Strategies to optimize a metabolic pathway often involve building a large collection of strains, each containing different versions of sequences that regulate the expression of pathway genes. Here we develop reagents and methods to carry out this process at high efficiency in the yeast Saccharomyces cerevisiae. We identify variants of the E. coli tet operator (tetO) sequence that bind a TetR-VP16 activator with differential affinity, and therefore different TetR-VP16 activator driven expression. By recombining these variants upstream of the genes of a pathway, we generate unique combinations of expression levels. Here, we built a tetO toolkit, which includes: the I-OnuI homing endonuclease to create double-strand breaks, which increases homologous recombination by 105; a plasmid carrying six variant tetO sequences flanked by I-OnuI sites, uncoupling transformation and recombination steps; an S. cerevisiae-optimized TetR-VP16 activator; and a vector to integrate constructs into the yeast genome. We introduce into the S. cerevisiae genome the three crt genes from Erwinia herbicola required for yeast to synthesize lycopene, and carry out the recombination process to produce a population of cells with permutations of tetO variants regulating the three genes. We identify 0.7% of this population as making detectable lycopene, of which the vast majority have undergone recombination at all three crt genes. We estimate a rate of ∼20% recombination per targeted site, much higher than obtained in other studies. Application of this toolkit to medically or industrially important endproducts could reduce the time and labor required to optimize the expression of a set of metabolic genes.
Purpose -The purpose of this paper is to assert that libraries should actively seek and request alternatives to purchasing older ebook content rather than buying large ebook backlists. Currently publishers are offering the equivalent of the "big deal" with monographs through ebook backlists, and this is not a model that can or should be sustained by libraries. Design/methodology/approach -This paper discusses experience the author gained through evaluating major ebook backlist promotional offers from 2012 and 2013, reviews library literature concerning ebook use, and reviews current purchase methods such as patron-driven acquisition and firm ordering as alternatives to purchasing large ebook backlist packages. Findings -There does not appear to be agreement in library literature about use and preference for ebooks, but there does appear to be a consensus that users expect the availability of ebook content. However, that expectation is not reason enough to purchase older content in large quantities in a format that still has problematic limitations. Patron-driven acquisition and targeted firm ordering allow libraries to buy older content in ebook format with more information about the need for that content. Also, publishers should become more involved in offering alternatives to ebook backlist purchase models. Originality/value -This discussion sheds light on the fact that while the ebook format may be new, the content and the purchase models often are not. Libraries need to advocate for effective purchase models now rather than becoming dependent on publishers to provide content libraries already own.
Purpose – This paper aims to examine demand-driven acquisition (DDA) models that require an initial or minimum investment through the University of Wyoming’s experience with Elsevier’s Evidence-Based Selection model. Design/methodology/approach – In an attempt to avoid title-by-title selection and a desire to explore an alternative to all or nothing e-book packages, the University of Wyoming Libraries (UWL) participated in Elsevier’s Evidence-Based Selection purchase model for 2011 and 2012 e-book content in the 2013 calendar year. After an initial investment, the library was given access to the content. At the end of one year, UWL was provided with use data for the content and could choose an amount of content to retain up to Elsevier’s established “access fee”. Findings – Many studies have shown that print monographs in academic libraries do not circulate in high volumes. The use data for the titles included in the Evidence-Based Selection model was congruous with studies of print monograph circulation. Through a review of the literature and an account of the UWL’ experience with Elsevier’s Evidence-Based Selection model, this paper advocates for libraries to exercise caution when considering a DDA model requiring an initial investment. Originality/value – DDA is a purchase model that is becoming immensely popular, and in some libraries, the primary mode of acquisition. The value of this paper lies in the examination of a DDA model of a major academic publisher and the account of one library’s experience with that model.
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