HaloTag: a new versatile reporter gene system in plant cells.
HaloTag Interchangeable Labeling Technology (HaloTag) was originally developed for mammalian cell analysis. In this report, the use of HaloTag is demonstrated in plant cells for the first time. This system allows different fluorescent colours to be used to visualize the localization of the non-fluorescent HaloTag protein within living cells. A vector was constructed which expresses the HaloTag protein under the control of the 35S promoter of cauliflower mosaic virus. The functionality of the HaloTag construct was tested in transient assays by (i) transforming tobacco protoplasts and (ii) using biolistic transformation of intact leaf cells of tobacco and poplar plants. Two to fourteen days after transformation, the plant material was incubated with ligands specific for labelling the HaloTag protein, and fluorescence was visualized by confocal laser scanning microscopy. The results demonstrate that HaloTag technology is a flexible system which generates efficient fluorescence in different types of plant cells. The ligand-specific labelling of HaloTag protein was not hampered by the plant cell wall.
Lang
Christina
C
Institut für Pflanzenbiologie, Technische Universität Braunschweig, Humboldtstrasse 1, D-38106 Braunschweig, Germany.
Schulze
Jutta
J
Mendel
Ralf-R
RR
Hänsch
Robert
R
D002473:Cell Wall / Q000378:metabolism; D002478:Cells, Cultured; D017930:Genes, Reporter; D005822:Genetic Vectors; D008164:Luminescent Proteins / Q000032:analysis; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D032107:Populus / Q000166:cytology / Q000235:genetics* / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods; D011523:Protoplasts / Q000166:cytology / Q000378:metabolism; D014026:Nicotiana / Q000166:cytology / Q000235:genetics* / Q000378:metabolism
D008164:Luminescent Proteins