Amplification of the DFR1 gene in Saccaromyces cerevisiae

A new system was employed to study amplification of t,he DHF'R gene DFB,1 ) in Sa<,:;charoillYCB§. .Q~~Yi...S!i<;1~. . This system consists of a series of yeast strains containing a casset,te which encodes t he yeast, D..ERl gene ttghtly linked tjO a f usion of the yeast 1EU2. regulat,ory region wi tJ1 the LAQZ str ctural gene from E. cO.1-1 (,) . M. Clement , unpubl i,::;hed) . Th's casset;t e was shown t.o be integrat,ed int o a unj que chromosomal l ocati on in each strain . Yeast cells were se l ected for MTX-resistance and overproduction of ~ galac t osi d se ( B-gal ). Since the inserted DF'Rl and ~ACZ genes are independently regulated, it was thought that cel l s with this phenotype probably contain e d ampl if ications of the cassette. A lar ge variat ion in the f requn y o f MTX-resistance was found between the di ff e r ent str ains. These freqlen c ~ es r anged from about 2 x 10 - 7 fo r a population of cells containing the cassette integrated at, the BI J2.l gene in t,he middle of the long arm of chromosome V, to about 5 x 10-4 for a strain with the cassette i nserted in the r DNA cluster Abo It 85% of the MTX- res i stcmt iso l ates examined showed enhanced B·-gal act i v ity rel a t ive t o the parental strain . For the ma jorit y of strains, the mean B- gal activity in drug-r sistant clones was about 3 times that o f the parent following a single se l ect i on step . I n con t r ast, primary MTX-resistant derivat~ves of cells with the cassette inserted 3 at the rDNA cluster showed inc r eases in B- gal activity ranging from 9 - 14 f old r elative to the parent. Analysis of the latte r s train by Southe rn hybr idization indicated that the cassette was inde e d amplified several fold in MTX-re sistant derivatives. A sing l e strain, in which the cassette was inserted at the !lEA;], loc u.s , was used to examine in more detai 1 , the parameters affecting DFRl gene amplificat~ion in yeast . The mean B- gal activity in drug-resistant derivatives of this strain could be increased from 3 to 6 or 7 fold relative to the parent, by stepwise sel ection using increasing MTX concentrations. B-gal overproduction was found to be un stable in all primary and highly -resistant isolates examined. There was no indication, h owever, of a decrease i n growth r a t e in MTX-res i s tant cells which overproduced B - gal.

The interpretation of environmental sustainability (ES) by the IOC/Olympic Games 1994-2008

The purpose of this qualitative multi-case study was to examine the interpretation of environmental sustainability (ES) within the Olympic 11 Movement. Two research questions guided the inquiry - first, how has the concept of ES been defined by the International Olympic Committee (lOC), and second, how has the concept of ES been defined and enacted by the Organizing Committees ofthe Olympic Games (OCOGs)? During the past two decades, the International Olympic Committee (lOC) established several policies and programs related to ES. Its actions reflect a broader trend of environmentalism within economic and social spheres around the world (Milton-Smith, 2002). Despite the numerous initiatives, the Olympic Games continue to cause significant environmental damage. Frey, et al. (2007) argued that the Olympic Movement contradicts the fundamental premises of ES because the Games are hosted in a two week time period, are situated in a confined area, and accumulate operating and infrastructure costs in the billions of dollars. Further, Etzion (2007) stated "there is positive and significant correlation between firm siz~ and environmental performance" (p. 642) and in the context of the Olympics the sizeimpact relation is striking. Since 1972, the year the UN launched its international environmental awareness efforts, the Summer Olympics grew to 201 nations (39% increase), 10,500 athletes (32% increase), 28 sports (30% increase), and 302 events (43% increase) (Johnson, 2004; Girginov & Parry, 2005; Upegui, 2008). The proliferation of Games activities counters the ES principles that exist within many of the IOC declarations, policies and programs.