Two types of transgenic lines for doxycycline-inducible, cell-specific gene expression in zebrafish ultraviolet cone photoreceptors

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• 作者：Megan C. West^a ; ^{mcwest@bio.umass.edu" class="auth_mail} ; Leah J. Campbell^a ; ^{leahc@bio.umass.edu" class="auth_mail} ; John J. Willoughby^a ; ^{johnw@bio.umass.edu" class="auth_mail} ; Abbie M. Jensen^a ; ^b ; ^{ajensen@bio.umass.edu" class="auth_mail}
• 关键词：Conditional expression ; Transgenic zebrafish ; Cone photoreceptor
• 刊名：Gene Expression Patterns
• 出版年：March, 2014
• 年：2014
• 卷：14
• 期：2
• 页码：96-104
• 全文大小：2622 K

文摘

Temporal and spatial control of gene expression is important for studying the molecular and cellular mechanisms of development, physiology, and disease. We used the doxycycline (Dox)-inducible, Tet-On system to develop transgenic zebrafish for inducible, cell specific control of gene expression in the ultraviolet (UV) cone photoreceptors. Two constructs containing the reverse tetracycline-controlled transcriptional transactivator (rtTA) gene driven by the UV opsin-specific promoter (opn1sw1) were used to generate stable transgenic zebrafish lines using the Tol2-based transgenesis method. One construct included a self-reporting GFP (opn1sw1:rtTA, TRE:GFP) and the other incorporated an epitope tag on the rtTA protein (opn1sw1:rtTA^flag). UV cone-specific expression of TRE-controlled transgenes was induced by Dox treatment in larvae and adults. Induction of gene expression was observed in 96% of all larval UV cones within 16 h of Dox treatment. UV cone-specific expression of two genes from a bidirectional TRE construct injected into one-cell Tg(opn1sw1:rtTA^flag) embryos were also induced by Dox treatment. In addition, UV cone-specific expression of Crb2a^IntraWT was induced by Dox treatment in progeny from crosses of the TRE-response transgenic line, Tg(TRE:HA-Crb2a^IntraWT), to the Tg(opn1sw1:rtTA, TRE:GFP) line and the Tg(opn1sw1:rtTA^flag) line. These lines can be used in addition to the inducible, rod-specific gene expression system from the Tet-On Toolkit to elucidate the photoreceptor-specific effects of genes of interest in photoreceptor cell biology and retinal disease.