Transformation of intact yeast cells treated with alkali cations.
TLDR
The transformation efficiency with Cs+ or Li+ was comparable with that of conventional protoplast methods for a plasmid containing ars1, although not for plasmids containing a 2 microns origin replication.Abstract:
Intact yeast cells treated with alkali cations took up plasmid DNA. Li+, Cs+, Rb+, K+, and Na+ were effective in inducing competence. Conditions for the transformation of Saccharomyces cerevisiae D13-1A with plasmid YRp7 were studied in detail with CsCl. The optimum incubation time was 1 h, and the optimum cell concentration was 5 x 10(7) cells per ml. The optimum concentration of Cs+ was 1.0 M. Transformation efficiency increased with increasing concentrations of plasmid DNA. Polyethylene glycol was absolutely required. Heat pulse and various polyamines or basic proteins stimulated the uptake of plasmid DNA. Besides circular DNA, linear plasmid DNA was also taken up by Cs+-treated yeast cells, although the uptake efficiency was considerably reduced. The transformation efficiency with Cs+ or Li+ was comparable with that of conventional protoplast methods for a plasmid containing ars1, although not for plasmids containing a 2 microns origin replication. Imagesread more
Citations
More filters
Journal ArticleDOI
A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae.
Robert S. Sikorski,Philip Hieter +1 more
TL;DR: A series of yeast shuttle vectors and host strains has been created to allow more efficient manipulation of DNA in Saccharomyces cerevisiae to perform most standard DNA manipulations in the same plasmid that is introduced into yeast.
Journal Article
A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae.
Journal ArticleDOI
A comprehensive analysis of protein–protein interactions in Saccharomyces cerevisiae
Peter Uetz,Loic Giot,Gerard Cagney,Traci A. Mansfield,Richard S. Judson,James R. Knight,Daniel Lockshon,Vaibhav A. Narayan,Maithreyan Srinivasan,Pascale Pochart,Alia Qureshi-Emili,Ying Li,Brian C. Godwin,Diana Conover,Theodore S. Kalbfleisch,Govindan Vijayadamodar,Meijia Yang,Mark Johnston,Stanley Fields,Jonathan M. Rothberg +19 more
TL;DR: Examination of large-scale yeast two-hybrid screens reveals interactions that place functionally unclassified proteins in a biological context, interactions between proteins involved in the same biological function, and interactions that link biological functions together into larger cellular processes.
Journal ArticleDOI
New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites.
R D Gietz,A Sugino +1 more
TL;DR: The production of new alleles of the LEU2, URA3 and TRP1 genes of Saccharomyces cerevisiae by in vitro mutagenesis is described and a unique series of yeast-Escherichia coli shuttle vectors derived from the plasmid pUC19 are constructed.
Journal ArticleDOI
New heterologous modules for classical or PCR‐based gene disruptions in Saccharomyces cerevisiae
TL;DR: A dominant resistance module, for selection of S. cerevisiae transformants, which entirely consists of heterologous DNA is constructed and tested, and some kanMX modules are flanked by 470 bp direct repeats, promoting in vivo excision with frequencies of 10–3–10–4.
References
More filters
Journal ArticleDOI
Calcium-dependent bacteriophage DNA infection.
Manley Mandel,A. Higa +1 more
TL;DR: Escherichia coli cells of strain K12 and C can be made competent to take up temperate phage DNA without the use of “helper phage”, and is effective for both linear and circular DNA molecules.
Journal ArticleDOI
Transformation of yeast
TL;DR: This work has used recently developed hybridization and restriction endonuclease mapping techniques to demonstrate directly the presence of the transforming DNA in the yeast genome and also to determine the arrangement of the sequences that were introduced.
Journal ArticleDOI
High-frequency transformation of yeast: autonomous replication of hybrid DNA molecules
TL;DR: A set of vector DNAs (Y vectors) useful for the cloning of DNA fragments in Saccharomyces cerevisiae (yeast) and in Escherichia coli are characterized in this paper.
Journal ArticleDOI
Simple agarose gel electrophoretic method for the identification and characterization of plasmid deoxyribonucleic acid.
TL;DR: Agarose gel electrophoresis may be employed effectively for the detection and preliminary characterization of plasmid deoxyribonucleic acid (DNA) present in clinical isolates and laboratory strains of gram-negative microorganisms.
Journal ArticleDOI
Analysis of endonuclease R-EcoRI fragments of DNA from lambdoid bacteriophages and other viruses by agarose-gel electrophoresis.
Robert B. Helling,Robert B. Helling,Howard M. Goodman,Howard M. Goodman,Herbert W. Boyer,Herbert W. Boyer +5 more
TL;DR: By means of agarose-gel electrophoresis, endonuclease R.EcoRI-generated fragments of DNA from various viruses were separated, their molecular weights were determined, and complete or partial fragment maps for lambda, phi80, and hybrid phages were constructed.