title: Improvements to the throughput of recombinant protein expression in the baculovirus/insect cell system.
abstract: Recombinant baculoviruses have proved to be a very useful means to express many proteins over the last 20 years. Since their introduction, there have been a number of significant improvements that have simplified and speeded up the construction of baculoviruses. One of the most commonly used methods relies upon recombination with the baculovirus genome maintained in Escherichia coli. In this paper, we report the conversion of nearly all the steps in this process including the expression testing and purification to a multi-well plate format. This enables a significant increase in the number of constructs that can be processed in a shorter period of time and an order of magnitude increase in the number of expression conditions that can be analysed. A key step in our process is that the transfection is done in suspension rather than adherent cells, which gives a much higher virus titre than in the standard methods.
authors.affiliation: Global Protein Science and Supply, Global Sciences and Information, AstraZeneca at Mereside, Alderley Park, Macclesfield, Cheshire, UK.
authors.forename: Eileen J
authors.initials: EJ
authors.lastname: McCall
authors.forename: Anette
authors.initials: A
authors.lastname: Danielsson
authors.forename: Ian M
authors.initials: IM
authors.lastname: Hardern
authors.forename: Christine
authors.initials: C
authors.lastname: Dartsch
authors.forename: Ryan
authors.initials: R
authors.lastname: Hicks
authors.forename: Johanna M
authors.initials: JM
authors.lastname: Wahlberg
authors.forename: W Mark
authors.initials: WM
authors.lastname: Abbott
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line; D049109:Cell Proliferation; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics; D006639:Histidine / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D015203:Reproducibility of Results; D018411:Spodoptera; D014162:Transfection / Q000379:methods
chemical_list: D011994:Recombinant Proteins; C033223:polyhistidine; D006639:Histidine