@article{halfhill_sutherland_moon_poppy_warwick_weissinger_rufty_raymer_stewart_2005, title={Growth, productivity, and competitiveness of introgressed weedy Brassica rapa hybrids selected for the presence of Bt cry1Ac and gfp transgenes}, volume={14}, ISSN={["1365-294X"]}, DOI={10.1111/j.1365-294X.2005.02649.x}, abstractNote={Abstract}, number={10}, journal={MOLECULAR ECOLOGY}, author={Halfhill, MD and Sutherland, JP and Moon, HS and Poppy, GM and Warwick, SI and Weissinger, AK and Rufty, TW and Raymer, PL and Stewart, CN}, year={2005}, month={Sep}, pages={3177–3189} }
 @article{halfhill_zhu bin_raymer_millwood_weissinger_stewart_2004, title={Hybridization and backcrossing between transgenic oilseed rape and two related weed species under field conditions}, volume={3}, ISBN={1635-7922}, DOI={10.1051/ebr:2004007}, abstractNote={Determining the frequency of crop-wild transgene flow under field conditions is a necessity for the development of regulatory strategies to manage transgenic hybrids. Gene flow of green fluorescent protein (GFP) and Bacillus thuringiensis (Bt) transgenes was quantified in three field experiments using eleven independent transformed Brassica napus L. lines and the wild relatives, B. rapa L. and Raphanus raphanistrum L. Under a high crop to wild relative ratio (600:1), hybridization frequency with B. rapa differed among the individual transformed B. napus lines (ranging from ca. 4% to 22%), however, this difference could be caused by the insertion events or other factors, e.g., differences in the hybridization frequencies among the B. rapa plants. The average hybridization frequency over all transformed lines was close to 10%. No hybridization with R. raphanistrum was detected. Under a lower crop to wild relative ratio (180:1), hybridization frequency with B. rapa was consistent among the transformed B. napus lines at ca. 2%. Interspecific hybridization was higher when B. rapa occurred within the B. napus plot (ca. 37.2%) compared with plot margins (ca. 5.2%). No significant differences were detected among marginal plants grown at 1, 2, and 3 m from the field plot. Transgene backcrossing frequency between B. rapa and transgenic hybrids was determined in two field experiments in which the wild relative to transgenic hybrid ratio was 5-15 plants of B. rapa to 1 transgenic hybrid. As expected, ca. 50% of the seeds produced were transgenic backcrosses when the transgenic hybrid plants served as the maternal parent. When B. rapa plants served as the maternal parent, transgene backcrossing frequencies were 0.088% and 0.060%. Results show that transgene flow from many independent transformed lines of B. napus to B. rapa can occur under a range of field conditions, and that transgenic hybrids have a high potential to produce transgenic seeds in backcrosses.}, number={2}, journal={Environmental Biosafety Research}, author={Halfhill, M. D. and Zhu Bin, Warwick S. I. and Raymer, P. L. and Millwood, R. J. and Weissinger, A. K. and Stewart, C. N.}, year={2004}, pages={73} }
 @article{halfhill_millwood_weissinger_warwick_stewart_2003, title={Additive transgene expression and genetic introgression in multiple green-fluorescent protein transgenic crop x weed hybrid generations}, volume={107}, ISSN={["1432-2242"]}, DOI={10.1007/s00122-003-1397-7}, abstractNote={The level of transgene expression in crop x weed hybrids and the degree to which crop-specific genes are integrated into hybrid populations are important factors in assessing the potential ecological and agricultural risks of gene flow associated with genetic engineering. The average transgene zygosity and genetic structure of transgenic hybrid populations change with the progression of generations, and the green fluorescent protein (GFP) transgene is an ideal marker to quantify transgene expression in advancing populations. The homozygous T(1) single-locus insert GFP/ Bacillus thuringiensis (Bt) transgenic canola ( Brassica napus, cv Westar) with two copies of the transgene fluoresced twice as much as hemizygous individuals with only one copy of the transgene. These data indicate that the expression of the GFP gene was additive, and fluorescence could be used to determine zygosity status. Several hybrid generations (BC(1)F(1), BC(2)F(1)) were produced by backcrossing various GFP/Bt transgenic canola ( B. napus, cv Westar) and birdseed rape ( Brassica rapa) hybrid generations onto B. rapa. Intercrossed generations (BC(2)F(2) Bulk) were generated by crossing BC(2)F(1) individuals in the presence of a pollinating insect ( Musca domestica L.). The ploidy of plants in the BC(2)F(2) Bulk hybrid generation was identical to the weedy parental species, B. rapa. AFLP analysis was used to quantify the degree of B. napus introgression into multiple backcross hybrid generations with B. rapa. The F(1) hybrid generations contained 95-97% of the B. napus-specific AFLP markers, and each successive backcross generation demonstrated a reduction of markers resulting in the 15-29% presence in the BC(2)F(2) Bulk population. Average fluorescence of each successive hybrid generation was analyzed, and homozygous canola lines and hybrid populations that contained individuals homozygous for GFP (BC(2)F(2) Bulk) demonstrated significantly higher fluorescence than hemizygous hybrid generations (F(1), BC(1)F(1) and BC(2)F(1)). These data demonstrate that the formation of homozygous individuals within hybrid populations increases the average level of transgene expression as generations progress. This phenomenon must be considered in the development of risk-management strategies.}, number={8}, journal={THEORETICAL AND APPLIED GENETICS}, author={Halfhill, MD and Millwood, RJ and Weissinger, AK and Warwick, SI and Stewart, CN}, year={2003}, month={Nov}, pages={1533–1540} }
 @article{richards_halfhill_millwood_stewart_2003, title={Quantitative GFP fluorescence as an indicator of recombinant protein synthesis in transgenic plants}, volume={22}, ISSN={["0721-7714"]}, DOI={10.1007/s00299-003-0638-1}, abstractNote={The utility of green fluorescent protein (GFP) for biological research is evident. A fluorescence-based method was developed to quantify GFP levels in transgenic plants and protein extracts. Fluorescence intensity was linear with increasing levels of GFP over a range that encompasses transgene expression in plants by the cauliflower mosaic virus 35S promoter. Standard curves were used to estimate GFP concentration in planta and in protein extracts. These values were consistent with ELISA measurements of GFP in protein extracts from transgenic plants, indicating that the technique is a reliable measure of recombinant GFP expression. The levels of in planta GFP expression in both homozygous and hemizygous plants was then estimated. Homozygous transgenic plants expressed twice the amount of GFP than hemizygous plants, suggesting additive transgene expression. This methodology may be useful to simplify the characterization of transgene expression in plants.}, number={2}, journal={PLANT CELL REPORTS}, author={Richards, HA and Halfhill, MD and Millwood, RJ and Stewart, CN}, year={2003}, month={Sep}, pages={117–121} }
 @article{halfhill_millwood_rufty_weissinger_stewart_2003, title={Spatial and temporal patterns of green fluorescent protein (GFP) fluorescence during leaf canopy development in transgenic oilseed rape, Brassica napus L.}, volume={22}, ISSN={["1432-203X"]}, DOI={10.1007/s00299-003-0696-4}, abstractNote={The green fluorescent protein (GFP) holds promise as a field-level transgene marker. One obstacle to the use of GFP is fluorescence variability observed within leaf canopies. In growth chamber and field experiments, GFP fluorescence in transgenic oilseed rape ( Brassica napus) was shown to be variable at each leaf position over time and among different leaves on the same plant. A leaf had its highest GFP fluorescence after emergence and, subsequently, its fluorescence intensity decreased. GFP fluorescence intensity was directly correlated with the concentration of soluble protein. The concentration of the genetically linked recombinant Bacillus thuringiensis (Bt) cry1Ac endotoxin protein also was examined, and GFP fluorescence was positively correlated with Bt throughout development. The results show that GFP can be used as an accurate transgene marker but that aspects of plant developmental should be taken into account when interpreting fluorescence measurements.}, number={5}, journal={PLANT CELL REPORTS}, author={Halfhill, MD and Millwood, RJ and Rufty, TW and Weissinger, AK and Stewart, CN}, year={2003}, month={Dec}, pages={338–343} }