internal standard for semi-quantitative pcr

Tim Spahlinger txs at po.cwru.edu
Tue Dec 12 11:53:19 EST 2000


Although I haven't used it myself, I have seen GAPDH 
mentioned in other newsgroups for Real-time PCR as a 
possible internal standard.  However, this was not mentioned 
specifically for work on material obtained from HIV+ 
individuals.

a.boasso wrote:

> We are using a semi-quantitative pcr assay to evaluate the expression level
> of genes involved in the immune system functions (like citokynes, receptors,
> co-stimulation molecules), in hiv+ individuals or aids patients.
> In this assay we perform a pcr reaction with two sets of primers:
> 1. a pair of primers for the target mRNA amplification (citokynes, receptors
> or co-stimulation molecules)
> 2. a pair of prime for the beta-actin mRNA, which will be the internal
> standard
> 
> The beta-actin mRNA concentration is determined in a previous step, with a
> specific competitive pcr assay, with a synthetic competitor, used in
> different reactions at different concentrations.
> 
> Some critics have been moved against the use of beta-actin as an internal
> standard, as its expression seems to change widely in pathologic cells.
> 
> Is anybody able to give me some hints or information about the use of some
> other internal standard genes in cells from hiv infected patients?
> 
> Thank's






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