anthocyanins and protein isolation

William.Cress Cress at botany.unp.ac.za
Tue Aug 30 02:39:54 EST 1994


I am cirrently working on isolating total protein from callus cultures of a 
anthocyanin pigmented (cyanidin-3-glucoside) cell line as well as a non-
pigmented line from the same plant species. My extraction buffer has 
protease inhibitors (PMSF, EDTA)included in it and contains 3,5 microlitres 
per 10ml mercaptoehtanol. After extraction I perform a Biorad assay to 
quantify protein isolated. I generally observe that yields are a lot less 
for the pigmented line. I then run 1D and 2D PAGE gelsusing apperently equal 
quantities  of protein for both lines. Bands obtainrd with red callus for 1D 
gels are always fainter or a smear as compared to the white . For the 2D 
gels only 1 or 2 spots are obtained with the anthocyanin -rich callus 
meanwhile the white yields plenty of spots as expected. I have tried to 
get rid of the pigment by running SEPHADEX G25 columns nad I concentrate my 
proteins by daialysing against sucrose . I have also tried acetone 
precipitation but I still obtain the same results. 

I would like to know if anyone has had the same or similar problem nad how 
did they overcome it.

N.P. MAKUNGA
DEPT .OF BOTANY
UNIVERSITY OF NATAL
PRIVATE BAG X 01
SCOTTSVILLE
3209
SOUTH AFRICA

E-MAIL: Makunga at AGRIC_PMBUNP.AC.ZA
 



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