Long RT-PCR

Martin Offterdinger a8803349 at unet.univie.ac.at
Thu Nov 14 03:19:46 EST 1996


On Tue, 12 Nov 1996 22:23:31 GMT, bionbkx at panther.gsu.edu wrote:

>I am planning on using reverse transcription PCR to detect mRNA.  I am
>hoping to amplify the whole message, which is about 4 kbs.  So far
>most of the kits I have looked at don't promise an extension as long
>as the one I want.  If anyone has any experience with long RT-PCR,
>please let me know of the source of protocoles or kits that have
>worked for you.  Also, any optimization hints would be helpful.
>
>Thak you
>NK
>Hi
I have used two kits for the amplification of about 2kB
transcripts.The first one Perkin Elmer XL RNA PCR kit, I used for
amplification and cloning of a 2kB transcript-without any mismatch!
The second one is Boehringer-Mannheim titan one tube RT-PCR seems to
be more sensitive and is easier to use.(With Perkin elmer you have to
change the reaction buffer after RT by filling up to 100ul-with Boe-MA
you dont.) Both kits offer enhanced fidelity and have been used for
RNA amplicons up to 6kB(according to the manufacturerers)
Hope this helps
martin



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