Subject: FlyBase query
Dear Gillian,
I got this message from Dirk Bohmann via Julia Zetlinger:
>>Dear Dr. Bohmann,
>>
>>I am curating your paper for FlyBase:
>>
>>Zeitlinger and Bohmann, 1999, Development 126(17): 3947--3956
>>
>>and I have a question about the GFP constructs you used.
>>
>>You used 2 constructs:
>>
>>UAS-GFP
>>UAS-EGFP
>>
>>I would be grateful if you could tell me some more details about the
>>UAS-EGFP construct, so that I can work out whether it corresponds to a
>>UAS-GFP line we have already in FlyBase or whether it is new.
>>
>>Specifically, could you tell me:
>>
>>a. a reference where UAS-EGFP has been used.
>>b. who you got the line from.
>>c. presumably the 'EGFP' means that this is not the unmodified GFP -
>>does EGFP have the S65T mutation, or some other mutation that enhances
>>the fluorescence ?
>>
>>I look forward to hearing from you,
>>
>>Gillian
It was simply done by cloning an EcoRI-NotI fragment (about 800 bp) from
the pEGFP-N1 vector of Clontech into the pUAST vector (Brand and Perrimon).
pEGFP-N is the one with the 'enhanced GFP', which contains two aa
substitutions:
Phe-64 to Leu and Ser65 to Thr.
We got a few transformants, all with inserts on the second chromosome only.
Maura