Molecular biology tools for the study and therapy of PDE6β mutations
Molecular biology tools for the study and therapy of PDE6β mutations
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DOI：
10.1016/j.jbiotec.2018.07.030
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发表时间：
2018-10-20
影响因子：
4.1
通讯作者：
Silva, Gabriela A.
中科院分区：
文献类型：
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作者：
Bitoque, Diogo B.;Silva, Gabriela A.
基因疗法具有治疗视网膜疾病的潜力，我们一直在为此目的开发递送载体和表达载体。在这篇简短的交流中，我们描述了用于疾病机制的体外研究和治疗方法的体内测试的工具的产生。我们克隆了PDE6 β基因，并使用优化的质粒载体在rd10小鼠中重现了相同的突变。为了实现视觉检测，我们还通过定点诱变生成了表达正常和突变PDE6 β基因与GFP基因融合的质粒。接下来，我们用不同的载体转染视网膜色素上皮细胞，检测正常和突变PDE6 β的蛋白表达。通过这项工作，我们已经创建了基因治疗工具，用于体外和体内研究视网膜致病突变，即与视网膜色素变性有关的PDE6 β。
Gene therapy has the potential for treating retinal diseases, and we have been developing delivery vehicles and expression vectors for this purpose. In this short communication, we describe the generation of tools for both in vitro studies of the disease mechanism and for in vivo testing of therapeutic approaches. We have cloned the PDE6 beta gene and also recreated the same mutation present in the rd10 mouse using an optimized plasmid vector. To allow visual detection, we have also generated, through site-directed mutagenesis, plasmids expressing the normal and mutated PDE6 beta gene fused with the GFP gene. Next, we have transfected retinal pigment epithelium cells with the different vectors and detected the protein expression of both the normal and mutated PDE6 beta. With this work we have created gene therapy tools for in vitro and in vivo studies of retinal disease-causing mutations, namely for the PDE6 beta, implicated in retinitis pigmentosa.