Generation of Baculovirus Vectors for the High-Throughput Production of Proteins in Insect Cells

被引：46

作者：

Possee, Robert D. ^{[2
]}

Hitchman, Richard B. ^{[1
]}

Richards, Kevin S. ^{[1
]}

Mann, Susan G. ^{[1
]}

Siaterli, Evangelia ^{[1
]}

Nixon, Clare P. ^{[1
]}

Irving, Helen ^{[1
]}

Assenberg, Rene ^{[3
]}

Alderton, David ^{[3
]}

Owens, Raymond J. ^{[1
]}

King, Linda A. ^{[1
]}

机构：

[1] Oxford Brookes Univ, Sch Life Sci, Oxford OX3 0BP, England

[2] Ctr Hydrol & Ecol CEH Oxford, NERC, Oxford, England

[3] Wellcome Trust Ctr Human Genet, Oxford Prot Prod Facil, Oxford OX3 7BN, England

来源：

BIOTECHNOLOGY AND BIOENGINEERING | 2008年 / 101卷 / 06期

基金：

英国生物技术与生命科学研究理事会; 英国医学研究理事会;

关键词：

baculovirus expression system; high throughput; automated; bacmid; robotic liquid handler; secreted protein;

D O I：

10.1002/bit.22002

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

The baculovirus expression system is one of the most popular methods used for the production of recombinant proteins but has several complex steps which have proved inherently difficult to adapt to a multi-parallel process. We have developed a bacmid vector that does not require any form of selection pressure to seperate recombinant virus from non-recombinant parental virus. The method relies on homologous recombination in insect cells between a transfer vector containing a gene to be expressed and a replication-deficient bacmid. The target gene replaces a bacterial replicon at the polyhedrin loci, simultaneously restoring a virus gene essential for replication. Therefore, only recombinant virus can replicate facilitating the rapid production of multiple recombinant viruses on automated platforms in a one-step procedure. Using this vector allowed us to automate the generation of multiple recombinant viruses with a robotic liquid handler and then rapidly screen infected insect cell supernatant for the presence of secreted proteins.

引用

页码：1115 / 1122

页数：8

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