Hi Dave,
I'm writing about a paper I am curating for FlyBase.
Davis et al., Neuron 19: 561
Genetic analysis of the mechanisms controlling target selection:
target-derived Fasciclin II regulates the pattern of synapse formation.
In their materials and methods they describe two things made by you:
UAS-FasII (PEST+)
and
UAS-B-galrep6
We have one UAS-Fas2 construct from you but it is not the same one (record
included at the end, for your info). It would be marvellous if you could
give me a line or two about its construction, then I can do a decent job of
introducing the PEST+ version to FlyBase. I'll curate your reply as a
personal communication to FlyBase.
The same goes for the UAS-lacZ.
Many thanks for your help,
Rachel.
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Hi Rachel-
UAS-rep6 (lacZ reporter construct)
I did not make this construct. It was originally made by Janice Fischer
and is described in her paper Nature, 1988 Apr 28, 332(6167):853-6. The
parent UAS-type vector was created by Janice to see if GAL4 works at all in
flies (the grandfather of all fly GAL4 papers), and is not the same as
Andrea Brand's UAST construct.
UAS-FasII PEST+
This was a little more complicated. I used a FasII PEST+ cDNA that Gregg
Helt in Corey's lab had isolated from a cDNA library. I first modified the
ends of the cDNA to remove all 5' and 3' untranslated sequences. At the 5'
end: Not I, Xba I, AGA TAA AA ATG..... At the 3' end: TAA GG EcoRV
HindIII. To create the UAST-FasII (PEST+) construct, the UAST vector was
first digested with Asp718 and the 5' overhangs filled in with Klenow. It
was then digested with NotI and CIAP treated. The FasII cDNA was cut with
HindIII, the overhangs were filled in, and subsequently digested with NotI.
The two fragments were then ligated together and the construct transformed.
hope all of this is helpful!
dave
Dave Lin phone: 510-642-9887
265 LSA FAX: 510-643-6791
UC Berkeley CA 94720
e-mail: davelin@XXXX