Reference Report

Reference
Citation	Petersen, L.K., Stowers, R.S. (2011). A Gateway MultiSite Recombination Cloning Toolkit. PLoS ONE 6(9): e24531. (Export to RIS)
FlyBase ID	FBrf0215584
Publication Type	Research paper
PubMed ID	21931740
PubMed Abstract	The generation of DNA constructs is often a rate-limiting step in conducting biological experiments. Recombination cloning of single DNA fragments using the Gateway system provided an advance over traditional restriction enzyme cloning due to increases in efficiency and reliability. Here we introduce a series of entry clones and a destination vector for use in two, three, and four fragment Gateway MultiSite recombination cloning whose advantages include increased flexibility and versatility. In contrast to Gateway single-fragment cloning approaches where variations are typically incorporated into model system-specific destination vectors, our Gateway MultiSite cloning strategy incorporates variations in easily generated entry clones that are model system-independent. In particular, we present entry clones containing insertions of GAL4, QF, UAS, QUAS, eGFP, and mCherry, among others, and demonstrate their in vivo functionality in Drosophila by using them to generate expression clones including GAL4 and QF drivers for various trp ion channel family members, UAS and QUAS excitatory and inhibitory light-gated ion channels, and QUAS red and green fluorescent synaptic vesicle markers. We thus establish a starter toolkit of modular Gateway MultiSite entry clones potentially adaptable to any model system. An inventory of entry clones and destination vectors for Gateway MultiSite cloning has also been established (www.gatewaymultisite.org).
DOI	10.1371/journal.pone.0024531
Related Publication(s)
Recent Updates
Description	What does this section display? This section contains items that were added to this record for each release. It currently only tracks new links between this FlyBase report and other FlyBase data classes (e.g. genes, references, stocks) or controlled vocabulary terms (e.g. GO, anatomy terms). What does this section not display? This section does not currently display links that were removed or gene model changes.
Update Feed	Click the icon below to subscribe to this FlyBase record and receive updates automatically through your feed reader.
FB2013_03
FB2013_02
All updates	Click here to see a list of all updates to this record from FB2010_08 and on.
Associated Information
Comments
Associated Files
Other Information
Secondary IDs
Language of Publication	English
Additional Languages of Abstract
Also Published As
Parent Publication
Publication Type	Journal
Abbreviation	PLoS ONE
Title	PLoS ONE
Publication Year	2006-
ISBN/ISSN	1932-6203
Data from Reference
Alleles (32)
	Crei\ChR2^{CHETA.Ncra\QUAS.T:Avic\GFP-YFP} Crei\ChR2^{CHETA.Scer\UAS.T:Avic\GFP-YFP} Disc\RFP^{tdTomato.Ncra\QUAS.T:Uuuu\MyrPalm2,T:Ivir\HA1} Hsap\CD4^{20x.Scer\UAS.4X.T:Avic\GFP-EGFP,T:Hsap\MYC} Hsap\CD4^{20x.Scer\UAS.4X.T:Disc\RFP-mCherry,T:Ivir\HA1} Mmus\Cd8a^{IVS.20xScer\UAS.T:Avic\GFP} Mmus\Cd8a^{Scer\UAS.T:Avic\GFP} Mmus\Cd8a^{Scer\UAS.T:Disc\RFP-mCherry} Ncra\QF^iav.PP Ncra\QF^nompC.PP Ncra\QF^synj.PP Ncra\QF^TrpA1.PP Ncra\QF^trpl.PP n-syb^{Ncra\QUAS.4X.T:Avic\GFP-EGFP,T:Hsap\MYC} n-syb^{Ncra\QUAS.4X.T:Disc\RFP-mCherry,T:Ivir\HA1} n-syb^{Ncra\QUAS.T:Avic\GFP-EGFP,T:Hsap\MYC} n-syb^{Ncra\QUAS.T:Disc\RFP-mCherry,T:Ivir\HA1} n-syb^{Scer\UAS.T:Avic\GFP-EGFP} Rab3^{Ncra\QUAS.T:Avic\GFP-EGFP} Rab3^{Ncra\QUAS.T:Disc\RFP-mCherry} Scer\GAL4^iav.PP Scer\GAL4^nompC.PP Scer\GAL4^ppk.PG Scer\GAL4^Rh6.PD Scer\GAL4^scrt-11-6 Scer\GAL4^trp.PP Scer\GAL4^TrpA1.PP T:Avic\GFP^EGFP T:Avic\GFP^YFP T:Disc\RFP^mCherry Zzzz\HR^{e.Ncra\QUAS.T:Avic\GFP-YFP} Zzzz\HR^{e.Scer\UAS.T:Avic\GFP-YFP}
Constructs (31)
	P{20XUAS-CHETA.YFP} P{20XUAS-eNpHR3.0.YFP} P{20XUAS-hCD4.4XGFP.Myc} P{20XUAS-hCD4.4XmCherry.HA} P{20XUAS-IVS-mCD8::GFP} P{iav-QF.P} P{nompC-GAL4.P} P{nompC-QF.P} P{ppk-GAL4.G} P{QUAS-CHETA.YFP} P{QUAS-eNpHR3.0.YFP} P{QUAS-mtdTomato-3xHA} P{QUAS-n-syb.4XGFP.Myc} P{QUAS-n-syb.4XmCherry.HA} P{QUAS-n-syb.GFP.Myc} P{QUAS-n-syb.mCherry.HA} P{QUAS-Rab3.eGFP} P{QUAS-Rab3.mCherry} P{Rh6-GAL4.D} P{synj-QF.P} P{TrpA1-QF.P} P{trpl-QF.P} P{UAS-mCD8::GFP.L} P{UAS-mCD8.ChRFP} P{UAS-n-syb.eGFP} PBac{iav-GAL4.P} PBac{nompC-GAL4.P} PBac{QUAS-Rab3.eGFP} PBac{QUAS-Rab3.mCherry} PBac{TrpA1-GAL4.P} PBac{trp-GAL4.P}
Genes (17)
	Crei\ChR2 Disc\RFP Hsap\CD4 iav Mmus\Cd8a Ncra\QF nompC n-syb Rab3 Scer\GAL4 synj T:Hsap\MYC T:Ivir\HA1 trp TrpA1 trpl Zzzz\HR
Insertions (37)
	P{20XUAS-CHETA.YFP}attP2 P{20XUAS-CHETA.YFP}attP40 P{20XUAS-eNpHR3.0.YFP}attP2 P{20XUAS-eNpHR3.0.YFP}attP40 P{20XUAS-hCD4.4XGFP.Myc}attP2 P{20XUAS-hCD4.4XmCherry.HA}attP2 P{CaryP}attP2 P{CaryP}attP33 P{CaryP}attP40 P{CaryP}attP154 P{GawB}scrt^11-6 P{iav-QF.P}attP154 P{nompC-GAL4.P}attP154 P{nompC-QF.P}attP40 P{nompC-QF.P}attP154 P{QUAS-CHETA.YFP}attP2 P{QUAS-CHETA.YFP}attP33 P{QUAS-eNpHR3.0.YFP}attP2 P{QUAS-eNpHR3.0.YFP}attP33 P{QUAS-n-syb.4XGFP.Myc}attP40 P{QUAS-n-syb.4XmCherry.HA}attP40 P{QUAS-n-syb.GFP.Myc}attP40 P{QUAS-n-syb.mCherry.HA}attP40 P{QUAS-Rab3.eGFP}attP40 P{QUAS-Rab3.mCherry}attP40 P{synj-QF.P}attP40 P{TrpA1-QF.P}attP40 P{TrpA1-QF.P}attP154 P{trpl-QF.P}attP154 PBac{iav-GAL4.P}VK00014 PBac{nompC-GAL4.P}VK00014 PBac{QUAS-Rab3.eGFP}VK00027 PBac{QUAS-Rab3.mCherry}VK00027 PBac{TrpA1-GAL4.P}VK00014 PBac{trp-GAL4.P}VK00014 PBac{y⁺-attP-9A}VK00014 PBac{y⁺-attP-9A}VK00027
Natural transposons (2)
	P-element Tni\piggyBac