FtsH1 + FtsH5 | ATP-dependent zinc metalloprotease FtsH1 + FtsH5 (chloroplastic)
AS16 3930 | clonality: polyclonal | host: rabbit | reactivity: Arabidopsis thaliana, Nicotiana tabacum, Spinacia oleracea
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|Recommended dilution||1 : 5000 (WB)|
|Expected | apparent MW||
67.1 kD (Arabidopsis thaliana)
|Confirmed reactivity||Arabidopsis thaliana, Nicotiana tabacum, Spinacia oleracea|
|Not reactive in||No confirmed exceptions from predicted reactivity are currently known.|
Both FtsH5 (VAR1) and FtsH1 ahare high degree of homology therefore this antibody recognizes both proteins.
Total proteins were isolated from Arabidopsis (Arabidopsis thaliana) wild type (Col) and mutant lacking FtsH2 (yellow variegated2 [var2]). Samples were immediately frozen in liquid nitrogen and pulverized with a microtube homogenizer. Proteins were extracted by adding appropriate extraction buffer. After measurement of chlorophyll concentration, equally loaded supernatants (based on chlorophyll [0.5 µg chlorophyll/lane]) were subjected to SDS-PAGE analysis. Proteins were separated on 12% SDS-PAGE gel and blotted 1h to PVDF membrane. Blots were blocked in 1% BSA in PBST buffer for 1 h at room temperature. Then, blots were incubated in the primary antibody (anti-VAR1) at a dilution of 1:5000 for 1 h. After washing 2 times for 10 min in PBST buffer, blots were incubated in the secondary antibody (anti Rabbit IgG) at a dilution of 1:5000 for 1 h. Blots were washed 2 times for 10 min in PBST buffer. Luminata crescendo (Millipore) was used for signal detection. Images of the blots were obtained using ChemiDoc™ XRS (Bio-rad). Exposure time was 2 seconds.
Absence of FtsH2 in var2 mutant results in decreased amount of FtsH1 which together form a hetero-hexamer complex.
Courtesy of Dr. Yusuke Kato, Plant Light Acclimation Research Group, Okayama University, Japan
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