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  • pCAG-FIP200 (1-640)

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    Plasmid for mammalian expression of FIP200 NTD:ATG13:ULK1 complex.

  • pGST2-TAX1BP1

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    Plasmid: For the purification of TAX1BP1 protein.

  • Introducing GolgiTag to Cells and Immunoprecipitation of Golgi

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    Protocol for generating GolgiTag cells and immunoprecipitation of Golgi for subsequent -omics analyses.

  • pLenti-CMV-DNAJC5

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    Mammalian expression of DNAJC5

  • Axonal and somatodendritic proteomes of dopamine neurons in the mouse brain

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    Dopamine (DA) neurons modulate neural circuits and behaviors via dopamine release from expansive, long range axonal projections. The elaborate cytoarchitecture of DA neurons is embedded within complex brain tissues, making it difficult to access the DA neuronal proteome using conventional methods. Here, we demonstrate APEX2 proximity labeling within genetically targeted neurons in the mouse brain, enabling subcellular proteomics with cell type-specificity. By combining APEX2 biotinylation with mass spectrometry, we mapped the somatodendritic and axonal proteomes of DA neurons. Our dataset reveals the proteomic architecture underlying axonal transport, dopamine transmission, and axonal metabolism in DA neurons. We find a significant enrichment of proteins encoded by Parkinson’s disease-linked genes in dopaminergic axons, including proteins with previously undescribed axonal localization. Our proteomic datasets comprise a significant resource for axonal and DA neuronal cell biology, while the methodology developed here will enable future studies of other neural cell types. This mass spectrometry proteomics dataset is a part of "Subcellular proteomics of dopamine neurons in the mouse brain" (Hobson et. al, 2022)

  • Cell line construction and maintenance for Lyso-IP with or without genes linked with lysosomal storage disease

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    Associated with the following preprint (published September 30th 2021): Progranulin deficiency results in reduced bis(monoacylglycero)phosphate (BMP) levels and gangliosidosis Sebastian Boland, Sharan Swarup, Yohannes A. Ambaw, Ruth C. Richards, Alexander W. Fischer, Shubham Singh, Geetika Aggarwal, Salvatore Spina, Alissa L. Nana, Lea T. Grinberg, William W. Seeley, Michal A. Surma, Christian Klose, Joao A. Paulo, Andrew D. Nguyen, J. Wade Harper, Tobias C. Walther, Robert V. Farese Jr. bioRxiv 2021.09.30.461806; doi: https://doi.org/10.1101/2021.09.30.461806

  • pCAG-VPS34

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    Plasmid for mammalian expression of VPS34

  • Immunohistochemistry Protocol – Chu lab (Wichman)

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    This protocol describes immunohistochemistry.

  • HyDrop-ATAC v1.0

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    Step-by-step protocol for execution of HyDrop-ATAC.

  • Golgi immunopurification (Golgi-IP) for subcellular lipid profiling

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    This protocol provides details for preparing Golgi-IP lipidomics samples.

  • pHAGE-eGFP-TAX1BP1 CC3Δ

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    Plasmid

  • pCAG-OSF-ATG13 (2-197)

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    Plasmid: To make ATG101/ATG13 HORMA complex for expression in mammalian cells.

  • Image quantification

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    Image quantification.

  • FM1-43 endocytic uptake assay in HIPSC derived neurons

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    FM1-43 is a lipophilic styryl dye, nontoxic to the cells, virtually nonfluorescent in aqueous media. The dye will become fluorescent once it binds to the plasma membrane. In neurons that are actively releasing neurotransmitters, these dyes become internalized within the recycled synaptic vesicles, and the nerve terminals become brightly stained. This shows an increase in the fluorescence intensity of FM1-43. When the cells undergo another stimulation (through KCL or electrical) the FM1-43 dye containing vesicles will be released through exocytosis. Thus, resulting in a decrease in the FM1-43 fluorescence intensity.

  • Flag co-immunoprecipitation

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    Protocol used to pull down V5-FLAG-Gcase interacting proteins in HEK cells

  • Freezing of hPSCs grown on MEFs

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    This protocol describes the standard procedure of freezing human pluripotent stem cells (hPSCs), which were grown on inactivated mouse embryonic fibroblasts (MEFs).

  • Rotarod Test to assess motor coordination in a mouse parkinsonian model

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    This protocol evaluates motor coordination in the rotarod assay in parkinsonian mice.

  • Cryo-ET of S. cerevisiae during starvation (tomograms)

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    Cryo-ET of S. cerevisiae during starvation (tomograms).

  • powereQTL – An R package for calculating sample size and power of bulk tissue and single-cell eQTL analysis

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    R package for calculating sample size and power of bulk tissue and single-cell eQTL analysis

  • pHAGE-APEX2-FLAG-OPTN

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