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Virion Display: A High-Throughput Method to Express Functional Membrane Proteins.

Current protocols in molecular biology (2020-09-24)
Guan-Da Syu, Eric Johansen, Heng Zhu
ABSTRACT

Transmembrane proteins are responsible for many critical cellular functions and represent one of the largest families of drug targets. However, these proteins, especially multipass transmembrane proteins, are difficult to study because they must be embedded in a lipid bilayer to maintain their native conformations. The development of the virion display (VirD) technology enables transmembrane proteins to be integrated into the viral envelope of herpes simplex virus 1 (HSV-1). Combining high-throughput cloning, expression, and purification techniques, VirD technology has been applied to the largest set of human transmembrane proteins, namely G-protein-coupled receptors, and has allowed the identification of interactions that are both specific and functional. This article describes the procedures to integrate an open reading frame for any transmembrane protein into the HSV-1 genome and produce recombinant HSV-1 virus to ultimately generate pure VirD virions for biological and pharmaceutical studies. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Gateway cloning of transmembrane proteins Support Protocol 1: Ethanol precipitation of bacterial artificial chromosomal DNA Support Protocol 2: Preparation of competent cells Basic Protocol 2: Production of recombinant HSV-1 virions.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
牛血清白蛋白 来源于牛血清, lyophilized powder, ≥96% (agarose gel electrophoresis)
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总蛋白提取细胞裂解缓冲液
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蛋白酶抑制剂混合物粉末, for general use, lyophilized powder