HAU16008-02-6 - NEWS AND VIEWS Either/or selection markers...

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N E W S A N D V I E W S Identifying plants that have successfully incorporated transgenic DNA is somewhat akin to finding a transgenic needle in a non- transgenic haystack. Until now, plant biolo- gists have commonly relied on positive selection of a marker gene—usually cotrans- ferred with a transgene of interest—that provides cells integrating the DNA with a growth advantage over nontransformed cells under selective conditions. In this issue, Erikson et al. 1 present a novel marker system that is based on the metabolism of D -amino acids (mirror images of L -amino acids). Their system allows both positive and negative selection, expanding the choice of markers available for creating transgenic plants. In contrast to the increase in number of transformable species, the choice of widely applicable marker genes has not expanded significantly. Traditionally, selection has been carried out with a handful of antibiotic- or herbicide-resistance genes 2 , most of which, whether justified or not, have become infa- mous for their potential danger to health or the environment. In addition, repetitive use of these same markers hampers the combination of transgenes and sequential transformation and also increases the risk of unwanted mutual gene silencing. As an alternative approach, Erikson et al. have generated transgenic Arabidopsis thal- iana plants expressing D -amino acid oxidase (DAAO) from the DAO1 gene of the yeast Rhodotorula gracilis . This enzyme catalyzes deamination of several amino acids with stereospecificity for their D enantiomer (the mirror image, at the asymmetric carbon atom, of the L configuration). Erikson et al. have demonstrated that development of non- transgenic A. thaliana seedlings is arrested soon after germination in the presence of D - serine or D -alanine, whereas plants expressing DAO1 detoxify these amino acids and grow without developmental abnormalities. In contrast, D -valine or D -isoleucine are non- toxic to wild-type plants but kill plantlets transgenic for DAO1 . Thus, the same trans- gene product allows both positive and nega- tive selection, depending on the selective agent applied ( Fig. 1a ). This system has obvious advantages. Namely, selective agents can be applied by spraying on to soil-grown seedlings—a pro- cedure that once made the bialaphos resist- ance genes ( PAT , encoding phosphinothricin- N -acetyl transferase, or BAR , encoding bialaphos resistance) and their selective agent phosphinothricin superior, for many pur- poses, to the antibiotic-resistance markers encoding neomycin phosphotransferase ( NPTII ) and hygromycin phosphotransferase ( HPT ) 1 . Additionally, the natural levels of D -amino acids and other DAO substrates in plants do not appear to be a problem, because even plants with high DAAO expression are indistinguishable from transgene-free plants under nonselective conditions. Selection
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  • Spring '10
  • TaiDat
  • Genetics, Embryonic stem cell, Somatic cell nuclear transfer

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