GFP- Cherry FRET construct

[Maria Calvo-2]

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Dear all,
We are doing intensity based FRET experiments between proteins tagged to
GFP and mCherry.
We would like to have an intramolecular FRET positive construct with GFP and
Cherry.
Does anyone have this construct or know of any group who has it?
Thank you,
Maria Calvo

___________________________________

Dra. Maria Calvo

Unitat de Microscòpia Confocal
:: CCiTUB- Centres Científics i Tecnològics
Facultat de Medicina
Universitat de Barcelona- IDIBAPS
C/ Casanova 143
Barcelona 08036

Tel: [phone]/39930
Fax: [phone]
E-mail: [hidden email]
___________________________________

[Ippei Kotera]

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Hi Maria,

The following group has published FLIM analysis of EGFP-mCherry.

Biophys J. 2008 Sep 15;95(6):2976-88

I should mention that my former colleagues had been trying to optimize
various Aequorea FP/non-Aequorea FP FRET pairs, but no pair had
surpassed the FRET efficiency of Aequorea CFP/YFP. Theoretically
speaking, red FPs like mCherry should be a good acceptor for GFP because
of the larger overlap integral, but I haven't heard any one achieving
the FRET efficiency even close to the CFP/YFP pair. In many imaging
applications, the GFP/RFP pairs give much less (if any) overall FRET
signal than equivalent CFP/YFP pair. Our conclusion was that Aequorea
FPs have weak tendency to dimerize, and that's one of the important
factors to achieve high FRET efficiency. See also:

ACS Chem Biol. 2010 Feb 19;5(2):215-22.

Best,


Ippei



(05/05/2011 7:33 AM),

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Dear all,
> We are doing intensity based FRET experiments between proteins tagged to
> GFP and mCherry.
> We would like to have an intramolecular FRET positive construct with GFP and
> Cherry.
> Does anyone have this construct or know of any group who has it?
> Thank you,
> Maria Calvo
>
> ___________________________________
>
> Dra. Maria Calvo
>
> Unitat de Microscòpia Confocal
> :: CCiTUB- Centres Científics i Tecnològics
> Facultat de Medicina
> Universitat de Barcelona- IDIBAPS
> C/ Casanova 143
> Barcelona 08036
>
> Tel: [phone]/39930
> Fax: [phone]
> E-mail: [hidden email]
> ___________________________________

[Maria Calvo-2]

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http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
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Thank you Ippei, I will take that into account!
Best regards,
Maria Calvo

Al 05/05/2011 17:17, En/na Ippei Kotera ha escrit:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Hi Maria,
>
> The following group has published FLIM analysis of EGFP-mCherry.
>
> Biophys J. 2008 Sep 15;95(6):2976-88
>
> I should mention that my former colleagues had been trying to optimize
> various Aequorea FP/non-Aequorea FP FRET pairs, but no pair had
> surpassed the FRET efficiency of Aequorea CFP/YFP. Theoretically
> speaking, red FPs like mCherry should be a good acceptor for GFP because
> of the larger overlap integral, but I haven't heard any one achieving
> the FRET efficiency even close to the CFP/YFP pair. In many imaging
> applications, the GFP/RFP pairs give much less (if any) overall FRET
> signal than equivalent CFP/YFP pair. Our conclusion was that Aequorea
> FPs have weak tendency to dimerize, and that's one of the important
> factors to achieve high FRET efficiency. See also:
>
> ACS Chem Biol. 2010 Feb 19;5(2):215-22.
>
> Best,
>
>
> Ippei
>
>
>
> (05/05/2011 7:33 AM),
>> *****
>> To join, leave or search the confocal microscopy listserv, go to:
>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>> *****
>>
>> Dear all,
>> We are doing intensity based FRET experiments between proteins tagged to
>> GFP and mCherry.
>> We would like to have an intramolecular FRET positive construct with GFP and
>> Cherry.
>> Does anyone have this construct or know of any group who has it?
>> Thank you,
>> Maria Calvo
>>
>> ___________________________________
>>
>> Dra. Maria Calvo
>>
>> Unitat de Microscòpia Confocal
>> :: CCiTUB- Centres Científics i Tecnològics
>> Facultat de Medicina
>> Universitat de Barcelona- IDIBAPS
>> C/ Casanova 143
>> Barcelona 08036
>>
>> Tel: [phone]/39930
>> Fax: [phone]
>> E-mail: [hidden email]
>> ___________________________________
>

--


___________________________________

Dra. Maria Calvo

Unitat de Microscòpia Confocal
:: CCiTUB- Centres Científics i Tecnològics
Facultat de Medicina
Universitat de Barcelona- IDIBAPS
C/ Casanova 143
Barcelona 08036

Tel: [phone]/39930
Fax: [phone]
E-mail: [hidden email]
___________________________________

[Vitaly Boyko]

Have your coleagues also tested EYFP with mRFP1 and/or mCh
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Hi Ippei,

Have your coleagues also tested EYFP with mRFP1 and/or mCherry?
I agree that CeFP-EYPF is the best pair for FRET. In my hands EYFP-mRFP1 was ca.
2-fold better than YFP-mCherry and ca. 10-fold weaker than CeFP-EYFP pair.

Best,

Vitaly
301-515-7833
 




________________________________
From: Maria Calvo <[hidden email]>
To: [hidden email]
Sent: Tue, May 10, 2011 11:39:38 AM
Subject: Re: GFP- Cherry FRET construct

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To join, leave or search the confocal microscopy listserv, go to:
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Thank you Ippei, I will take that into account!
Best regards,
Maria Calvo

Al 05/05/2011 17:17, En/na Ippei Kotera ha escrit:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> *****
>
> Hi Maria,
>
> The following group has published FLIM analysis of EGFP-mCherry.
>
> Biophys J. 2008 Sep 15;95(6):2976-88
>
> I should mention that my former colleagues had been trying to optimize
> various Aequorea FP/non-Aequorea FP FRET pairs, but no pair had
> surpassed the FRET efficiency of Aequorea CFP/YFP. Theoretically
> speaking, red FPs like mCherry should be a good acceptor for GFP because
> of the larger overlap integral, but I haven't heard any one achieving
> the FRET efficiency even close to the CFP/YFP pair. In many imaging
> applications, the GFP/RFP pairs give much less (if any) overall FRET
> signal than equivalent CFP/YFP pair. Our conclusion was that Aequorea
> FPs have weak tendency to dimerize, and that's one of the important
> factors to achieve high FRET efficiency. See also:
>
> ACS Chem Biol. 2010 Feb 19;5(2):215-22.
>
> Best,
>
>
> Ippei
>
>
>
> (05/05/2011 7:33 AM),
>> *****
>> To join, leave or search the confocal microscopy listserv, go to:
>> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
>> *****
>>
>> Dear all,
>> We are doing intensity based FRET experiments between proteins tagged to
>> GFP and mCherry.
>> We would like to have an intramolecular FRET positive construct with GFP and
>> Cherry.
>> Does anyone have this construct or know of any group who has it?
>> Thank you,
>> Maria Calvo
>>
>> ___________________________________
>>
>> Dra. Maria Calvo
>>
>> Unitat de Microscòpia Confocal
>> :: CCiTUB- Centres Científics i Tecnològics
>> Facultat de Medicina
>> Universitat de Barcelona- IDIBAPS
>> C/ Casanova 143
>> Barcelona 08036
>>
>> Tel: [phone]/39930
>> Fax: [phone]
>> E-mail: [hidden email]
>> ___________________________________
>

--


___________________________________

Dra. Maria Calvo

Unitat de Microscòpia Confocal
:: CCiTUB- Centres Científics i Tecnològics
Facultat de Medicina
Universitat de Barcelona- IDIBAPS
C/ Casanova 143
Barcelona 08036

Tel: [phone]/39930
Fax: [phone]
E-mail: [hidden email]
___________________________________