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Results for “reverse genetic manipulation”

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PubMed1975

Selection of spontaneous mutants by inositol starvation in yeast.

A new method for the routine isolation of mutations of spontaneous origin in the yeast, Saccharomyces cerevisiae, is reported. The technique is based on the observation that inositol auxotrophs die when deprived of inositol. However, if macromolecular synthesis is inhibited, most of the cells survive. Appropriate manipulation of inositol requiring mutants can therefore result in the selective survival of cells possessing mutations which affect macromolecular synthesis. Since reversion to inositol prototrophy can be a major source of interference in efficient selection, a haploid double mutant strain has been constructed which reverts to inositol prototrophy with a frequency estimated to be several orders of magnitude lower than the expected frequency of single, spontaneous mutational events. Using this strain, enrichment in excess of 10,000 fold has been obtained for various classes of auxotrophic mutants. Spontaneous temperature sensitive mutants have also been obtained.

Genetic Techniques↗

PubMed2026

Plant U-box E3 ligases: Versatile regulators of environmental stress adaptation and ABA signaling.

Ubiquitination is a reversible post-translational modification that orchestrates a wide spectrum of fundamental processes throughout the plant life cycle. Executed by a hierarchical E1-E2-E3 cascades, this modification tags targets with ubiquitin to modulate their turnover, activity, or subcellular compartmentalization. Among the diverse E3 ligase families, plant U-box (PUB) proteins stand out as a prominent class that determines substrate selection and has emerged as a focal point of stress biology. In this review, we first delineate the structural features of PUB proteins, highlighting their conserved domains and associated regulatory motifs. We then systematically dissect their multifaceted functions in abiotic stress adaptation, encompassing drought, salinity, extreme temperatures, oxidative stress, heavy metal toxicity, with particular emphasis on their integration with ABA signaling networks. We further outline critical knowledge gaps and propose future strategies to decode the regulatory architecture of PUBs. Collectively, this review provides a theoretical foundation and new insights for facilitating the genetic improvement of crop resilience in the face of continuously intensifying environmental stresses through the manipulation of PUB-mediated ubiquitination networks.

ABA signaling↗