Making long transcripts in vitro

[Bruce Phillips]

	There has recently been some discussion of how to synthesize in vitro
long cDNAs with RT/PCR.  I wonder if anyone has prepared very long RNAs
using plasmids with SP6 or T7 promoter-driven plasmids.  I am faced with
synthesizing in vitro a 8 kb RNA from a SP6 promoter.  I have succeeded but
the long transcript is only <5% of the yield.  The average size transcript is
about 5- 5.5 kb.  Any ideas for enhancing the yield of larger size molecules
would be greatly appreciated.  Incubation time is not the solution so I'm
interested in cocktail conditions or pretreatments of the plasmid.

	Thanks.

                     Bruce Phillips -  bap at med.pitt.edu