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A question for microRNA TLDA


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#1 yueh76

yueh76

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Posted 30 September 2010 - 07:23 PM

I used ABI microRNA TLDA to research my study, but no biological replicate. I transfected a mock plasmid and a plasmid expressing specific gene in Huh7 cells. MicroRNA-122 is known to highly express in Huh7 cells. In my results, the CT values (miR-122) are about 32 in mock and experiment, but the CT value seems to be high. Now, I focused on miR-XXX, and the CT value is about 33 in experiment and CT value in mock is about 37. It is reported that the expression levels of miR-XXX in Huh7 is 5-fold low compared with miR-122 in Huh7. ABI told me that "For TaqMan Low Density Arrays, precision falls off at Ct values >30. On TaqMan Low Density Arrays, a Ct of >30 reflects a very low copy number of the target molecule (less than 30) per well; Ct=30 is equivalent to Ct ~33 on plate. Ct=33 on TaqMan Low Density Arrays is approaching the limit of detection because it is equivalent to one copy of the target
molecule." ABI also told me that CT value between 33 to 40 in TLDA means one copy number of target microRNA.
My question is that whether delta-delta-CT value >1 is significant (If CT value between 37 to 40 in mock, and CT value between 32 to 33 in experiment) because of the similar results are not less in my study.
I used Trizon to isolate total RNA for 60 mm dish.
Thank you for your help.




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