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However, there is currently a lack of comprehensive evaluation and guidance for available reference-based deconvolution methods in cell-type deconvolution analysis. In this study, we introduce Deconvolution Evaluator (Deconer), a comprehensive toolkit for the evaluation of reference-based deconvolution methods. Deconer provides various simulated and real gene expression datasets, including both bulk and single-cell sequencing data, and offers multiple visualization interfaces. By utilizing Deconer, we conducted systematic comparisons of 16 reference-based deconvolution methods from different perspectives, including method robustness, accuracy in deconvolving rare components, signature gene selection performance, and external reference construction capability. We also performed an in-depth analysis of the application scenarios and challenges in cell-type deconvolution methods. Finally, we provided constructive suggestions for users to select and develop cell-type deconvolution algorithms. This study provides novel insights for researchers, assisting them in choosing appropriate toolkits, applying solutions in clinical contexts, and advancing the development of deconvolution tools tailored to gene expression data. The tutorials, manual, source code, and demo data of Deconer are publicly available at https:\/\/honchkrow.github.io\/Deconer\/ and https:\/\/ngdc.cncb.ac.cn\/biocode\/tool\/7577.<\/jats:p>","DOI":"10.1093\/gpbjnl\/qzaf009","type":"journal-article","created":{"date-parts":[[2025,2,18]],"date-time":"2025-02-18T07:02:42Z","timestamp":1739862162000},"source":"Crossref","is-referenced-by-count":5,"title":["Deconer: An Evaluation Toolkit for Reference-based Deconvolution Methods Using Gene Expression Data"],"prefix":"10.1093","volume":"23","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-4264-5758","authenticated-orcid":false,"given":"Wei","family":"Zhang","sequence":"first","affiliation":[{"name":"Center of Intelligent Medicine, School of Control Science and Engineering, Shandong University , Jinan 250061,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8585-1917","authenticated-orcid":false,"given":"Xianglin","family":"Zhang","sequence":"additional","affiliation":[{"name":"Department of Clinical Laboratory, The Second Hospital, Cheeloo College of Medicine, Shandong University , Jinan 250033,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9781-3360","authenticated-orcid":false,"given":"Qiao","family":"Liu","sequence":"additional","affiliation":[{"name":"Department of Statistics, Stanford University , Stanford, CA 94305,","place":["USA"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1546-6458","authenticated-orcid":false,"given":"Lei","family":"Wei","sequence":"additional","affiliation":[{"name":"Ministry of Education Key Laboratory of Bioinformatics; Center for Synthetic and Systems Biology; Bioinformatics Division, Beijing National Research Center for Information Science and Technology; Department of Automation, Tsinghua University , Beijing 100084,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6854-7270","authenticated-orcid":false,"given":"Xu","family":"Qiao","sequence":"additional","affiliation":[{"name":"Center of Intelligent Medicine, School of Control Science and Engineering, Shandong University , Jinan 250061,","place":["China"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3599-7678","authenticated-orcid":false,"given":"Rui","family":"Gao","sequence":"additional","affiliation":[{"name":"Center of Intelligent Medicine, School of Control Science and Engineering, Shandong University , Jinan 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