| Autor: |
M S, Longtine, A, McKenzie, D J, Demarini, N G, Shah, A, Wach, A, Brachat, P, Philippsen, J R, Pringle |
| Rok vydání: |
1998 |
| Předmět: |
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| Zdroj: |
Yeast (Chichester, England). 14(10) |
| ISSN: |
0749-503X |
| Popis: |
An important recent advance in the functional analysis of Saccharomyces cerevisiae genes is the development of the one-step PCR-mediated technique for deletion and modification of chromosomal genes. This method allows very rapid gene manipulations without requiring plasmid clones of the gene of interest. We describe here a new set of plasmids that serve as templates for the PCR synthesis of fragments that allow a variety of gene modifications. Using as selectable marker the S. cerevisiae TRP1 gene or modules containing the heterologous Schizosaccharomyces pombe his5+ or Escherichia coli kan(r) gene, these plasmids allow gene deletion, gene overexpression (using the regulatable GAL1 promoter), C- or N-terminal protein tagging [with GFP(S65T), GST, or the 3HA or 13Myc epitope], and partial N- or C-terminal deletions (with or without concomitant protein tagging). Because of the modular nature of the plasmids, they allow efficient and economical use of a small number of PCR primers for a wide variety of gene manipulations. Thus, these plasmids should further facilitate the rapid analysis of gene function in S. cerevisiae. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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