The generation and characterization of a transgenic zebrafish line with lens-specific Cre expression.

IF 1.8 3区医学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY

Molecular Vision Pub Date : 2024-03-19 eCollection Date: 2024-01-01

Xuyan Peng, Xiaolin Jia, Guohui Shang, Mengjiao Xue, Mingjun Jiang, Dandan Chen, Fengyan Zhang, Yanzhong Hu

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引用次数: 0

Abstract

Purpose: Danio rerio zebrafish constitute a popular model for studying lens development and congenital cataracts. However, the specific deletion of a gene with a Cre/LoxP system in the zebrafish lens is unavailable because of the lack of a lens-Cre-transgenic zebrafish. This study aimed to generate a transgenic zebrafish line in which Cre recombinase was specifically expressed in the lens.

Methods: The pTol2 cryaa:Cre-polyA-cryaa:EGFP (enhanced green fluorescent protein) plasmid was constructed and co-injected with Tol2-transposase into one-to-two-cell-stage wild-type (WT) zebrafish embryos. Whole-mount in situ hybridization (ISH), tissue section, hematoxylin and eosin staining, a Western blot, a split-lamp observation, and a grid transmission assay were used to analyze the Cre expression, lens structure, and lens transparency of the transgenic zebrafish.

Results: In this study, we generated a transgenic zebrafish line, zTg(cryaa:Cre-cryaa:EGFP), in which Cre recombinase and EGFP were driven by the lens-specific cryaa promoter. zTg(cryaa:Cre-cryaa:EGFP) began to express Cre and EGFP specifically in the lens at the 22 hpf stage, and this ectopic Cre could efficiently and specifically delete the red fluorescent protein (RFP) signal from the lens when zTg(cryaa:Cre-cryaa:EGFP) embryos were injected with the loxP-flanked RFP plasmid. The overexpression of Cre and EGFP did not impair zebrafish development or lens transparency. Accordingly, this zTg(cryaa:Cre-cryaa:EGFP) zebrafish line is a useful tool for gene editing, specifically with zebrafish lenses.

Conclusions: We established a zTg(cryaa:Cre-cryaa:EGFP) zebrafish line that can specifically express an active Cre recombinase in lens tissues. This transgenic zebrafish line can be used as a tool to specifically manipulate a gene in zebrafish lenses.

本刊更多论文

具有晶状体特异性 Cre 表达的转基因斑马鱼品系的产生和特征。

目的：斑马鱼是研究晶状体发育和先天性白内障的常用模型。然而，由于缺乏晶状体-Cre 转基因斑马鱼，因此无法利用 Cre/LoxP 系统在斑马鱼晶状体中特异性地删除一个基因。本研究旨在产生一种在晶状体中特异表达 Cre 重组酶的转基因斑马鱼品系：方法：构建了 pTol2 cryaa:Cre-polyA-cryaa:EGFP（增强型绿色荧光蛋白）质粒，并将其与 Tol2-转座酶共同注射到一至二细胞期的野生型（WT）斑马鱼胚胎中。通过原位杂交（ISH）、组织切片、苏木精和伊红染色、Western印迹、分光灯观察和网格透射试验，分析了转基因斑马鱼的Cre表达、晶状体结构和晶状体透明度：在这项研究中，我们产生了一个转基因斑马鱼品系zTg（cryaa:Cre-cryaa:EGFP），其中Cre重组酶和EGFP由晶状体特异性cryaa启动子驱动。当向zTg(cryaa:Cre-cryaa:EGFP)胚胎注射loxP-fanked RFP质粒时，这种异位的Cre能高效、特异地删除晶状体中的红色荧光蛋白（RFP）信号。过表达 Cre 和 EGFP 不会影响斑马鱼的发育或晶状体的透明度。因此，这种zTg(cryaa:Cre-cryaa:EGFP)斑马鱼品系是一种有用的基因编辑工具，特别是在斑马鱼晶状体方面：我们建立了一个能在晶状体组织中特异表达活性 Cre 重组酶的 zTg（cryaa:Cre-cryaa:EGFP）斑马鱼品系。该转基因斑马鱼品系可用作特异性操纵斑马鱼晶状体基因的工具。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Molecular Vision 生物-生化与分子生物学

CiteScore

4.40

自引率

0.00%

发文量

审稿时长

1 months

期刊介绍： Molecular Vision is a peer-reviewed journal dedicated to the dissemination of research results in molecular biology, cell biology, and the genetics of the visual system (ocular and cortical). Molecular Vision publishes articles presenting original research that has not previously been published and comprehensive articles reviewing the current status of a particular field or topic. Submissions to Molecular Vision are subjected to rigorous peer review. Molecular Vision does NOT publish preprints. For authors, Molecular Vision provides a rapid means of communicating important results. Access to Molecular Vision is free and unrestricted, allowing the widest possible audience for your article. Digital publishing allows you to use color images freely (and without fees). Additionally, you may publish animations, sounds, or other supplementary information that clarifies or supports your article. Each of the authors of an article may also list an electronic mail address (which will be updated upon request) to give interested readers easy access to authors.