Salk-CIRM Shared Resources Laboratory
The Cell Technologies and Engineering Core Facility (CTEC) at the Salk Institute supports research on human fibroblasts and stem cells, providing dedicated lab space, equipment, validated reagents, and extensively characterized cell lines. It generates patient-derived fibroblasts from healthy donors and Alzheimer’s patients, which are either studied directly, converted to induced pluripotent stem cells (iPSCs), or transdifferentiated into brain cell types for aging and neurodegeneration studies. iPSCs reset aging hallmarks, while directly reprogrammed cells retain donor aging signatures, making them ideal for studying the aging brain. The core ensures rigor and reproducibility through standardized protocols, reagent packages, and rigorous quality control.
Cell lines available for modeling of aging and Alzheimer’s and related dementias (ADRDs)
- Primary patient-derived dermal fibroblasts
- “iN-ready” dermal fibroblasts
- Patient-derived iPSCs
- Biopsy processing (fibroblast derivation)
- iPSC Derivation (Reprogramming from fibroblasts or PBMCs)-with full QC
- Custom genome editing
iPSC Quality Control (Core-generated lines tested by default; user lines, by request)
- Karyotyping
- Mycoplasma testing
- Short tandem repeat analysis
- Undifferentiated state panel
- Trilineage differentiation assay
- Sendai virus+Yamanaka factor retention
Fibroblast QC (by request)
- Mycoplasma
- Growth rate
- STR profiling (human only)
- Genome editing QC (by request)
- Mycoplasma/sterility testing
- Allele sequencing
- Cell line authentication by short tandem repeat analysis
Training
- Basic Stem Cell culture and maintenance
- Transdifferentiation of fibroblasts to neurons
Salk-CIRM SRL Training Course
The Salk Institute has designed a detailed course with two optional subcourses that will introduce researchers to foundational stem cell concepts and techniques and then delve into the more specialized topics of genome editing and the creation of stem cell-based neuronal models for studying aging and disorders of the brain. The course will be offered annually, beginning in Summer of 2026.
Basic Stem Cell Techniques Course
Techniques that will be taught include:
- Basic hPSC culture techniques including feeder free culture as well as culture on feeders
- Cryopreservation and thawing of hPSCs
- Basic characterization of pluripotency by immunocytochemistry and alkaline phosphatase
- Differentiation via embryoid body formation
Transdifferentiation of Somatic Cells to Neurons (led by Salk with contributions from UCSD)
Topics will include:
- History of direct conversion/transdifferentiation technology
- Deriving fibroblasts from skin biopsies that will be used for transdifferentiation
- How to assess the quality of fibroblasts prior to transdifferentiation
- The rationale for choosing different transcription factors for neuronal conversion
- Making and stable fibroblast lines for transdifferentiation
- Using cell sorting to select PSA-NCAM-expressing neurons after transdifferentiation
- Phenotyping neuronal cells (e.g., via immunochemistry and live cell imaging)
- Using transdiferentiation technology to study human aging and neurodegenerative conditions
- Latest technological advances to improve neuronal conversion
Genome Editing of Stem Cells and Primary Fibroblasts
Discussion topics include:
- The rationale and choice of iPSCs for genome editing
- Genome editing technologies and formats, when each is most appropriate
- The latest technological advances, CRISPR spin-off technologies
- Designing and generating DNA donor molecules for homologous recombination
- Special considerations when single-cell cloning iPSCs
- Editing primary fibroblasts prior to neural conversion
Hands-on activities will include:
- Delivery of CRISPR materials into cells in different formats, by different methods
- Validation by target site PCR and Sanger sequencing
- Colony picking to isolate clones
- Generation of a ssDNA donor molecule for knockin of a transgene
- Special considerations when single-cell cloning iPSCs
- Editing primary fibroblasts prior to neural conversion
- Basic Stem Cell Techniques will be designed and co-taught by Drs. Maria Carolina Marchetto and Jennifer Page.
- Genome Editing of Stem Cells and Primary Fibroblasts will be taught by Dr. Jennifer Page.
- Transdifferentiation of Somatic Cells to Neurons will be designed and taught by Drs. Gage, Allen, and Maria Carolina Marchetto.
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Basic Stem Cell Techniques [View References]
- Takahashi, K. et al. Induction of Pluripotent Stem Cells from Adult Human Fibroblasts by Defined Factors. Cell 131, 861–872 (2007).
- Mali, P. et al. Butyrate greatly enhances derivation of human induced pluripotent stem cells by promoting epigenetic remodeling and the expression of pluripotency-associated genes. Stem Cells 28, 713–720 (2010).
- Watanabe, K. et al. A ROCK inhibitor permits survival of dissociated human embryonic stem cells. Nat Biotechnol 25, 681–686 (2007).
- Codner, G. F. et al. Aneuploidy screening of embryonic stem cell clones by metaphase karyotyping and droplet digital polymerase chain reaction. BMC Cell Biol 17, (2016).
Genome Editing [View References]
- Cong, L., Ran, F.A., Cox, D., Lin, S., Barretto, R., Habib, N., Hsu, P.D., Wu, X., Jiang, W., Marraffini, L.A., et al. (2013). Multiplex genome engineering using CRISPR/Cas systems. Science 339, 819–823. 10.1126/science.1231143.
- Lin, S., Staahl, B.T., Alla, R.K., and Doudna, J.A. (2014). Enhanced homology-directed human genome engineering by controlled timing of CRISPR/Cas9 delivery. eLife 3, e04766. 10.7554/eLife.04766.
- Hockemeyer, D., and Jaenisch, R. (2016). Induced pluripotent stem cells meet genome editing. Cell Stem Cell 18, 573–586. 10.1016/j.stem.2016.04.013.
- Kime, C., Mandegar, M.A., Srivastava, D., Yamanaka, S., Conklin, B.R., and Rand, T.A. (2016).
- Efficient CRISPR/Cas9-Based Genome Engineering in Human Pluripotent Stem Cells. Curr. Protoc. Hum. Genet. 88, 21.4.1-21.4.23. 10.1002/0471142905.hg2104s88.
Direct Reprogramming (Fibro-iN) [View References]
- Mertens J, Paquola ACM, Ku M, Hatch E, Böhnke L, Ladjevardi S, McGrath S, Campbell B, Lee H, Herdy JR, Gonçalves JT, Toda T, Kim Y, Winkler J, Yao J, Hetzer MW, Gage FH. Directly Reprogrammed Human Neurons Retain Aging-Associated Transcriptomic Signatures and Reveal Age-Related Nucleocytoplasmic Defects. Cell Stem Cell. 2015 Dec 3;17(6):705-718. doi: 10.1016/j.stem.2015.09.001. Epub 2015 Oct 8. PMID: 26456686; PMCID: PMC5929130.
- Zheng X, Boyer L, Jin M, Mertens J, Kim Y, Ma L, Ma L, Hamm M, Gage FH, Hunter T. Metabolic reprogramming during neuronal differentiation from aerobic glycolysis to neuronal oxidative phosphorylation. Elife. 2016 Jun 10;5:e13374. doi: 10.7554/eLife.13374. PMID: 27282387; PMCID: PMC4963198.
- Mertens J, Reid D, Lau S, Kim Y, Gage FH. Aging in a Dish: iPSC-Derived and Directly Induced Neurons for Studying Brain Aging and Age-Related Neurodegenerative Diseases. Annu Rev Genet. 2018 Nov 23;52:271-293. doi: 10.1146/annurev-genet-120417-031534. Epub 2018 Sep 12. PMID: 30208291; PMCID: PMC6415910.
- Mertens J, Herdy JR, Traxler L, Schafer ST, Schlachetzki JCM, Böhnke L, Reid DA, Lee H, Zangwill D, Fernandes DP, Agarwal RK, Lucciola R, Zhou-Yang L, Karbacher L, Edenhofer F, Stern S, Horvath S, Paquola ACM, Glass CK, Yuan SH, Ku M, Szücs A, Goldstein LSB, Galasko D, Gage FH. Age-dependent instability of mature neuronal fate in induced neurons from Alzheimer’s patients. Cell Stem Cell. 2021 Sep 2;28(9):1533-1548.e6. doi: 10.1016/j.stem.2021.04.004. Epub 2021 Apr 27. PMID: 33910058; PMCID: PMC8423435.
Astrocytes [View References]
- Santos R, Vadodaria KC, Jaeger BN, Mei A, Lefcochilos-Fogelquist S, Mendes APD, Erikson G, Shokhirev M, Randolph-Moore L, Fredlender C, Dave S, Oefner R, Fitzpatrick C, Pena M, Barron JJ, Ku M, Denli AM, Kerman BE, Charnay P, Kelsoe JR, Marchetto MC, Gage FH. Differentiation of Inflammation-Responsive Astrocytes from Glial Progenitors Generated from Human Induced Pluripotent Stem Cells. Stem Cell Reports. 2017 Jun 6;8(6):1757-1769. doi: 10.1016/j.stemcr.2017.05.011. PMID: 28591655; PMCID: PMC5470172.
- Santos R, Mei A, Marchetto MC. Generation of inflammation-responsive astrocytes from glial progenitors derived from human pluripotent stem cells. STAR Protoc. 2022 Mar 17;3(2):101261. doi: 10.1016/j.xpro.2022.101261. PMID: 35313707; PMCID: PMC8933838.
- Patani R, Hardingham GE, Liddelow SA. Functional roles of reactive astrocytes in neuroinflammation and neurodegeneration. Nat Rev Neurol. 2023 Jul;19(7):395-409. doi: 10.1038/s41582-023-00822-1. Epub 2023 Jun 12. PMID: 37308616.
- Derevyanko A, Tao T, Allen NJ. Common alterations to astrocytes across neurodegenerative disorders. Curr Opin Neurobiol. 2025 Feb;90:102970. doi: 10.1016/j.conb.2025.102970. Epub 2025 Jan 28. PMID: 39879721.
Microglia [View References]
- Abud EM, Ramirez RN, Martinez ES, Healy LM, Nguyen CHH, Newman SA, Yeromin AV, Scarfone VM, Marsh SE, Fimbres C, Caraway CA, Fote GM, Madany AM, Agrawal A, Kayed R, Gylys KH, Cahalan MD, Cummings BJ, Antel JP, Mortazavi A, Carson MJ, Poon WW, Blurton-Jones M. iPSC-Derived Human Microglia-like Cells to Study Neurological Diseases. Neuron. 2017 Apr 19;94(2):278-293.e9. doi: 10.1016/j.neuron.2017.03.042. PMID: 28426964; PMCID: PMC5482419.
- Munro DAD, Bestard-Cuche N, McQuaid C, Chagnot A, Shabestari SK, Chadarevian JP, Maheshwari U, Szymkowiak S, Morris K, Mohammad M, Corsinotti A, Bradford B, Mabbott N, Lennen RJ, Jansen MA, Pridans C, McColl BW, Keller A, Blurton-Jones M, Montagne A, Williams A, Priller J. Microglia protect against age-associated brain pathologies. Neuron. 2024 Aug 21;112(16):2732-2748.e8. doi: 10.1016/j.neuron.2024.05.018. Epub 2024 Jun 18. PMID: 38897208.