Transcription Questions

[Robert Schoenfeld]

Hi -
I've got some questions about transcription reactions.  After doing them
for several years, I now find myself having to calculate specific
activity.  Realizing that there's probably a range of values, what
percent incorporation is considered "good"?  What are typical levels of
specific activity?

After TCA preciptating 1 ul of my transcription, I cleaned up the
remainder via a G25 spin column.  After counting the eluate in a
scintillation counter, I found that the cpm/ul for the column was about
50% higher than that calculated from specific activity.  Are spin
columns "leaky" to dNTPs, rNTPs?

Thanks in advance,
Robert Schoenfeld

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