[{"data":1,"prerenderedAt":-1},["ShallowReactive",2],{"detail-sidebar-cat-0-en-105":3,"doc-seo-451088-105":59,"doc-detail-451088-en":130},{"code":4,"msg":5,"data":6},0,"success",[7,13,18,23,28,33,38,43,48,51,55],{"id":8,"doc_module":4,"doc_module_name":9,"category_name":10,"show_sort_weight":11,"slug":12},1,"Document","Story & Novel",90,"story-novel",{"id":14,"doc_module":4,"doc_module_name":9,"category_name":15,"show_sort_weight":16,"slug":17},2,"Literature",80,"literature",{"id":19,"doc_module":4,"doc_module_name":9,"category_name":20,"show_sort_weight":21,"slug":22},4,"Exam",70,"exam",{"id":24,"doc_module":4,"doc_module_name":9,"category_name":25,"show_sort_weight":26,"slug":27},5,"Comic",60,"comic",{"id":29,"doc_module":4,"doc_module_name":9,"category_name":30,"show_sort_weight":31,"slug":32},6,"Technology",50,"technology",{"id":34,"doc_module":4,"doc_module_name":9,"category_name":35,"show_sort_weight":36,"slug":37},7,"Healthcare",40,"healthcare",{"id":39,"doc_module":4,"doc_module_name":9,"category_name":40,"show_sort_weight":41,"slug":42},8,"Research & Report",30,"research-report",{"id":44,"doc_module":4,"doc_module_name":9,"category_name":45,"show_sort_weight":46,"slug":47},9,"Religion & Spirituality",20,"religion-spirituality",{"id":46,"doc_module":4,"doc_module_name":9,"category_name":49,"show_sort_weight":46,"slug":50},"World Cup","world-cup",{"id":52,"doc_module":4,"doc_module_name":9,"category_name":53,"show_sort_weight":52,"slug":54},10,"Lifestyle","lifestyle",{"id":56,"doc_module":4,"doc_module_name":9,"category_name":57,"show_sort_weight":24,"slug":58},19,"General","general",{"code":4,"msg":60,"data":61},"ok",{"site_id":62,"language":63,"slug":64,"title":65,"keywords":66,"description":67,"schema_data":68,"social_meta":123,"head_meta":125,"extra_data":127,"updated_unix":129},105,"en","protocol-for-differentiating-patient-derived-ipscs-into-photoreceptor-like-cells","Protocol for differentiating patient-derived iPSCs into photoreceptor-like cells","","Protocol modeling occult macular dystrophy uses induced pluripotent stem cells (iPSCs) generated from patient blood cells to produce photoreceptor-like cells. The method introduces a tetracycline-inducible PiggyBac system to stably deliver doxycycline-inducible CRX and NEUROD1, then differentiates iPSCs into iPRCs and validates photoreceptor marker expression. The workflow includes guidance for doxycycline-inducible induction, neomycin selection, single-cell cloning, colony expansion, and marker-based assessment to support mutation-focused studies of inherited retinal dystrophies.",{"@graph":69,"@context":122},[70,84,105],{"@type":71,"itemListElement":72},"BreadcrumbList",[73,77,79,82],{"item":74,"name":75,"@type":76,"position":8},"https://docshare.wps.com","Home","ListItem",{"item":78,"name":9,"@type":76,"position":14},"https://docshare.wps.com/document/",{"item":80,"name":35,"@type":76,"position":81},"https://docshare.wps.com/document/healthcare/",3,{"item":83,"name":65,"@type":76,"position":19},"https://docshare.wps.com/document/protocol-for-differentiating-patient-derived-ipscs-into-photoreceptor-like-cells/451088/",{"url":83,"name":65,"@type":85,"image":86,"author":91,"headline":65,"publisher":94,"fileFormat":97,"inLanguage":63,"description":67,"dateModified":98,"datePublished":99,"encodingFormat":97,"isAccessibleForFree":100,"interactionStatistic":101},"DigitalDocument",{"url":87,"@type":88,"width":89,"height":90},"https://docshare.wps.com/thumbnails/protocol-for-differentiating-patient-derived-ipscs-into-photoreceptor-like-cells/451088.png","ImageObject",300,407,{"name":92,"@type":93},"Chumphorn","Person",{"url":74,"name":95,"@type":96},"DocShare","Organization","application/pdf","2026-11-01","2026-09-30",true,{"@type":102,"interactionType":103,"userInteractionCount":29},"InteractionCounter",{"@type":104},"ViewAction",{"@type":106,"mainEntity":107},"FAQPage",[108,114,118],{"name":109,"@type":110,"acceptedAnswer":111},"What is the main purpose of this protocol?","Question",{"text":112,"@type":113},"To differentiate patient-derived iPSCs into photoreceptor-like cells using a doxycycline-inducible CRX and NEUROD1 PiggyBac system for inherited retinal dystrophy research.","Answer",{"name":115,"@type":110,"acceptedAnswer":116},"How are CRX and NEUROD1 introduced into iPSCs?",{"text":117,"@type":113},"They are delivered via a tetracycline-inducible PiggyBac vector using doxycycline induction, enabling stable expression of a doxycycline-inducible CRX-T2A-NEUROD1 construct.",{"name":119,"@type":110,"acceptedAnswer":120},"What steps are included to obtain and verify differentiated photoreceptor-like cells?",{"text":121,"@type":113},"The workflow includes neomycin selection of stably transduced iPSCs, single-cell cloning and colony expansion, differentiation of selected clones, and validation of photoreceptor markers to assess differentiation.","https://schema.org",{"og:url":83,"og:type":124,"og:title":65,"og:site_name":95,"og:description":67},"article",{"robots":126,"canonical":83},"index,follow",{"doc_id":128,"site_id":62},451088,1791046492,{"code":4,"msg":5,"data":131},{"doc_id":128,"user_id":132,"nickname":92,"user_avatar":133,"doc_module":4,"category_id":34,"category_name":35,"doc_title":65,"doc_description":67,"doc_content":134,"file_id":135,"file_url":136,"file_type":137,"file_size":138,"view_count":29,"is_deleted":4,"is_public":8,"is_downloadable":8,"audit_status":8,"page_count":46,"language":139,"language_code":63,"site_id":62,"html_lang":63,"table_of_contents":140,"faqs":141,"seo_title":142,"seo_description":67,"update_tm":143,"read_time":31,"preview_page_count":4},2336475401981,"https://ap-avatar.wpscdn.com/avatar/22000c94efd8d5204d?x-image-process=image/resize,m_fixed,w_180,h_180&k=1786935347598174694","ll  \nOPEN ACCESS  \nProtocol  \nProtocol for differentiating patient-derived iPSCs into photoreceptor-like cells  \nRetinal biopsies are rarely feasible, limiting direct studies of inherited retinal dystrophies (IRDs) . We present a protocol modeling occult macular dystrophy, an autosomal-dominant IRD, using induced pluripotent stem cells (iPSCs) generated from patient blood cells. We describe steps to differentiate iPSCs into photoreceptor-like cells by transducing CRX and NEUROD1 via a tetracycline-inducible PiggyBac vector. We also detail procedures for the validation of photoreceptor markers to assess differentiation. This protocol enables mutation-focused investigation of IRD pathogenesis.  \nPublisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.  \nYang Pan, Takeshi Iwata  \n[takeshi.iwata@kankakuki.jp](takeshi.iwata@kankakuki.jp)  \nHighlights  \nSteps for inducing iPSCs with PiggyBac delivery of doxycyclineinducible CRXNEUROD1  \nInstructions for neomycin selection of stably transducediPSCs  \nGuidance on singlecell cloning and colony expansion  \nSteps for differentiating selected iPSC clones into photoreceptorlike cells (iPRCs)  \nPan & Iwata, STAR Protocols 7, 104283  \nMarch 20, 2026 © 2025 The Author(s) . Published by Elsevier Inc.  \n[https://doi.org/10.1016/](https://doi.org/10.1016/)[ ](https://doi.org/10.1016/)[j.xpro.2025.104283](j.xpro.2025.104283)  \nll  \nOPEN ACCESS  \nProtocol  \nProtocol for differentiating patient-derived iPSCs into photoreceptor-like cells  \nYang Pan 1,2 and Takeshi Iwata 1,3,*  \n1Division of Molecular and Cellular Biology, National Institute of Sensory Organs, NHO Tokyo Medical Center, Tokyo 152- 8902, Japan  \n2Technical contact  \n3Lead contact  \n*Correspondence: [takeshi.iwata@kankakuki.jp](takeshi.iwata@kankakuki.jp)  \n[https://doi.org/10.1016/j.xpro.2025.104283](https://doi.org/10.1016/j.xpro.2025.104283)  \nSUMMARY  \nRetinal biopsies are rarely feasible, limiting direct studies of inherited retinal dystrophies (IRDs) . We present a protocol modeling occult macular dystrophy, an autosomal-dominant IRD, using induced pluripotent stem cells (iPSCs) generated from patient blood cells. We describe steps to differentiate iPSCs into photoreceptor-like cells by transducing CRX and NEUROD1 via a tetracycline-inducible PiggyBac vector. We also detail procedures for the validation of photoreceptor markers to assess differentiation. This protocol enables mutation-focused inves  \ntigation of IRD pathogenesis.  \nFor complete details on the use and execution of this protocol, please refer to  \nPan et al.1  \nBEFORE YOU BEGIN  \nInduced pluripotent stem cells (iPSCs) can be generated from patient-derived cells.2 We have established multiple iPSC lines from circulating T cells of patients with inherited retinal diseases and their unaffected family members using the Sendai virus reprogramming system.1 ,3–5 Initially, iPSCs are cultured on mitomycin C-inactivated mouse embryonic fibroblast feeder cells, then transferred to a feeder-free DEF-CS culture system. Although direct feeder-free human iPSC generation methods are now available,6 feeder cells still improve reprogramming efficiency, especially for older patients commonly seen in inherited retinal diseases.  \nOverexpression of CRX and NEUROD1 in human iPSCs enables reproducible generation of induced photoreceptor-like cells (iPRCs) .7 This protocol describes an induction of iPRCs from patient-derived iPSCs using a PiggyBac delivery system to stably introduce a doxycycline-inducible CRXT2A-NEUROD1 construct (Figures 1A and 1B) . The protocol is optimized for iPSCs derived from an R45W-carrying occult macular dystrophy (OMD) patient and a related control, 1 but is also applicable to other iPSC lines.  \nAll cell culture steps should be performed under sterile conditions in a biosafety cabinet, and cells should be maintained in a humidified 37◦ C incubator with 5% CO2.  \nInnovation ","cbCaig9Q4M4TRi1l","https://ap.wps.com/l/cbCaig9Q4M4TRi1l","pdf",4560953,"English","# Summary\n# Before you begin\n## iPSC generation and culture background\n## CRX/NEUROD1 induction rationale\n# Innovation\n# Institutional permissions\n# Preparation of iPSC culture\n## Timing","[{\"question\":\"What is the main purpose of this protocol?\",\"answer\":\"To differentiate patient-derived iPSCs into photoreceptor-like cells using a doxycycline-inducible CRX and NEUROD1 PiggyBac system for inherited retinal dystrophy research.\"},{\"question\":\"How are CRX and NEUROD1 introduced into iPSCs?\",\"answer\":\"They are delivered via a tetracycline-inducible PiggyBac vector using doxycycline induction, enabling stable expression of a doxycycline-inducible CRX-T2A-NEUROD1 construct.\"},{\"question\":\"What steps are included to obtain and verify differentiated photoreceptor-like cells?\",\"answer\":\"The workflow includes neomycin selection of stably transduced iPSCs, single-cell cloning and colony expansion, differentiation of selected clones, and validation of photoreceptor markers to assess differentiation.\"}]","Protocol for differentiating patient-derived iPSCs into photoreceptor-like cells | PDF",1790734348]