bp) within cDNA clones might affect the efficiency
>of subtraction. Are there any ways to avoid the formation of this kind
>of long poly (dA/T) tails with the oligo(dT)-primed synthesis appproach?
>And are there available commercial kits for random-primed cDNA synthesis?
>
>Many thanks!
>
>Deshui Zhang
>Texas Tech University
>Lubbock, TX 79409-2122
>
>---


Michael L. Sullivan, Ph.D

U.S. Dairy Forage Research Center
1925 Linden Drive West
Madison WI, 53706

(608) 264-5144 Phone
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