Genetics & Molecular Biology

CRISPR & Gene Perturbation

I have generated multiple CRISPR-edited iPSC cell lines (Parkin KO, α-synuclein KO, LRRK2 KO, GBA KO) to assess protein function and mimic genetic forms of neurodegenerative diseases. GBA knockout heightened vulnerability to fibril-induced inclusions, whereas α-synuclein knockout abolished pathology entirely. Validation includes PCR, Western blotting, karyotyping, and ICC for pluripotency markers.

RNA Interference & Oligonucleotide Biology

I design, optimize, and validate siRNA, shRNA, ASO, and SSO-mediated knockdown assays in iPSC-derived neurons and cancer cell lines.

Delivery methods include gymnotic uptake (media addition), lipid-based transfection, and electroporation/nucleofection. I integrate RT-qPCR, protein-level (Western/ELISA), and functional neuronal readouts to assess potency, durability, specificity, and mechanism-of-action.

I have miniaturized assays into 96/384-well formats with SOP and repeatability metrics for scalable, screening-ready execution, translating RNAi results into clear go/no-go and target-prioritization recommendations for preclinical programs.

Molecular Cloning & Construct Design

End-to-end cloning workflows including restriction cloning, Gibson assembly, Golden Gate cloning, PCR/primer design, bacterial transformation, colony screening, plasmid preparation (MiniPrep/MaxiPrep), Sanger sequencing, and endotoxin-free DNA prep. I design constructs for membrane-associated and endolysosomal proteins, with expression verification by flow cytometry and/or immunoassay. Stable cell line generation uses selection markers (Puromycin, G418, Blasticidin) with single-cell cloning and pool evaluation.

Viral Transduction & Expression

I produce and optimize lentiviral, adenoviral, and AAV/rAAV constructs for transduction in iPSC-derived neurons and mammalian cell lines. Key applications include: (1) lentiviral shRNA knockdown of LAMP2, which I demonstrated is sufficient to drive α-synuclein inclusion formation even without immune challenge — a critical finding from my Nature Neuroscience 2024 paper; (2) adenoviral overexpression of α-synuclein-HA to show that endogenous protein is actively recruited into pathological inclusions; and (3) reporter construct expression for live-cell tracking. I handle full vector design (cassettes, promoters, tags), MOI optimization, titering, and biosafety compliance across BSL-2 workflows.

Lentiviral shRNA-mediated LAMP2 knockdown in iPSC-derived DA neurons. Western blot confirms >80% reduction in LAMP2 protein. Confocal shows that LAMP2 loss alone — without immune challenge — is sufficient to induce α-synuclein inclusion formation, establishing lysosomal dysfunction as the critical vulnerability in Lewy body pathogenesis. From Bayati et al., Nature Neuroscience 2024.

Adenoviral overexpression of α-synuclein-HA in iPSC-derived DA neurons. Phospho-synuclein (pSyn S129) accumulation is dramatically enhanced under dual-hit conditions (PFF + IFN-γ), confirming that endogenous α-synuclein is recruited into pathological inclusions. Western blot quantification shows dose-dependent pSyn increase. Bayati et al., Nature Neuroscience 2024.

Lysosomal Biology & Protein Biochemistry

My work on lysosomal biology spans from organelle-level mechanistic studies to biochemical characterization. I showed that IFN-γ specifically downregulates LAMP1, LAMP2, TFEB, and NRF2 in DA neurons — but not cortical neurons — creating a selective vulnerability window for Lewy body formation (Nature Neuroscience 2024). I use galectin-3 as a reporter for lysosomal membrane permeabilization, revealing that fibrils rupture lysosomes and escape into the cytoplasm. My Lyso-IP approach (HA-tagged TMEM192-RFP + magnetic bead immunoprecipitation) confirmed PFF arrival at lysosomes within 2 minutes of uptake.

For protein biochemistry, I perform recombinant α-synuclein purification (Ni-NTA affinity), preformed fibril (PFF) generation (37°C shaking, 5 days, sonication), size exclusion chromatography (SEC), and DLS particle sizing. I developed a custom nanogold-PFF conjugation protocol — published in STAR Protocols (2023) as first author — enabling direct EM visualization of fibril trafficking without relying on antibody specificity.

Immunoprecipitation & Biochemical Fractionation

I apply biochemical fractionation and immunoprecipitation (IP/Co-IP) techniques to resolve protein compartmentalization and interaction networks. Key applications include: (1) sequential detergent extraction (Triton-soluble → SDS-soluble → insoluble pellet) to separate soluble, membrane-associated, and aggregated protein pools; (2) Lyso-IP (HA-tagged TMEM192-RFP + magnetic bead immunoprecipitation) to isolate intact lysosomes and confirm fibril cargo arrival within 2 minutes; and (3) native co-IP workflows for capturing transient protein–protein interactions under physiological conditions. Downstream readouts include Western blot, mass spectrometry, and activity assays.

Biochemical fractionation of α-synuclein inclusions from iPSC-derived DA neurons. Sequential extraction (Triton-soluble → SDS-soluble → pellet) reveals enrichment of α-synuclein and HDAC6 (an aggresome marker) in the insoluble pellet fraction under dual-hit conditions, confirming the formation of detergent-resistant aggregates. Bayati et al., Nature Neuroscience 2024.

RT-qPCR & Gene Expression Data

RT-qPCR datasets from iPSC-derived neuronal, cardiac, and pancreatic differentiation experiments as well as viral host-response profiling. Expression is normalized to geometric mean of housekeeping genes (GAPDH, ACTB, RPLP0) and presented as fold change (2^−ΔΔCt) relative to undifferentiated iPSCs or vehicle controls. All primer pairs validated with efficiency 90–110% and single melt-curve peaks. 500+ primer pairs validated across the lab.

Flow Cytometry Quantitative Data

Multicolor flow cytometry datasets from immunophenotyping experiments across immune cell profiling (lymph node and spleen tissue from LRRK2 mouse models), iPSC differentiation QC, and cancer immunology co-cultures. Acquired on BD LSRFortessa and Cytek Aurora platforms. Analyzed in FlowJo v10 with tSNE/UMAP dimensionality reduction. All data gated on live singlets (Zombie Aqua⁻, FSC-A/FSC-H).

Virology — Viral Internalization, Entry & Transduction

Quantitative virology datasets spanning SARS-CoV-2 spike internalization via clathrin-mediated endocytosis (CME), pseudoviral infectivity assays, host-directed antiviral screening, and viral vector transduction optimization. This work led to the first paper demonstrating the cellular entry mechanism of SARS-CoV-2 (J. Biol. Chem. 2021; 500+ citations) and contributed to host-directed antiviral strategies targeting PIKfyve and CSNK2 kinases in collaboration with UNC Chapel Hill, GSK, and the Structural Genomics Consortium.