{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,5,12]],"date-time":"2026-05-12T23:04:06Z","timestamp":1778627046996,"version":"3.51.4"},"reference-count":51,"publisher":"MDPI AG","issue":"14","license":[{"start":{"date-parts":[[2024,7,11]],"date-time":"2024-07-11T00:00:00Z","timestamp":1720656000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Key Research and Development Program of China","award":["2022YFD2400100"],"award-info":[{"award-number":["2022YFD2400100"]}]},{"name":"National Key Research and Development Program of China","award":["2024XT0103"],"award-info":[{"award-number":["2024XT0103"]}]},{"name":"National Key Research and Development Program of China","award":["2023TD19"],"award-info":[{"award-number":["2023TD19"]}]},{"name":"National Key Research and Development Program of China","award":["32002371"],"award-info":[{"award-number":["32002371"]}]},{"name":"National Key Research and Development Program of China","award":["tstp20221149"],"award-info":[{"award-number":["tstp20221149"]}]},{"name":"National Key Research and Development Program of China","award":["2017ASTCP-ES06"],"award-info":[{"award-number":["2017ASTCP-ES06"]}]},{"name":"Central Public-interest Scientific Institution Basal Research Fund CAFS","award":["2022YFD2400100"],"award-info":[{"award-number":["2022YFD2400100"]}]},{"name":"Central Public-interest Scientific Institution Basal Research Fund CAFS","award":["2024XT0103"],"award-info":[{"award-number":["2024XT0103"]}]},{"name":"Central Public-interest Scientific Institution Basal Research Fund CAFS","award":["2023TD19"],"award-info":[{"award-number":["2023TD19"]}]},{"name":"Central Public-interest Scientific Institution Basal Research Fund CAFS","award":["32002371"],"award-info":[{"award-number":["32002371"]}]},{"name":"Central Public-interest Scientific Institution Basal Research Fund CAFS","award":["tstp20221149"],"award-info":[{"award-number":["tstp20221149"]}]},{"name":"Central Public-interest Scientific Institution Basal Research Fund CAFS","award":["2017ASTCP-ES06"],"award-info":[{"award-number":["2017ASTCP-ES06"]}]},{"name":"National Nature Science Foundation of China","award":["2022YFD2400100"],"award-info":[{"award-number":["2022YFD2400100"]}]},{"name":"National Nature Science Foundation of China","award":["2024XT0103"],"award-info":[{"award-number":["2024XT0103"]}]},{"name":"National Nature Science Foundation of China","award":["2023TD19"],"award-info":[{"award-number":["2023TD19"]}]},{"name":"National Nature Science Foundation of China","award":["32002371"],"award-info":[{"award-number":["32002371"]}]},{"name":"National Nature Science Foundation of China","award":["tstp20221149"],"award-info":[{"award-number":["tstp20221149"]}]},{"name":"National Nature Science Foundation of China","award":["2017ASTCP-ES06"],"award-info":[{"award-number":["2017ASTCP-ES06"]}]},{"name":"Taishan Scholars Program","award":["2022YFD2400100"],"award-info":[{"award-number":["2022YFD2400100"]}]},{"name":"Taishan Scholars Program","award":["2024XT0103"],"award-info":[{"award-number":["2024XT0103"]}]},{"name":"Taishan Scholars Program","award":["2023TD19"],"award-info":[{"award-number":["2023TD19"]}]},{"name":"Taishan Scholars Program","award":["32002371"],"award-info":[{"award-number":["32002371"]}]},{"name":"Taishan Scholars Program","award":["tstp20221149"],"award-info":[{"award-number":["tstp20221149"]}]},{"name":"Taishan Scholars Program","award":["2017ASTCP-ES06"],"award-info":[{"award-number":["2017ASTCP-ES06"]}]},{"name":"National Ten-Thousands Talents Special Support Program","award":["2022YFD2400100"],"award-info":[{"award-number":["2022YFD2400100"]}]},{"name":"National Ten-Thousands Talents Special Support Program","award":["2024XT0103"],"award-info":[{"award-number":["2024XT0103"]}]},{"name":"National Ten-Thousands Talents Special Support Program","award":["2023TD19"],"award-info":[{"award-number":["2023TD19"]}]},{"name":"National Ten-Thousands Talents Special Support Program","award":["32002371"],"award-info":[{"award-number":["32002371"]}]},{"name":"National Ten-Thousands Talents Special Support Program","award":["tstp20221149"],"award-info":[{"award-number":["tstp20221149"]}]},{"name":"National Ten-Thousands Talents Special Support Program","award":["2017ASTCP-ES06"],"award-info":[{"award-number":["2017ASTCP-ES06"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>DNA methylation is a key epigenetic mechanism orchestrating gene expression networks in many biological processes. Nonetheless, studying the role of specific gene methylation events in fish faces challenges. In this study, we validate the regulation of DNA methylation on empty spiracles homeobox 2 (emx2) expression with decitabine treatment in Chinese tongue sole testis cells. We used the emx2 gene as the target gene and developed a new DNA methylation editing system by fusing dnmt3a with catalytic dead Cas9 (dCas9) and demonstrated its ability for sequence-specific DNA methylation editing. Results revealed that utilizing dCas9-dnmt3a to target emx2 promoter region led to increased DNA methylation levels and decreased emx2 expression in Chinese tongue sole testis cells. More importantly, the DNA methylation editing significantly suppressed the expression of MYC proto-oncogene, bHLH transcription factor (myc), one target gene of emx2. Furthermore, we assessed the off-target effects of dCas9-dnmt3a and confirmed no significant impact on the predicted off-target gene expression. Taken together, we developed the first DNA methylation editing system in marine species and demonstrated its effective editing ability in Chinese tongue sole cells. This provides a new strategy for both epigenetic research and molecular breeding of marine species.<\/jats:p>","DOI":"10.3390\/ijms25147637","type":"journal-article","created":{"date-parts":[[2024,7,12]],"date-time":"2024-07-12T08:47:41Z","timestamp":1720774061000},"page":"7637","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["CRISPR\/dCas9-Mediated DNA Methylation Editing on emx2 in Chinese Tongue Sole (Cynoglossus semilaevis) Testis Cells"],"prefix":"10.3390","volume":"25","author":[{"given":"Yanxu","family":"Sun","sequence":"first","affiliation":[{"name":"College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China"},{"name":"State Key Laboratory of Mariculture Biobreeding and Sustainable Goods, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1754-4018","authenticated-orcid":false,"given":"Hong-Yan","family":"Wang","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Mariculture Biobreeding and Sustainable Goods, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Binghua","family":"Liu","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Mariculture Biobreeding and Sustainable Goods, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Bowen","family":"Yue","sequence":"additional","affiliation":[{"name":"College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China"},{"name":"State Key Laboratory of Mariculture Biobreeding and Sustainable Goods, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Qian","family":"Liu","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Mariculture Biobreeding and Sustainable Goods, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yuyan","family":"Liu","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Mariculture Biobreeding and Sustainable Goods, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6128-0099","authenticated-orcid":false,"given":"Ivana F.","family":"Rosa","sequence":"additional","affiliation":[{"name":"Department of Structural and Functional Biology, Institute of Biosciences, S\u00e3o Paulo State University (UNESP), Botucatu 01049-010, Brazil"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0620-7526","authenticated-orcid":false,"given":"Lucas B.","family":"Doretto","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Mariculture Biobreeding and Sustainable Goods, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shenglei","family":"Han","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Mariculture Biobreeding and Sustainable Goods, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Lei","family":"Lin","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Mariculture Biobreeding and Sustainable Goods, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xiaoling","family":"Gong","sequence":"additional","affiliation":[{"name":"College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306, China"},{"name":"Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources (Shanghai Ocean University), Ministry of Education, Shanghai 201306, China"},{"name":"National Demonstration Center for Experimental Fisheries Science Education, Shanghai Ocean University, Shanghai 201306, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6953-2203","authenticated-orcid":false,"given":"Changwei","family":"Shao","sequence":"additional","affiliation":[{"name":"State Key Laboratory of Mariculture Biobreeding and Sustainable Goods, Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Qingdao 266071, China"},{"name":"Laboratory for Marine Fisheries Science and Food Production Processes, Qingdao Marine Science and Technology Center, Qingdao 266237, China"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2024,7,11]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"777","DOI":"10.1038\/nrc907","article-title":"Deregulated homeobox gene expression in cancer: Cause or consequence?","volume":"2","year":"2002","journal-title":"Nat. Rev. Cancer"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"427","DOI":"10.1007\/s004290050110","article-title":"Emx2 developmental expression in the primordia of the reproductive and excretory systems","volume":"196","author":"Pellegrini","year":"1997","journal-title":"Anat. Embryol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1653","DOI":"10.1242\/dev.124.9.1653","article-title":"Defects of urogenital development in mice lacking Emx2","volume":"124","author":"Miyamoto","year":"1997","journal-title":"Dev. (Camb. Engl.)"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"5893","DOI":"10.1210\/en.2010-0915","article-title":"Abnormal epithelial cell polarity and ectopic epidermal growth factor receptor (EGFR) expression induced in Emx2 KO embryonic gonads","volume":"151","author":"Kusaka","year":"2010","journal-title":"Endocrinology"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"247","DOI":"10.1186\/s13046-021-02048-9","article-title":"Empty spiracles homeobox genes EMX1 and EMX2 regulate WNT pathway activation in sarcomagenesis","volume":"40","author":"Carnero","year":"2021","journal-title":"J. Exp. Clin. Cancer Res. CR"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"176","DOI":"10.1016\/j.mce.2011.06.010","article-title":"Interaction between sex hormones and WNT\/\u03b2-catenin signal transduction in endometrial physiology and disease","volume":"358","author":"Wang","year":"2012","journal-title":"Mol. Cell. Endocrinol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"515","DOI":"10.1038\/s41419-021-03801-w","article-title":"Regulation of sarcomagenesis by the empty spiracles homeobox genes EMX1 and EMX2","volume":"12","author":"Carnero","year":"2021","journal-title":"Cell Death Dis."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"604","DOI":"10.1101\/gr.162172.113","article-title":"Epigenetic modification and inheritance in sexual reversal of fish","volume":"24","author":"Shao","year":"2014","journal-title":"Genome Res."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"469","DOI":"10.1038\/s41588-022-01040-z","article-title":"DNMT3A-dependent DNA methylation is required for spermatogonial stem cells to commit to spermatogenesis","volume":"54","author":"Dura","year":"2022","journal-title":"Nat. Genet."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1703","DOI":"10.1093\/nar\/gkw1112","article-title":"Efficient targeted DNA methylation with chimeric dCas9-Dnmt3a-Dnmt3L methyltransferase","volume":"45","author":"Stepper","year":"2017","journal-title":"Nucleic Acids Res."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"41","DOI":"10.1186\/s40035-022-00314-0","article-title":"CRISPR\/dCas9-Dnmt3a-mediated targeted DNA methylation of APP rescues brain pathology in a mouse model of Alzheimer\u2019s disease","volume":"11","author":"Park","year":"2022","journal-title":"Transl. Neurodegener."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"233","DOI":"10.1016\/j.cell.2016.08.056","article-title":"Editing DNA Methylation in the Mammalian Genome","volume":"167","author":"Liu","year":"2016","journal-title":"Cell"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"193","DOI":"10.1080\/03602532.2018.1437446","article-title":"Effects of 5-Aza-2\u2019-deoxycytidine (decitabine) on gene expression","volume":"50","author":"Seelan","year":"2018","journal-title":"Drug Metab. Rev."},{"key":"ref_14","first-page":"315","article-title":"Chapter 14\u2014Impact of epigenetic modifiers on the immune system","volume":"Volume 16","author":"Kabelitz","year":"2020","journal-title":"Epigenetics of the Immune System"},{"key":"ref_15","first-page":"9","article-title":"Chapter 2\u2014DNA methyltransferase inhibitors (DNMTis) as sensitizing agents to overcome chemoresistance","volume":"Volume 15","author":"Wu","year":"2021","journal-title":"Epigenetic Regulation in Overcoming Chemoresistance"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"2503","DOI":"10.1016\/j.cell.2021.03.025","article-title":"Genome-wide programmable transcriptional memory by CRISPR-based epigenome editing","volume":"184","author":"Nunez","year":"2021","journal-title":"Cell"},{"key":"ref_17","first-page":"E2125","article-title":"Targeted DNA demethylation of the Arabidopsis genome using the human TET1 catalytic domain","volume":"115","author":"Gardiner","year":"2018","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"5615","DOI":"10.1093\/nar\/gkw159","article-title":"Repurposing the CRISPR-Cas9 system for targeted DNA methylation","volume":"44","author":"Vojta","year":"2016","journal-title":"Nucleic Acids Res."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"2119","DOI":"10.1038\/s41467-019-10146-8","article-title":"Targeted removal of epigenetic barriers during transcriptional reprogramming","volume":"10","author":"Baumann","year":"2019","journal-title":"Nat. Commun."},{"key":"ref_20","doi-asserted-by":"crossref","unstructured":"Marx, N., Gr\u00fcnwald-Gruber, C., Bydlinski, N., Dhiman, H., Ngoc Nguyen, L., Klanert, G., and Borth, N. (2018). CRISPR-Based Targeted Epigenetic Editing Enables Gene Expression Modulation of the Silenced Beta-Galactoside Alpha-2,6-Sialyltransferase 1 in CHO Cells. Biotechnol. J., 13.","DOI":"10.1002\/biot.201700217"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"11","DOI":"10.1038\/s41556-020-00620-7","article-title":"CRISPR technologies for precise epigenome editing","volume":"23","author":"Nakamura","year":"2021","journal-title":"Nat. Cell Biol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"1525","DOI":"10.1093\/cvr\/cvy111","article-title":"DNA methylation of a PLPP3 MIR transposon-based enhancer promotes an osteogenic programme in calcific aortic valve disease","volume":"114","author":"Mkannez","year":"2018","journal-title":"Cardiovasc. Res."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"S168","DOI":"10.1111\/raq.12020","article-title":"What determines growth potential and juvenile quality of farmed fish species?","volume":"5","author":"Valente","year":"2013","journal-title":"Rev. Aquac."},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Aykut, B., Ochs, M., Radhakrishnan, P., Brill, A., H\u00f6cker, H., Schwarz, S., Weissinger, D., Kehm, R., Kulu, Y., and Ulrich, A. (2017). EMX2 gene expression predicts liver metastasis and survival in colorectal cancer. BMC Cancer, 17.","DOI":"10.1186\/s12885-017-3556-2"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"347","DOI":"10.1016\/S0166-2236(00)01608-8","article-title":"Emx homeogenes and mouse brain development","volume":"23","author":"Cecchi","year":"2000","journal-title":"Trends Neurosci."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"8","DOI":"10.1016\/j.mod.2017.06.006","article-title":"EMX2 activates slow myosin heavy chain 2 gene expression in embryonic muscle fibers","volume":"147","author":"Hatch","year":"2017","journal-title":"Mech. Dev."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"451","DOI":"10.1016\/S0092-8674(00)81532-9","article-title":"Methylation-induced repression\u2014Belts, braces, and chromatin","volume":"99","author":"Bird","year":"1999","journal-title":"Cell"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1335","DOI":"10.1038\/s41467-022-28940-2","article-title":"DNA methylation-free Arabidopsis reveals crucial roles of DNA methylation in regulating gene expression and development","volume":"13","author":"He","year":"2022","journal-title":"Nat. Commun."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"661","DOI":"10.1038\/s41551-023-01037-x","article-title":"Stable expression of large transgenes via the knock-in of an integrase-deficient lentivirus","volume":"7","author":"Chavez","year":"2023","journal-title":"Nat. Biomed. Eng."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"435","DOI":"10.1007\/978-1-0716-0290-4_24","article-title":"CRISPR\/Cas9-Mediated Genome Engineering of Primary Human B Cells","volume":"2115","author":"Laoharawee","year":"2020","journal-title":"Methods Mol. Biol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"191","DOI":"10.1146\/annurev-biochem-013118-111730","article-title":"Evaluating and Enhancing Target Specificity of Gene-Editing Nucleases and Deaminases","volume":"88","author":"Kim","year":"2019","journal-title":"Annu. Rev. Biochem."},{"key":"ref_32","first-page":"109","article-title":"CRISPR\/Cas9-based epigenome editing: An overview of dCas9-based tools with special emphasis on off-target activity","volume":"164\u2013165","author":"Vojta","year":"2019","journal-title":"Methods"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1093\/gigascience\/giy011","article-title":"Genome-wide determination of on-target and off-target characteristics for RNA-guided DNA methylation by dCas9 methyltransferases","volume":"7","author":"Lin","year":"2018","journal-title":"GigaScience"},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Yamazaki, T., Hatano, Y., Handa, T., Kato, S., Hoida, K., Yamamura, R., Fukuyama, T., Uematsu, T., Kobayashi, N., and Kimura, H. (2017). Targeted DNA methylation in pericentromeres with genome editing-based artificial DNA methyltransferase. PLoS ONE, 12.","DOI":"10.1371\/journal.pone.0177764"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"49","DOI":"10.1016\/j.aquaculture.2013.01.024","article-title":"Identification of two novel female-specific DNA sequences in half-smooth tongue sole, Cynoglossus semilaevis","volume":"388\u2013391","author":"Wang","year":"2013","journal-title":"Aquaculture"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"673","DOI":"10.1007\/s10695-015-0037-x","article-title":"Establishment and characterization of a gonad cell line from half-smooth tongue sole Cynoglossus semilaevis pseudomale","volume":"41","author":"Sun","year":"2015","journal-title":"Fish Physiol. Biochem."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"935","DOI":"10.1016\/j.bbrc.2007.05.208","article-title":"Follicle-stimulating hormone signaling and Foxl2 are involved in transcriptional regulation of aromatase gene during gonadal sex differentiation in Japanese flounder, Paralichthys olivaceus","volume":"359","author":"Yamaguchi","year":"2007","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"333","DOI":"10.1095\/biolreprod.107.064246","article-title":"Sexual dimorphic expression of genes in gonads during early differentiation of a teleost fish, the Nile tilapia Oreochromis niloticus","volume":"78","author":"Ijiri","year":"2008","journal-title":"Biol. Reprod."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"149","DOI":"10.1006\/dbio.2000.0129","article-title":"Two sox9 genes on duplicated zebrafish chromosomes: Expression of similar transcription activators in distinct sites","volume":"231","author":"Chiang","year":"2001","journal-title":"Dev. Biol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"950","DOI":"10.1016\/j.bbrc.2005.03.066","article-title":"Gene structure, multiple alternative splicing, and expression in gonads of zebrafish Dmrt1","volume":"330","author":"Guo","year":"2005","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"167","DOI":"10.1016\/S0925-4773(01)00613-X","article-title":"Two isoforms of vasa homologs in a teleost fish: Their differential expression during germ cell differentiation","volume":"111","author":"Kobayashi","year":"2002","journal-title":"Mech. Dev."},{"key":"ref_42","doi-asserted-by":"crossref","unstructured":"Burge, S., Kelly, E., Lonsdale, D., Mutowo-Muellenet, P., McAnulla, C., Mitchell, A., Sangrador-Vegas, A., Yong, S.Y., Mulder, N., and Hunter, S. (2012). Manual GO annotation of predictive protein signatures: The InterPro approach to GO curation. Database J. Biol. Databases Curation, 2012.","DOI":"10.1093\/database\/bar068"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"319","DOI":"10.24272\/j.issn.2095-8137.2021.406","article-title":"Identification of global alternative splicing and sex-specific splicing via comparative transcriptome analysis of gonads of Chinese tongue sole (Cynoglossus semilaevis)","volume":"43","author":"Lu","year":"2022","journal-title":"Zool. Res."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"1427","DOI":"10.1093\/bioinformatics\/18.11.1427","article-title":"MethPrimer: Designing primers for methylation PCRs","volume":"18","author":"Li","year":"2002","journal-title":"Bioinformatics"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"402","DOI":"10.1006\/meth.2001.1262","article-title":"Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method","volume":"25","author":"Livak","year":"2001","journal-title":"Methods"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"4067","DOI":"10.1093\/bioinformatics\/bti652","article-title":"BiQ Analyzer: Visualization and quality control for DNA methylation data from bisulfite sequencing","volume":"21","author":"Bock","year":"2005","journal-title":"Bioinformatics"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"D265","DOI":"10.1093\/nar\/gkz991","article-title":"CDD\/SPARCLE: The conserved domain database in 2020","volume":"48","author":"Lu","year":"2020","journal-title":"Nucleic Acids Res."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"W327","DOI":"10.1093\/nar\/gkh454","article-title":"CD-Search: Protein domain annotations on the fly","volume":"32","author":"Bryant","year":"2004","journal-title":"Nucleic Acids Res."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"D225","DOI":"10.1093\/nar\/gkq1189","article-title":"CDD: A Conserved Domain Database for the functional annotation of proteins","volume":"39","author":"Lu","year":"2011","journal-title":"Nucleic Acids Res."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"1530","DOI":"10.1093\/molbev\/msaa015","article-title":"IQ-TREE 2: New Models and Efficient Methods for Phylogenetic Inference in the Genomic Era","volume":"37","author":"Minh","year":"2020","journal-title":"Mol. Biol. Evol."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"W276","DOI":"10.1093\/nar\/gkac240","article-title":"Search and sequence analysis tools services from EMBL-EBI in 2022","volume":"50","author":"Madeira","year":"2022","journal-title":"Nucleic Acids Res."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/25\/14\/7637\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T15:15:26Z","timestamp":1760109326000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/25\/14\/7637"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,7,11]]},"references-count":51,"journal-issue":{"issue":"14","published-online":{"date-parts":[[2024,7]]}},"alternative-id":["ijms25147637"],"URL":"https:\/\/doi.org\/10.3390\/ijms25147637","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2024,7,11]]}}}