NtMMP1 expression in wild type BY-2 suspension cells. A: Northern blot analysis of the endogenous NtMMP1 mRNA during BY-2 suspension cell cultivation. Total RNA (12 μg) was loaded for each time point and the blot was hybridized with a BglII/HindIII NtMMP1 probe. The ethidium bromide bands confirm equal loading. Lane 1: day 4, 2: day 5; 3: day 6; 4: day 7; 5: day 8; 6: day 9; 7: day 10 after sub-culturing. B: Endogenous NtMMP1 protein was detected during BY-2 suspension cell cultivation by immunoblot analysis. Equal amounts of BY-2 cell extracts were separated by 12% SDS PAGE and blotted onto a nitrocellulose membrane. NtMMP1 was detected with anti-LeMMP antiserum diluted 1:2000 and a goat anti-rabbit HRP-labeled Fc-specific antibody diluted 1:5000 (Dianova) followed by the ECL procedure. C: recombinant NtMMP1-apo transiently produced in tobacco leaves as positive control. Lane 1: day 4; 2: day 5; 3: day 6; 4: day 7; 5: day 8; 6: day 9; 7: day 10 after sub-culturing.