Generation of an expression library in the baculovirus expression vector system

Vernon K. Ward, Sabine B. Kreissig, Bruce D. Hammock, Prabhakara V Choudary

Research output: Contribution to journalArticle

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Abstract

The construction and screening of a small cDNA library consisting of 2 × 104 clones in the baculovirus expression vector system are described. This library consists of antibody heavy chain sequences isolated from the spleen of a mouse immunized with tetanus toxoid fragment C. A portion of this library was used to produce a pool of recombinant baculoviruses which were screened for production of antibody fragments reactive to tetanus toxoid without prior expression in Escherichia coli. The pool of 30 clones was found to contain at least 6 different populations of antibody indicating that diversity existed within the library. Positive clones were isolated from the baculovirus system and confirmed as being capable of producing a tetanus reactive antibody by expression as a β-lactamase fusion protein in E. coli. One of these clones was returned to the baculovirus system using a different transfer vector, and tetanus binding reconfirmed. The results presented here show that the concept of the construction and screening of a baculovirus expression library is feasible even with "difficult" proteins, such as antibody heavy chain fragments, and that the baculovirus expression vector system has the potential to produce cDNA expression libraries which can be screened directly for the desired protein.

Original languageEnglish (US)
Pages (from-to)263-272
Number of pages10
JournalJournal of Virological Methods
Volume53
Issue number2-3
DOIs
StatePublished - 1995

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Keywords

  • Antibody expression
  • Baculovirus
  • Eukaryotic expression
  • Expression library

ASJC Scopus subject areas

  • Virology

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