Cloning of the LEU2 gene of Saccharomyces cerevisiae by in vivo recombination.
We describe a convenient method for the in vivo construction of large plasmids that possess a multitude of restriction sites. A large (23 kbases) circular self-replicating plasmid carrying a partial LEU2-d gene was cotransformed with a circular non-replicating plasmid carrying the entire LEU2 gene. In vivo recombination results preferentially in a plasmid that carries both the LEU2-d and the entire LEU2 gene. In addition we also found one plasmid with a tandem LEU2 insertion and one plasmid where the LEU2-d gene was replaced by the entire LEU2 gene.
Valinger
R
R
Mikrobiologisches Institut, Eidgenössische Technische Hochschule, Zürich, Switzerland.
Braus
G
G
Niederberger
P
P
Künzler
M
M
Paravicini
G
G
Schmidheini
T
T
Hütter
R
R
D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D005800:Genes, Fungal; D007930:Leucine / Q000096:biosynthesis; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D011995:Recombination, Genetic; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D007930:Leucine