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Populus trichocarpa Coproporphyrinogen III oxidase (PtCPO) localizes in sub-chloroplastic structures

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posted on 2014-05-30, 17:56 authored by Cécile LorrainCécile Lorrain, Sophien KamounSophien Kamoun, Ben PetreBen Petre

The open reading frame (ORF) of PtCPO (see sequence below) was amplified by PCR from the hybrid poplar Beaupré (P. trichocarpa x P. deltoides) leaf cDNAs. The ORF was fused with the N-terminus of a green fluorescent protein (GFP) into the pICH86988 in order to create an in-frame PtCPO-GFP protein fusion, under the control of a 35S-Ω promoter. The final vector was inserted into Agrobacterium tumefaciens (GV3101 strain). The PtCPO-GFP protein fusion was transiently expressed in N. benthamiana leaf cells using A. tumefaciens leaf infiltration. Two days after infiltration, pavement cells were observed by live-cell imaging with a laser-scanning confocal microscope. Chlorophyll and GFP were excited at 488 nm, and their fluorescent signals were collected between 505-525 nm and 680-700 nm, respectively. Images present a single optical section of 0.8 µm. White arrowheads indicate discrete areas (diameter: 0.1 to 0.3 µm) within the chloroplasts. Scale bar: 5 µm. >PtCPO (98.7 % amino acid identity with Potri.011G023900.1, Phytozome portal http://www.phytozome.net/genePage.php?search=1&detail=1&crown&method=0&searchText=transcriptid%3A27003187/)

 

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