{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,12]],"date-time":"2025-10-12T02:59:49Z","timestamp":1760237989439,"version":"build-2065373602"},"reference-count":20,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2022,6,2]],"date-time":"2022-06-02T00:00:00Z","timestamp":1654128000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"National Genetically Modified Organisms Breeding Major Projects program","award":["2018ZX8010-08B","31671313"],"award-info":[{"award-number":["2018ZX8010-08B","31671313"]}]},{"DOI":"10.13039\/501100001809","name":"National Natural Science Foundation of China","doi-asserted-by":"publisher","award":["2018ZX8010-08B","31671313"],"award-info":[{"award-number":["2018ZX8010-08B","31671313"]}],"id":[{"id":"10.13039\/501100001809","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Priority Academic Program Development of Jiangsu Higher Education Institutions","award":["2018ZX8010-08B","31671313"],"award-info":[{"award-number":["2018ZX8010-08B","31671313"]}]},{"name":"High-end Talent Support Program of Yangzhou University","award":["2018ZX8010-08B","31671313"],"award-info":[{"award-number":["2018ZX8010-08B","31671313"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["CIMB"],"abstract":"<jats:p>An enhancer trap (ET) mediated by a transposon is an effective method for functional gene research. Here, an ET system based on a PB transposon that carries a mini Krt4 promoter (the keratin4 minimal promoter from zebrafish) and the green fluorescent protein gene (GFP) has been used to produce zebrafish ET lines. One enhancer trap line with eye-specific expression GFP named EYE was used to identify the trapped enhancers and genes. Firstly, GFP showed a temporal and spatial expression pattern with whole-embryo expression at 6, 12, and 24 hpf stages and eye-specific expression from 2 to 7 dpf. Then, the genome insertion sites were detected by splinkerette PCR (spPCR). The Krt4-GFP was inserted into the fourth intron of the gene itgav (integrin, alpha V) in chromosome 9 of the zebrafish genome, with the GFP direction the same as that of the itgav gene. By the alignment of homologous gene sequences in different species, three predicted endogenous enhancers were obtained. The trapped endogenous gene itgav, whose overexpression is related to hepatocellular carcinoma, showed a similar expression pattern as GFP detected by in situ hybridization, which suggested that GFP and itgav were possibly regulated by the same enhancers. In short, the zebrafish enhancer trap lines generated by the PB transposon-mediated enhancer trap technology in this study were valuable resources as visual markers to study the regulators and genes. This work provides an efficient method to identify and isolate tissue-specific enhancer sequences.<\/jats:p>","DOI":"10.3390\/cimb44060178","type":"journal-article","created":{"date-parts":[[2022,6,2]],"date-time":"2022-06-02T11:05:02Z","timestamp":1654167902000},"page":"2614-2621","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["The Annotation of Zebrafish Enhancer Trap Lines Generated with PB Transposon"],"prefix":"10.3390","volume":"44","author":[{"given":"Wenzhu","family":"Jia","sequence":"first","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Zhongxia","family":"Guan","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shasha","family":"Shi","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kuilin","family":"Xiang","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Peihong","family":"Chen","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Fen","family":"Tan","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Numan","family":"Ullah","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4044-4469","authenticated-orcid":false,"given":"Mohamed","family":"Diaby","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Mengke","family":"Guo","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0488-4718","authenticated-orcid":false,"given":"Chengyi","family":"Song","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4029-1258","authenticated-orcid":false,"given":"Bo","family":"Gao","sequence":"additional","affiliation":[{"name":"College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2022,6,2]]},"reference":[{"key":"ref_1","first-page":"589","article-title":"Advances in research methods for gene function","volume":"20","author":"Du","year":"2008","journal-title":"Chin. Bull. Life Sci."},{"key":"ref_2","doi-asserted-by":"crossref","unstructured":"Liu, C., Song, G., Mao, L., Long, Y., Li, Q., and Cui, Z. (2015). Generation of an Enhancer-Trapping Vector for Insertional Mutagenesis in Zebrafish. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0139612"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"1288","DOI":"10.1101\/gad.3.9.1288","article-title":"P-element-mediated enhancer detection: A versatile method to study development in Drosophila","volume":"3","author":"Bellen","year":"1989","journal-title":"Genes Dev."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1101","DOI":"10.1126\/science.aah6192","article-title":"Lentiviral vectors, two decades later","volume":"353","author":"Naldini","year":"2016","journal-title":"Science"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"932","DOI":"10.1074\/jbc.RA118.005568","article-title":"Tol2 Improved -mediated enhancer trap identifies weakly expressed genes during liver and \u03b2 cell development and regeneration in zebrafish","volume":"294","author":"Zhong","year":"2019","journal-title":"J. Biol. Chem."},{"key":"ref_6","first-page":"449","article-title":"Tol2 transposon-mediated enhancer trap to identify developmentally regulated zebrafish genes in vivo","volume":"231","author":"Parinov","year":"2004","journal-title":"Dev. Dyn. Off. Publ. Am. Assoc. Anat."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"57","DOI":"10.1016\/j.gene.2003.09.009","article-title":"Transposon-mediated enhancer trapping in medaka","volume":"322","author":"Grabher","year":"2003","journal-title":"Gene"},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Shen, D., Xue, S., Chan, S., Sang, Y., Wang, S., Wang, Y., Chen, C., Gao, B., Mueller, F., and Song, C. (2018). Enhancer Trapping and Annotation in Zebrafish Mediated with Sleeping Beauty, piggyBac and Tol2 Transposons. Genes, 9.","DOI":"10.3390\/genes9120630"},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Balciunas, D., Davidson, A., Sivasubbu, S., Hermanson, S.B., Welle, Z., and Ekker, S.C. (2008). Enhancer trapping in zebrafish using the Sleeping Beauty transposon. BMC Genom., 5.","DOI":"10.1186\/1471-2164-5-62"},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Potter, C., and Luo, L. (2010). Splinkerette PCR for mapping transposable elements in Drosophila. PLoS ONE, 5.","DOI":"10.1371\/journal.pone.0010168"},{"key":"ref_11","first-page":"449","article-title":"Enhancer trapping nearby rps26 gene in zebrafish mediated by the Tol2 transposon and it\u2019s annotation","volume":"34","author":"Sang","year":"2018","journal-title":"Sheng Wu Gong Cheng Xue Bao Chin. J. Biotechnol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"59","DOI":"10.1038\/nprot.2007.514","article-title":"High-resolution in situ hybridization to wholemount zebrafish embryos","volume":"3","author":"Thisse","year":"2008","journal-title":"Nat. Protoc."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"287","DOI":"10.1128\/jvi.47.2.287-300.1983","article-title":"Acquisition of Host Cell DNA Sequences by Baculoviruses: Relationship Between Host DNA Insertions and FP Mutants of Autographa californica and Galleria mellonella Nuclear Polyhedrosis Viruses","volume":"47","author":"Fraser","year":"1983","journal-title":"J. Virol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"e13503","DOI":"10.7554\/eLife.13503","article-title":"A Mammalian enhancer trap resource for discovering and manipulating neuronal cell types","volume":"5","author":"Shima","year":"2016","journal-title":"Elife"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"9290","DOI":"10.1073\/pnas.0801017105","article-title":"Chromosomal transposition of PiggyBac in mouse embryonic stem cells","volume":"105","author":"Wang","year":"2008","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"404","DOI":"10.1002\/dvg.20508","article-title":"Chromosomal mobilization and reintegration of Sleeping Beauty and PiggyBac transposons","volume":"47","author":"Liang","year":"2009","journal-title":"Genesis"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"1317","DOI":"10.1128\/MCB.00670-12","article-title":"The piggyBac Transposon Displays Local and Distant Reintegration Preferences and Can Cause Mutations at Noncanonical Integration Sites","volume":"33","author":"Li","year":"2013","journal-title":"Mol. Cell. Biol."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"434","DOI":"10.1111\/dgd.12055","article-title":"Enhancer and gene traps for molecular imaging and genetic analysis in zebrafish","volume":"55","author":"Trinh","year":"2013","journal-title":"Dev. Growth Differ."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"265","DOI":"10.1016\/j.cell.2008.03.024","article-title":"The 3D Structure of the Immunoglobulin HeavyChain Locus: Implications for LongRange Genomic Interactions","volume":"133","author":"Jhunjhunwala","year":"2008","journal-title":"Cell"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"299","DOI":"10.1007\/s11248-009-9312-x","article-title":"The role of post-transcriptional RNA processing and plasmid vector sequences on transient transgene expression in zebrafish","volume":"19","author":"Chatterjee","year":"2010","journal-title":"Transgenic Res."}],"container-title":["Current Issues in Molecular Biology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1467-3045\/44\/6\/178\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T23:24:04Z","timestamp":1760138644000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1467-3045\/44\/6\/178"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,6,2]]},"references-count":20,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2022,6]]}},"alternative-id":["cimb44060178"],"URL":"https:\/\/doi.org\/10.3390\/cimb44060178","relation":{},"ISSN":["1467-3045"],"issn-type":[{"type":"electronic","value":"1467-3045"}],"subject":[],"published":{"date-parts":[[2022,6,2]]}}}