Abstract：
The baculovirus expression vector system (BEVS) has now found acceptance in both research laboratories and industry, which can be attributed to many of its key features including the limited host range of the vectors, their non-pathogenicity to humans, and the mammalian-like post-translational modification (PTMs) that can be achieved in insect cells. In fact, this system acts as a middle ground between prokaryotes and higher eukaryotes to produce complex biologics. Still, industrial use of the BEVS lags compared to other platforms. We have postulated that one reason for this has been a lack of genetic tools that can complement the study of baculovirus vectors, while a second reason is the co-production of the baculovirus vector with the desired product. While some genetic enhancements have been made to improve the BEVS as a production platform, the genome remains under-scrutinized. This chapter outlines the methodology for a CRISPR-Cas9-based transfection-infection assay to probe the baculovirus genome for essential/nonessential genes that can potentially maximize foreign gene expression under a promoter of choice.
摘要：
杆状病毒表达载体系统（BEVS）现已在研究实验室和行业中得到认可，这可以归因于它的许多关键特征，包括载体的有限宿主范围，它们对人类的非致病性，以及可以在昆虫细胞中实现的哺乳动物样翻译后修饰(PTM)。事实上,该系统充当原核生物和高等真核生物之间的中间地带，以产生复杂的生物制品。尽管如此，与其他平台相比，BEVS的工业使用滞后。我们推测，造成这种情况的原因之一是缺乏可以补充杆状病毒载体研究的遗传工具，而第二个原因是杆状病毒载体与所需产物的共同生产。虽然已经进行了一些遗传增强以改善BEVS作为生产平台，基因组仍未得到严格审查。本章概述了基于CRISPR-Cas9的转染感染测定的方法，以探测杆状病毒基因组中必需/非必需基因，这些基因可以在选择的启动子下潜在地最大化外源基因表达。
