CHS | Chalcone synthase

application example

 0.5 to 7 µg of protein from Arabidopsis thaliana Col0 leaf tissue, extracted with Agrisera PEB protein extraction buffer 1X were separated on 12 % SDS-PAGE using tank (BioRad system) to transfer to to nitrocellulose membrane during 1 hour. Blots were blocked with BSA (Sigma) for 1h at room temperature (RT) with agitation. Blot was incubated in the primary antibody at a dilution of 1: 1 000 for 1h at RT with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, then washed once for 15 min and 3 times for 5 min in TBS-T at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated, from Agrisera AS09 602) diluted to 1:20 000 in for 1h at RT with agitation. The blot was washed as above and developed for 5 min with ECL according to the manufacturer's instructions. Exposure time was 20 seconds. For transference control the membrane was stained with Ponceau red and integrity of proteins was evaluated using 12% SDS-PAGE silver stained.