Plant Transformation

Location

Morgenstelle 1
2. Stock, Raum Nr.218

Morgenstelle 32
5. Stock, Raum 5X17

phone 73223

Booking Trafos - click me
 

Equipment

 

2 sterile benches, 3 growth rooms

 

Coordinator

Dr. Kenneth W Berendzen


 

Transformation technicians

 

Transformation Unit Protocols


Cell Culture 01.doc
Cell Culture Maintenance (Arabidopsis, BY2)


BY-2-protopl.doc
BY-2 Tobacco Cell Suspension Protoplasts

 

PEG transfection protocol 2018

Arabodopsis Cell Culture Protoplasts
 

REFERENCING

Please cite for the standard protoplast transfection protocol after Feb. 2017:

Mehlhorn D.G., Wallmeroth N., Berendzen K.W., Grefen C. (2018) 2in1 Vectors Improve In Planta BiFC and FRET Analyses. In: Hawes C., Kriechbaumer V. (eds) The Plant Endoplasmic Reticulum. Methods in Molecular Biology, vol 1691. Humana Press, New York, NY.

Please cite for the 96-well transfection protocol:

Berendzen KW, Böhmer M, Wallmeroth N, Peter S, Vesić M, Zhou Y, Tiesler FK, Schleifenbaum F, Harter K. Screening for in planta protein-protein interactions combining bimolecular fluorescence complementation with flow cytometry. Plant Methods. 2012 Jul 12;8(1):25.

 

Transfections performed before Feb.2017 cite:

Schütze K, Harter K, Chaban C (2009) Bimolecular fluorescence complementation (BiFC) to study protein-protein interactions in living plant cells. Methods Mol Biol 479: 189-202.

 

STABLE TRANSFORMATIONS
PROTOCOLS


PotatoStabile.doc
Stable Transformation protocol for potato

Transformation protocol for tobacco

Protocol for tomato stable transformation
Protocol for tomato stable transformation

 

REFERENCING

Please cite for tomato transformations:

Wittmann et al. 2015. Plant Pathology. Doi: 10.1111/ppa.12417

http://onlinelibrary.wiley.com/doi/10.1111/ppa.12417/abstract

 

OTHER PROTOCOLS

Protoplasts isolation from leaves of Arabidopsis thaliana
 

 

 

1. The unit offers Agrobacterium-mediated transformation of potato, tobacco and tomato.
2. Regenaration and cultivation of transformants.
3. PEG-mediated transformation of protoplasts from cell cultures or mesophyll cells (Arabidopsis or tobacco).
4. Production of cell cultures (wild type).

 

Fig. Z-stack image of protoplasts liberated from Arabidopsis thaliana leaf tissue.

 

Facs

Fig. Examination of the protoplat transformation efficiency by flow cytometric anaylsis (FCA).

Protoplasts isolated from root cell culture are transformed with a 35S::GFP construct (12kb) as a routine control. As an alternative to calculating the transformation efficiency, we have used FCA as a means for detecting cells expressing GFP. The negative population is marked in black. Cells that are strongly expressing GFP have a fluoresence signal that is well above that of the total plant cell population‘s green autofluoresence (cells marked in dark green); cells marked in light green are those with weaker GFP fluoresence emission. In this example 23% of the cells were transformed with GFP, and 50% of those are strongly expressing GFP.

 

 


Current Publications


Albert I, Böhm H, Albert M, Feiler CE, Imkampe J, Wallmeroth N, Brancato C, Raaymakers TM, Oome S, Zhang H, Krol E, Grefen C, Gust A, Chai J, Hedrich R, Van den Ackerveken G, Nürnberger T (2015). An RLP23-SOBIR1-BAK1 complex mediates NLP-triggered immunity. Nature Plants, 15140, doi: 10.1038/NPLANTS.2015.140.

J. Wittmann, C. Brancato, K. W. Berendzen and B. Dreiseikelmann (2015).
Development of a tomato plant resistant to Clavibacter michiganensis using the endolysin gene of bacteriophage CMP1 as a transgene.
Plant Pathology DOI: 10.1111/ppa.12417