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在没有UTR序列信息的情况下,工程人类/猿类轮状病毒VP7重组剂

Roman Valusenko-Mehrkens¹, Reimar Johne¹, Alexander Falkenhagen²

¹Department of Biological Safety, German Federal Institute for Risk Assessment, Max-Dohrn-Str. 8-10, 10589, Berlin, Germany.

Applied microbiology and biotechnology

|February 27, 2025

概括

生成轮状病毒A (RVA) 重组物具有挑战性,因为基因组序列不完整. 这项研究表明,猿类RVA未翻译区域 (UTR) 可以替代缺失的人类RVA UTR,从而使多样化的疫苗菌株开发成为可能.

科学领域:

病毒学病毒学
分子生物学分子生物学
疫苗学疫苗学疫苗学

背景情况:

基于等离子体的反转基因系统用于罗塔病毒A (RVA) 促进与人类RVA抗原重组病毒的产生.
一个显著的局限性是经常缺少来自野外菌株的完整基因组段序列,特别是未翻译区域 (UTR).
这就需要替代策略来产生RVA复合剂,用于疫苗开发.

研究的目的:

建立一个系统,只使用开放式读取框架 (ORF) 核酸序列信息来生成RVA重组剂.
评估替换缺失的人类RVA UTRs的SA11 UTRs的可行性.
扩大抗原谱,用于设计新型RVA疫苗菌株.

主要方法:

构建含有6个人类RVA野外菌株 (G2,G5,G8,G9,G12,G29) 的VP7-ORF核酸序列的等离子体.
人类RVA ORF序列与来自猿类RVA菌株SA11的UTR序列的并排.
使用这些工程塑生成VP7重组剂,并随后对生存能力和遗传完整性进行分析.

主要成果:

在SA11骨干中成功生成了四个VP7复合物 (G2,G5,G9,G12).
无法产生可行的G8和G29重组剂,与异常的氨基酸替代有关,可能是由测序错误或内在限制引起的.
证明在G8中对特定突变 (C656T) 的位点定向逆转挽救了复杂病毒的生存能力,而在G29 (G84C) 中的逆转却没有.

结论:

大多数人类RVA VP7 UTRs可以有效地被猿RVA菌株SA11的RVA菌株所取代.
潜在的测序错误或内在的重组限制可能会阻碍生成可行的RVA重组剂.
开发的系统为快速设计多种RVA重组剂提供了有价值的工具,可能加速新型RVA疫苗菌株的开发.

关键词:

进行重新分类.反向遗传学是一种反向遗传学.罗塔病毒A型病毒没有翻译的地区,点突变.在VP7中,我们可以使用VP7.

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