[Arabidopsis] Can overly expressed GFP by 35S be a secretory protein?

Sarah Wyatt via arab-gen%40net.bio.net (by wyatts At ohio.edu)
Tue Dec 19 08:41:43 EST 2006


you both are correct.  It is quenched but can be "rescued" if you will by 
bathing cells in a pH buffer.  here is the reference to confirm those 
observations.  Scott, A. Wyatt, S.E., Tsou, P.-L., Robertson, D., and 
Allen, N.S. (1999) A model system for plant cell biology:  GFP imaging in 
living onion epidermal cells.  BioTechniques 26, 1127-1132

sincerely, sarah



--On Monday, December 18, 2006 11:58 AM +0000 John Runions 
<jrunions At brookes.ac.uk> wrote:

>
>    Dear Hee-Jin,
>    I suspect that Tobias is absolutely correct about the low pH quenching
>    the fluorescence of GFP in the apoplast.  As evidence of this, we have
>    two fusions that should mark the Golgi bodies; 35s ST-GFP and 35s
>    ST-mRFP.  Both mark Golgi bodies beautifully and the ST-mRFP shows up
>    very brightly in the apoplast marking all of the cell boundaries.
>    There is no reason why one version of this should be secreted when the
>    other isn't.
>    Regards, John.
>    Hee-Jin Kim wrote:
>
> Dear Arab-gen and colleagues
>
> How are you doing?
>
> Does anybody know if overly expressed GFP with 35S can be found
> extracellular f raction of cells in addition to the typical localizations
> (cytoplasm and nucleo plasm) of GFP ?  One of my soluble protein (no
> signal peptide), abundantly ove r-expressed mGFP5 fusion with 35S  was
> found in the cell wall fractions by West ern blot although microscopic
> picture showed most portion of the fusion protein  is in cytoplasm.
>
> I found a paper that 31% of overly expressed mGFP5-ER with 35S in BY2 was
> secre ted into medium according to Western blot analyses (Liu et al.,
> 2001, J Biotech nology 87: 1-16).  However,  I could not find so far if
> overly expressed GFP al one in plant cells can be secreted.
> Any comment how the levels of GFP expressions can affect localization
> would be  appreciated.
>
> Hee-Jin
>
>
>
>
>
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>
>    --
>
>    *********************************
>    C. John Runions, Ph.D.
>    School of Life Sciences
>    Oxford Brookes University
>    Oxford, UK
>    OX3 0BP
>    email:  [3]jrunions At brookes.ac.uk
>    phone: +44 (0) 1865 483 964
>
>    web: [4]http://www.brookes.ac.uk/lifesci/runions/HTMLpages/index.html
>
>
>    New - Oxford Brookes Master's in [5]Bioimaging with Molecular
>    Technology
>
> References
>
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________________________________

Sarah Wyatt, PhD
Associate Professor
Environmental and Plant Biology
Ohio University
Athens, OH 45701
phone:(740) 593-1133
FAX: (740) 593-1130
http://www.plantbio.ohiou.edu/epb/faculty/faculty/sew.htm



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