Available Technology

A FLEX switch targets Channelrhodopsin-2 to multiple cell types for imaging and long-range circuit mapping.

Plasmids are available from Addgene.com

• AAV-FLEX-rev-ChR2(H134R)-mCherry
• AAV-FLEX-rev-ChR2-tdtomato (recommended)
• pJ241-FLEX

These vectors are prone to recombination. This is a well known issue with these AAV vectors and is due to the inverted terminal repeats (ITRs) required for rAAV production. To minimize recombination, we propagate these plasmids in Stbl2 cells from Invitrogen. Also, to minimize recombination, cells should be cultured at 30 ºC.

Note that these cultures will grow slowly (20 h for minipreps). Better yields and culture times are obtained with 2xYT as the media. This is strongly recommended.

Because recombination may still happen occasionally, we do a panel of restriction digestions to assess whether the ITRs are in tact. Separate digestions with PvuII, Sma1, and SnaB1 should be performed. The expected patterns can be calculated from the attached sequence available on addgene.com.

Viruses for Cre-dependent optogentics based on FLEX switch available from University of Pennsylvania Vector Core

Fly stocks generated in the Rubin lab are generally deposited in the Bloomington Drosophila Stock Center. For more specialized stocks please contact us directly.

In January 2011, the Rubin lab began depositing our collection of GAL4 driver lines in the Bloomington Stock Center at the rate of approximately 100 lines a week and all lines should be available for distribution by August 31, 2012.

Rubin lab Plasmid Constructs are available from Addgene, which has distributed over 430 of the plasmids listed below.

Plasmids from Pfeiffer et al. 2008 and Pfeiffer et al. 2010   

BP plasmid vector backbones are derived from pBPGUw and contain the pUC19-derived bacterial origin of replication and ampicillin resistance gene, the PhiC31 attB site, the mini-white marker for identification of transformants in Drosophila, and the DSCP basal promoter. Abbreviations:  U, DSCP basal promoter; w, mini-white marker; nls, nuclear localization signal; IVS, intervening sequence within the 5’ UTR; WPRE, a Woodchuck Hepatitis Virus Posttranscriptional Regulatory Element within the 3’ UTR; 3’ UTR Term., the transcriptional terminator; and pBP, plasmid BP backbone.

Janelia Farm Reporter Construct (JFRC) backbones are derived from pBDP. In addition, all vectors also contain a basal promoter derived from hsp70 and an SV40 transcriptional terminator.

a = Drosophila codon-optimized transgene 

 Plasmids from Nern et al PNAS 2011: Multiple new site-specific recombinases for use in manipulating animal genomes

 Plasmids from Pfeiffer et al PNAS 2012: Using translational enhancers to increase transgene expression in Drosophila 

Enables one to overlay images acquired in the same focal plane via (1) laser scanning fluorescence microscopy and (2) conventional Infrared Differential Interference Microscopy (IR-DIC).

Written by Lakshmi Ramasamy in ID&F for the Murphy Lab.

Lead author:  Sean Eddy

Lead author:  Eric Nawrocki

The Multi-Worm Tracker is available on Sourceforge.

Frank Midgley (Janelia Scientific Computing) worked with Vivek Jayaraman, Mitya Chklovskii and others at Janelia on this freely available software tool for neural circuit visualization. It allows users to dynamically represent connectivity and information flow at different levels of a nervous system, and can also serve as a front-end for storage of other types of data (e.g., physiological or anatomical). More information available at: Neuroptikon.org

FlyFizz is an evolving webspace dedicated to enabling the exchange of information relating to one growing subfield of Drosophila brain physiology: understanding how neural circuits generate behavior by applying electrophysiological and optical imaging techniques, particularly in behaving flies. We hope this space will become a repository for supplemental information regarding published techniques, as well as a community forum for discussion, software distribution and job postings.

Learn more at FlyFizz.org

Plasmids available from Addgene.com:
    CMV-GCaMP3
    CMV-GCaMP5G  (improved deltaF/F0, lower F0)

Viruses available from University of Pennsylvania Viral Vector Core:
    AAV2/1.hSynap.GCaMP3.3.SV40
    AAV2/5.hSynap.GCaMP3.3.SV40
    AAV2/9.hSynap.GCaMP3.3.SV40
    AAV2/1.hSynap.Flex.GCaMP3.3.SV40
    AAV2/5.hSynap.Flex.GCaMP3.3.SV40
    AAV2/9.hSynap.Flex.GCaMP3.3.SV40

Flies available from Bloomington Stock Center:
    P{UAS-GCaMP3.T}attP40
    P{20XUAS-GCaMP3}su(Hw)attP8
    P{20XUAS-GCaMP3}attP18
    P{20XUAS-GCaMP3}attP2
    PBac{20XUAS-GCaMP3}VK00005

C. elegans distributed upon request:
    none