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Title: Assessment of long-term transgene expression in barley: Ds-mediated delivery of bar results in robust, stable, and heritable expression

item Bregitzer, Paul
item Brown, Ryan

Submitted to: In Vitro Cellular and Developmental Biology - Plants
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/1/2013
Publication Date: 3/22/2013
Citation: Bregitzer, P.P., Brown, R.H. 2013. Assessment of long-term transgene expression in barley: Ds-mediated delivery of bar results in robust, stable, and heritable expression. In Vitro Cellular and Developmental Biology - Plants. DOI: 10.1007/S11627-013-9507-Y.

Interpretive Summary: Transgenic technologies are powerful techniques for creating useful, genetically modified plants, also known as GMOs. Although there are many examples of widely used, and successful, GMO varieties, there are still problems and limitations of the technology that must be solved. For instance, consumers would prefer GMOs that do not contain selectable marker genes, used in the creation of the GMO crop, that do not have anything to do with the improved trait that the GMO crop delivers. Another problem is that for every successful GMO crop, there are many that are unsuccessful, often because the new gene fails to function properly and contribute the new trait. We have discovered that by using a natural genetic phenomenon, called transposition, GMO barley can be produced where the transgene is included in such a way that predictable, stable trait expression occurs 95% of the time. Transposition can also be used to eliminate the selectable marker gene, and because this is a natural genetic system derived from corn, GMO barley plants can be produced that do not contain any bacterial DNA.

Technical Abstract: The utility of transgenic plants for both experimental and practical agronomic purposes is highly dependent on stable, predictable, and heritable expression of the introduced genes. This requirement is frequently unfulfilled, and transgenes are frequently subject to silencing. Studies of the characteristics of transgenic loci and RNA interference have shown that rearrangements of transgenes occur during the integration process that are potent cues that induce silencing. Techniques such as site-specific recombination and transposon-mediated gene delivery are capable of routinely inserting intact, unrearranged, single-copy transgenes that subsequently display favorable expression characteristics. In this study, the expression of independently transposed Ds-bar cassettes was shown to be stable among 107 barley (Hordeum vulgare L.) lines that had been advanced to at least F5, and in 31 near-isogenic lines (NILs) derived from eight backcrosses of independent transposition events, based on qualitative assessment of herbicide resistance. Heritability, based on qRT-PCR measurement of bar expression, was studied in eight NILs and in their donor parents. The expression of bar was highly correlated in parent and progeny NILs, showing high heritability of bar expression. These data demonstrate the expectation of stable, predictable transgene expression following Ds-mediated delivery.