A general method for polyethylene-glycol-induced genetic transformation of bacteria and yeast

Klebe, Robert J.; Harriss, June V.; Sharp, Zelton Dave; Douglas, Michael G.

doi:10.1016/0378-1119(83)90238-x

Cited by 449 publications

(213 citation statements)

References 23 publications

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“…E. coli cells were transformed by the CaC12 heat shock method (Maniatis et al 1982). For transformation of H. polymorpha, frozen cells (Klebe et al 1983) treated with polyethylene glycol were used as described earlier (Roggenkamp et al 1986) with minor modifications.…”

Section: Methodsmentioning

confidence: 99%

Targeting sequences of the two major peroxisomal proteins in the methylotrophic yeast Hansenula polymorpha

et al. 1992

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Section: Methodsmentioning

confidence: 99%

Targeting sequences of the two major peroxisomal proteins in the methylotrophic yeast Hansenula polymorpha

et al. 1992

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“…All 112::pAS17L(2x) Genetic manipulations, transformations and DNA preparations E. coli was transformed by the method of Hanahan (1985). Yeasts were transformed using the freeze (Klebe et al, 1983) or the lithium acetate method (Gietz et al, 1992). Chromosomal DNA for Southern analyses or as template for some PCR reactions was prepared from yeast according to Hoffman and Winston (1987).…”

Section: Media and Culture Conditionsmentioning

confidence: 99%

The effect of tea tree oil and antifungal agents on a reporter for yeast cell integrity signalling

Straede

Corran

Bundy

et al. 2007

Yeast

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Cell integrity in Saccharomyces cerevisiae is ensured by a rigid cell wall whose synthesis is controlled by a highly conserved MAP kinase signal transduction cascade. Stress at the cell surface is detected by a set of sensors and ultimately transmitted through this cascade to the transcription factor Rlm1, which governs expression of many genes encoding enzymes of cell wall biosynthesis. We here report on a number of versatile reporter constructs which link activation of a hybrid, Rlm1-lexA, by the MAP kinase Mpk1/Slt2 to the expression of the bacterial lacZ gene. This system was adapted to automated microwell screening and shown to be activated by a number of compounds inhibiting cell wall biosynthesis or interfering with plasma membrane function. In addition, we tested tea tree oil and two of its purified constituents (α-terpineol, terpinen-4-ol) for their effects on growth and on cell integrity signalling using such reporter strains. Tea tree oil was found to inhibit growth of wild-type and slg1/wsc1 mutant cells at a threshold of approximately 0.1% v/v, with the purified compounds acting already at half these concentrations. A mid2 deletion displayed hyper-resistance. Tea tree oil also induces the signalling pathway in a dose-dependent manner.

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“…The yeast strain Y190, which is Leu À and Trp À , and contains LacZ and HIS3 reporter genes for GAL4 transcriptional activity, was transformed with pGBT9-Apoptin according to the Klebe protocol. 27 In a bGal colony filter assay, 28 the bait plasmid was shown to have no transactivation activity on its own. Next, the cDNA library was transformed into Y190 containing pGBT9-Apoptin, with an efficiency of 1 Â 10 6 , and grown on plates lacking Leu, Trp and His.…”

Section: Methodsmentioning

confidence: 99%

Human death effector domain-associated factor interacts with the viral apoptosis agonist Apoptin and exerts tumor-preferential cell killing

Oorschot

Voskamp²,

Seelen³

et al. 2004

Cell Death Differ

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Apoptin, a protein from chicken anemia virus without an apparent cellular homologue, can induce apoptosis in mammalian cells. Its cytotoxicity is limited to transformed or tumor cells, making Apoptin a highly interesting candidate for cancer therapy. To elucidate Apoptin's mechanism of action, we have searched for binding partners in the human proteome. Here, we report that Apoptin interacts with DEDAF, a protein previously found to associate with death effector domain (DED)-containing pro-apoptotic proteins, and to be involved in regulation of transcription. Like Apoptin, after transient overexpression, DEDAF induced apoptosis in various human tumor cell lines, but not in primary fibroblasts or mesenchymal cells. DEDAF-induced cell death was inhibited by the caspase inhibitor p35. Together with the reported association of DEDAF with a DED-containing DNA-binding protein in the nucleus and the transcription regulatory activity, our findings may provide a clue for the mechanism of Apoptin's actions in mammalian cells.

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A general method for polyethylene-glycol-induced genetic transformation of bacteria and yeast

Cited by 449 publications

References 23 publications

Targeting sequences of the two major peroxisomal proteins in the methylotrophic yeast Hansenula polymorpha

Targeting sequences of the two major peroxisomal proteins in the methylotrophic yeast Hansenula polymorpha

The effect of tea tree oil and antifungal agents on a reporter for yeast cell integrity signalling

Human death effector domain-associated factor interacts with the viral apoptosis agonist Apoptin and exerts tumor-preferential cell killing

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