| Advanced Design of Minimalistic Dumbbell-shaped Gene Expression Vectors
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Author:
Date:
2017-08-05
[Abstract] Minimal DNA vectors exclusively comprising therapeutically relevant sequences hold great promise for the development of novel therapeutic regimen. Dumbbell-shaped vectors represent non-viral non-integrating DNA minimal vectors which have entered an advanced stage of clinical development (Hardee et al., 2017). Spliceable introns and DNA nuclear import signals such as SV40 enhancer sequences are molecular features that have found multiple applications in plasmid vectors to improve transgene expression. In dumbbells however, effects triggered by introns were not investigated and DNA-based nuclear import sequences have not found applications yet, presumably because dumbbell vectors have continuously been minimized with regard to size. We investigated the effects of an intron and/or ...
[摘要] 唯一包含治疗相关序列的最小DNA载体对于新型治疗方案的发展具有很大的希望。哑铃型载体代表已经进入临床发展的晚期阶段的非病毒非整合DNA最小载体（Hardee等人，2017）。可接合的内含子和DNA核输入信号如SV40增强子序列是在质粒载体中发现多个应用以改善转基因表达的分子特征。然而，在哑铃中，由内含子引发的效应未被研究，基于DNA的核导入序列尚未发现应用，可能是因为哑铃载体在尺寸上不断被最小化。我们调查内含子和/或SV40增强子衍生序列对哑铃载体驱动的报告基因表达的影响。发现可拼接内含子的实现在所有研究的细胞系中无条件地增强基因表达。相反，SV40增强子的使用改善了细胞类型依赖性的基因表达。虽然这两个特征显着增加哑铃载体大小，但内含子和增强子或两者的组合都不会对基因表达产生负面影响。相反，与质粒或对照哑铃相比，这两个特征在一起改善了哑铃驱动的基因表达，高达160-或56倍。因此，强烈建议考虑用于哑铃矢量设计的内含子和SV40增强子。这种先进的设计可以促进哑铃型DNA载体的临床前和临床应用。
【背景】虽然许多基因已经使用哑铃形DNA载体表达，但大多数这些应用使用包括启动子，编码序列（CDS）和转录终止子的基本设计。一些载体包括嵌合内含子，然而，没有报道通过这种设计增强了转基因表达（Schirmbeck等人，2001）。在这里，我们研究了分子特征引发的效应，这些特征经常在哑铃驱动基因表达的质粒设计中得到应用：1.已知来自人β-珠蛋白基因剪接的嵌合内含子促进RNA加工，核出口，随后基因表达（Luo ...
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| Mouse Subependymal Zone Explants Cultured on Primary Astrocytes
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Author:
Date:
2016-07-20
[Abstract] Neuroblast migration is a major component in the process of neuronal circuit assembly. In the rodent postnatal brain, the subependymal zone (SEZ) is the largest neurogenic niche where new neurons are born throughout life. These cells migrate several millimeters following a specific pathway called the rostral migratory stream (RMS) towards the olfactory bulb. Once they reach their final destination, they differentiate mainly as interneurons and integrate into already established neuronal circuits. Along the RMS, neuroblasts use a particular mode of migration known as chain migration. They stay attached to each other while migrating and are intimately associated with adjacent astrocytic processes. To dissect the molecular and cellular signals involved in neuroblast migration, we developed ...
[摘要] 神经母细胞迁移是神经元电路组装过程中的主要组成部分。 在啮齿动物出生后的大脑中，子宫颈区（SEZ）是新生神经元生命中最大的神经源性生态位。 这些细胞沿着称为传播流（RMS）的特定途径朝向嗅球迁移几毫米。 一旦达到其最终目的地，它们主要区分为中间神经元并整合到已建立的神经元回路中。 沿着RMS，神经母细胞使用称为链转移的特定迁移模式。 它们在迁移时保持相互依赖，并与相邻的星形细胞过程密切相关。 为了解剖神经母细胞迁移中涉及的分子和细胞信号，我们开发了一种体外系统，其由在星形胶质细胞单层顶部共培养的经SEZ衍生的外植体组成。
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