Subject: FlyBase query
Dear Dr. Klaembt,
I wrote to A. Michelson about a construct he used in a paper (as I
think it is a new construct that FlyBase doesn't have a record of yet)
that he obtained from you and he suggest I write to you to confirm
details of this construct.
The paper was:
Halfon et al, 2000, Cell 103(1): 63--74
and the construct is an activated UAS-pnt construct, which was called
'UAS-pntP2Act' or 'UAS-pntP2VP16' (<up></up> = superscript) in the paper.
here is what I asked A. Michelson about the construct:
> 2. UAS-pntAct (<up></up> = superscript)
>
> This construct is called 'UAS-pntP2VP16' in the Materials and Methods.
>
> I cannot find any record of an activated UAS-pnt construct in FlyBase,
> so I think it must be new (if it has been published previously, please
> could you tell me the reference).
>
> If it is new, I would be grateful if you could provide some details
> about the construct (which I would curate as a personal communication
> from you to FlyBase) so that users of FlyBase can see what the
> construct consists of.
>
> Could you tell me:
>
> a. who made the construct (this helps in naming it) ?
> b. is it the P2 isoform of pnt (as P2 was in the symbol) ?
> c. what is the nature of the activating mutation \- presumably this is
> what the VP16 stands for ?
> d. what vector was used to make the construct, e.g. pUAST, pCaSpeR
> (this helps in working out the marker allele in the construct) ?
here is what he replied:
> 2. UAS-PntP2VP16 = UAS-PntP2Act was constructed and transgenic lines
> were generated by Christian Klambt; to my knowledge, the details of its
> contruction have not been published; however, my understanding is that it
> contains the strong transcriptional activation domain of the herpes simplex
> virus VP16 protein fused to a part of the PntP2 coding region; you would
> have to contact Christian if you really need to know the precise details of
> the construct; in any case, you should verify this question with him since
> he is the actual source of the materials
>
> 3. I assume that pUAST must have been the vector used to make
> UAS-PntP2VP16, but again Christian would be the one to confirm that
> question.
I would be grateful if you could confirm these details, specifically
whether it is the 'P2' isoform of pnt that is used, whether 'VP16' does
mean the transcriptional activation domain of herpes VP16 protein, and
whether the vector used was pUAST.
The information about this construct would be recorded in FlyBase as a
personal communication from you to FlyBase,
I look forward to hearing from you,
Gillian
\--------------------------------------------------------------
Gillian Millburn.
FlyBase (Cambridge),
\--------------------------------------------------------------
>
Subject: Re: FlyBase query
>Dear Dr. Klaembt,
>
>I wrote to A. Michelson about a construct he used in a paper (as I
>think it is a new construct that FlyBase doesn't have a record of yet)
>that he obtained from you and he suggest I write to you to confirm
>details of this construct.
>
>The paper was:
>
>Halfon et al, 2000, Cell 103(1): 63--74
>
>and the construct is an activated UAS-pnt construct, which was called
>'UAS-pntP2Act' or 'UAS-pntP2VP16' (<up></up> = superscript) in the paper.
>
>here is what I asked A. Michelson about the construct:
>
>> 2. UAS-pntAct (<up></up> = superscript)
>>
>> This construct is called 'UAS-pntP2VP16' in the Materials and Methods.
>>
>> I cannot find any record of an activated UAS-pnt construct in FlyBase,
>> so I think it must be new (if it has been published previously, please
>> could you tell me the reference).
>>
>> If it is new, I would be grateful if you could provide some details
>> about the construct (which I would curate as a personal communication
>> from you to FlyBase) so that users of FlyBase can see what the
>> construct consists of.
>>
>> Could you tell me:
>>
>> a. who made the construct (this helps in naming it) ?
the construct was made by Klaus Dücker from the Hafen lab in Zürich, the
transgenic flies were generated in my lab-
>> b. is it the P2 isoform of pnt (as P2 was in the symbol) ?
it is the pointed P2 isoform
>> c. what is the nature of the activating mutation \- presumably this is
>> what the VP16 stands for ?
the activation domain is from VP16, it relaces the pointed domain including
the MAPkinase phosphorylation site
>> d. what vector was used to make the construct, e.g. pUAST, pCaSpeR
the construct was inserted in pUAST and injected in a w1118 background.
best wishes christian klaembt
>--------------------------------------------------------------
>Gillian Millburn.
>
>FlyBase (Cambridge),
>--------------------------------------------------------------
Dr. Christian Klämbt
Institut fuer Neurobiologie
Universität Münster
Badestrasse 9
D-48149 Münster Germany