[BioC] RT-PCR normalization & stats reference
anthonyb at ichr.uwa.edu.au
Fri Mar 25 07:07:22 CET 2005
18S rRNA is a good starting point because it is stable, but the
disadvantage is that the expression level is very high.
Ideally, the housekeeping gene and target gene should be expressed at
a similar level, therefore you should target a stable mRNA rather
than 18S rRNA.
I followed the literature below to get a stable gene for human T-cells
Warrington JA, Nair A, Mahadevappa M, Tsyganskaya M.
Comparison of human adult and fetal expression and identification of
535 housekeeping/maintenance genes.
Physiol Genomics. 2000 Apr 27;2(3):143-7.
PMID: 11015593 [PubMed - indexed for MEDLINE]
Hamalainen HK, Tubman JC, Vikman S, Kyrola T, Ylikoski E, Warrington
JA, Lahesmaa R.
Identification and validation of endogenous reference genes for
expression profiling of T helper cell differentiation by quantitative
Anal Biochem. 2001 Dec 1;299(1):63-70.
PMID: 11726185 [PubMed - indexed for MEDLINE]
The following website also has everything you need to know about real
time PCR including house keeping gene section, validation of
microarray, software etc........
>Anyone have a good reference for how to normalize and determine
>significance of fold change for RT-PCR data?
>Basically, we have 6 control individuals and 6 experimentals. We
>normalize using 18s, but our RNA measurements going into the machine
>are pretty noisy, so we need to know what people who
>**successfully** confirm microarray data with RT-PCR are up to...
>Bioconductor mailing list
>Bioconductor at stat.math.ethz.ch
Anthony Bosco - PhD Student
Institute for Child Health Research
(Company Limited by Guarantee ACN 009 278 755)
Subiaco, Western Australia, 6008
Ph 61 8 9489 , Fax 61 8 9489 7700
email anthonyb at ichr.uwa.edu.au
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