cDNA Synthesis Problem

hezma · #1 (**permalink**) 07-23-2008, 11:53 AM

Hello i am having problems synthesizing full length cDNAs. what could be the problem? I am extracting RNA from cells then using Reverse transcriptase to get the RNA to cDNA however when i clone the cDNA into plasmids, the ones i am looking for are not there or short.

plz help

danfive · #2 (**permalink**) 07-23-2008, 07:49 PM

Improve the quality of your RNA and the problem should go away.
I use Trizol, but a good kit can give you even better results. Also try to use fresh RNA since it will degrade, even if frozen.

vimlendusinha · #3 (**permalink**) 07-29-2008, 12:41 PM

just use GTC or any kit for rna isolation and do the RT. check ur cdna synthesis protocol u have to denature rna with oligo dt, cool it and then add dntp, buffer and enzyme and keep at 42 for 1hr u wil get it. The quality of rna should be good

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cDNA Synthesis Problem

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