Hi,
The taqman I ran has very high CT's. Even RPL19 is very high. I checked the RNA concs again and everything seems fine. What else could be causing these high CT's? I am clueless. Thank you for any help
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#2
cmgibson
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Posted 12 August 2004 - 03:40 AM
How many different primer sets have you used? Maybe you're reading on the wrong wavelength. We were having the same problem, granted it was DNA, and found out the fluorescence was off. Hope that helps.
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jadefalcon
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Posted 12 August 2004 - 04:14 AM
hi!
when our RT lost it's activity, i got high CTs, too. i found out that they correllated to not DNaseI digested DNA copys in the RNA prep. There always were 40-100 copys of DNA left in the sample, ever after digestion. They gave me positive signals in taqman assays at >38 cycles...
so maybe your sample doesn't hold rna or the RT reaction won't work, for whatever reasons...
mike
when our RT lost it's activity, i got high CTs, too. i found out that they correllated to not DNaseI digested DNA copys in the RNA prep. There always were 40-100 copys of DNA left in the sample, ever after digestion. They gave me positive signals in taqman assays at >38 cycles...
so maybe your sample doesn't hold rna or the RT reaction won't work, for whatever reasons...
mike
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