T4 RNA Ligase

Thomas J. Albert tjalbert at students.wisc.edu
Sun Feb 9 23:53:22 EST 1997


Does anyone have any experience with using T4 RNA ligase to ligate a
single stranded  DNA leader onto single stranded DNA? We are attempting
to make a cDNA library using poly T linked Dynal Beads, and wish to
place a spice leader on the 3' (comp strand) end for easy
reamplification by PCR. Although this reaction sounds inefficient, T4
RNA ligase is supposed to have this ability and we have seen references
where it is used for just this purpose. After several attempts, however,
PCR seems to indicate that the leader is not present, while internal
primers to known genes produce nice products. Any help on optimization
of this rection would be appreciated.

Tom



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