Housekeeping gene: actin
adobrovic at medicine.adelaide.edu.au
Wed Nov 12 16:59:56 EST 1997
Whereas B-actin and GAPDH are useful controls for Northerns, they are close
to useless for RT-PCR because of the presence of numerous pseudogenes in
humans and presumably other species. Run a RT negative control with just
about any primers and you still will get a signal.
Controls that you could consider are housekeeping genes without pseudogenes
such as abl or B2-microglobulin though the presence of pseudogenes will
vary from species to species.
For human work see Kidd V, 1997 Debate round-table. Appropriate controls
for RT-PCR. Leukemia 11(6), 871-881 (1997)
> Z. Huang (huangz at hotmail.com) wrote: : Hi, bionetters,
> : Now I am looking for primers for house keeping genes such as beta
> actin or : GAPDH in rat and guinea pig. However, in case of guinea pig
> I can only find : a short fragment of cDNA encoding GAPDH from
> genebank. And it's even worse
> : that I can't find published sequence data for rat beta actin. My
> problem is: : I want to design a set of primer to amplify cDNA of
> above house-keeping gene : from rat, guinea pig. If it's possible this
> set of primers can also be used : on human and rabbit. Is there anyone
> who know such a "universal primers for : house keeping gene"? I would
> be very appreciate to any suggestions. Thanks.
> I cant find your original posting, so i'm not sure of your original
> question but hope this helps...
> I thought actin had been sequenced in all species, but there you go!
> If you want to use b-actin, any exonic b-actin sequence you can find
> in any species will work in your system because of the high degree of
> conservation: human and yeast beta actins are still 98% identical in
> coding sequences, such is the selective pressure....
> Generally actin is a good because it can be used as an "all-species"
> control but be aware that levels of non-muscle actins can vary in some
> conditions (eg low serum)...
> Personally i have been using 18S rRNA as a control, see previous
> recent discussion at this site...
Alexander Dobrovic, Ph.D.
Department of Haematology-Oncology
The Queen Elizabeth Hospital
Woodville, SA 5011, Australia
Affiliate Senior Lecturer
Department of Medicine
University of Adelaide
Tel 61-8-8222 6884
Fax 61-8-8222 6046
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