The seeds of a number of Brassica L. species are cultivated for the production of oil. Collectively, the oilseed Brassica sp., generally referred to as rapeseed, supply more than 13% of the world's supply of edible oils and rank third behind soybean and oil palm in importance. The term "canola" was adopted by the Canadians in 1979 and used to describe oilseed Brassica cultivars that produce oils containing less than 2% erucic acid and to describe defatted seed meals with less than 30 pmol g-' of aliphatic glucosinolates. Brassica napus L. canola cultivars are currently dominant in U.S. production, although canola-quality Brassica rapa (synonymous with Brassica campestris) cultivars also exist (Raymer et al., 1990).Production of canola in the U.S. has grown at a modest rate during the last 10 years from virtually O in 1985 to 165,000 ha in 1995 (C. Boynton, U.S. Canola Association, personal communication). As production of canola continues to grow, insect problems are expected to become more serious (Lamb, 1989). This may be particularly true as canola production expands in the southeastern United States and California, where mild winter temperatures are likely to lead to increased herbivory, as compared with the much cooler areas (e.g. Canada), where canola has historically been produced. Ubiquitous lepidopteran Brassica specialists, such as the DBM and the CBL, and generalist lepidop-terans, such as the BAW and CEW, may increase in importance where canola is grown in warmer regions (Buntin and Raymer, 1994). This scenario may be especially viable with regard to generalist herbivores, since glucosinolates, a hypothesized antiherbivorant to generalist insects (Giamoustaris and Mithen, 1995), have been bred out of canola-quality rapeseed. Transgenic canola cultivars with insecticidal properties will certainly play a major role in integrated pest management strategies for canola pests (Talekar and Shelton, 1993;Evans and Scarisbrick, 1994). The objectives for this study were 2-fold. (a) To determine the effect of Bt expression in B. napus on antibiosis for severa1 lepidopteran insects. Unlike the related Bt soybean study (Stewart et al., 1996), in which only few, low-expressing synthetic Bacillus thuringiensis insecticidal crystal protein ( B t crylAc) plants were produced, the likelihood of obtaining a wide range of B t expression is greater using a species that is more amenable to genetic transformation, such as B. napus. (b) To develop biological tools to test models pertaining to biotechnological risk assessment. It is possible that a plant species, such as B. napus, that is able to persist in nonagricultural environments could become more weedy in a transgenic form if the transgene confers an increment of fitness and the plant is naturalized in areas of its cultivation. Thus, we developed the B t canola to ultimately test population-leve1 ecological hypotheses.

Rapeseed Brassica napus L. transgenic for a Bacillus thuringiensis (Bt) transgene was developed and was shown to be insecticidal towards certain caterpillars including the diamondback moth Plutella xylostella L. and the corn earworm Helicoverpa zea Boddie. To simulate an escape of the transgenics from cultivation, a field experiment was performed in which transgenic and nontransgenic rapeseed plants were planted in natural vegetation and cultivated plots and subjected to various selection pressures in the form of herbivory from insects. Only two plants, both transgenic, survived the winter to reproduce in the natural‐vegetation plots which were dominated by grasses such as crabgrass. However, in plots that were initially cultivated then allowed to naturalize, medium to high levels of defoliation decreased survivorship of nontransgenic plants relative to Bt‐transgenic plants and increased differential reproduction in favour of Bt plants. Thus, where suitable habitat is readily available, there is a likelihood of enhanced ecological risk associated with the release of certain transgene/crop combinations such as insecticidal rapeseed. This is the first report of a field study demonstrating the effect of a fitness‐increasing transgene in plants.

Concerns exist that transgenic crop x weed hybrid populations will be more vigorous and competitive with crops compared with the parental weed species. Hydroponic, glasshouse, and field experiments were performed to evaluate the effects of introgression of Bacillus thuringiensis (Bt) cry1Ac and green fluorescent protein (GFP) transgenes on hybrid productivity and competitiveness in four experimental Brassica rapa x transgenic Brassica napus hybrid generations (F1, BC1F1, BC2F1 and BC2F2). The average vegetative growth and nitrogen (N) use efficiency of transgenic hybrid generations grown under high N hydroponic conditions were lower than that of the weed parent (Brassica rapa, AA, 2n = 20), but similar to the transgenic crop parent, oilseed rape (Brassica napus, AACC, 2n = 38). No generational differences were detected under low N conditions. In two noncompetitive glasshouse experiments, both transgenic and nontransgenic BC2F2 hybrids had on average less vegetative growth and seed production than B. rapa. In two high intraspecific competition field experiments with varied herbivore pressure, BC2F2 hybrids produced less vegetative dry weight than B. rapa. The competitive ability of transgenic and nontransgenic BC2F2 hybrids against a neighbouring crop species were quantified in competition experiments that assayed wheat (Triticum aestivum) yield reductions under agronomic field conditions. The hybrids were the least competitive with wheat compared with parental Brassica competitors, although differences between transgenic and nontransgenic hybrids varied with location. Hybridization, with or without transgene introgression, resulted in less productive and competitive populations.

The movement of transgenes from crops to weeds and the resulting consequences are concerns of modern agriculture. The possible generation of "superweeds" from the escape of fitness-enhancing transgenes into wild populations is a risk that is often discussed, but rarely studied. Oilseed rape, Brassica napus (L.), is a crop with sexually compatible weedy relatives, such as birdseed rape (Brassica rapa (L.)). Hybridization of this crop with weedy relatives is an extant risk and an excellent interspecific gene flow model system. In laboratory crosses, T 3 lines of seven independent transformation events of Bacillus thuringiensis (Bt) oilseed rape were hybridized with two weedy accessions of B. rapa. Transgenic hybrids were generated from six of these oilseed rape lines, and the hybrids exhibited an intermediate morphology between the parental species. The Bt transgene was present in the hybrids, and the protein was synthesized at similar levels to the corresponding independent oilseed rape lines. Insect bioassays were performed and confirmed that the hybrid material was insecticidal. The hybrids were backcrossed with the weedy parent, and only half the oilseed rape lines were able to produce transgenic backcrosses. After two backcrosses, the ploidy level and morphology of the resultant plants were indistinguishable from B. rapa. Hybridization was monitored under field conditions (Tifton, GA, USA) with four independent lines of Bt oilseed rape with a crop to wild relative ratio of 1200:1. When B. rapa was used as the female parent, hybridization frequency varied among oilseed rape lines and ranged from 16.9% to 0.7%.

As genetically engineered crop varieties near widespread cultivation, both agronomic and environmental concerns mandate the development of effective strategies for isolating transgenic varieties from related non-transgenic varieties or cross-fertile weeds. We present the results of the first field experiment designed to test the effectiveness of two containment strategies that are commonly used in field trials of transgenic crops: (1) an isolation zone devoid of vegetation to discourage emigration of insect pollinators from transgenic plots; and (2) trap crops (non-transgenic varieties of the same crop planted adjacent to the transgenic plot that can "cleanse" emigrating pollinators of transgenic pollen). In conjunction with field trials of genetically engineered canola (Brassica napus) conducted by Calgene, Inc., in California and Georgia, we varied both the width of the barren zone and the presence or absence of a trap crop, and measured the effects on gene escape. Escape was easily detected since the genetic construct inserted into the transgenic canola contained a gene that rendered seedlings resistant to the normally lethal antibiotic kanamycin. Our results suggest that barren zones 4-8 m in width may actually increase seed contamination over what would be expected if the intervening ground were instead planted entirely with a trap crop. When trap crops occupied a limited portion of the isolation zone separating transgenic and non-transgenic varieties, the effectiveness of the trap depended on the width of the isolation zone: they reduced gene escape when the two varieties were separated by 8 m, but increased escape across a 4-m isolation zone. We conclude that, for the relatively short isolation distances we examined, the most effective strategy for reducing the escape of transgenic pollen is to devote the entire region between transgenic and non-transgenic varieties to a trap crop.

arm-season grasses are characterized by the C 4 photosynthetic pathway. This pathway occurs in 18 families of fl owering plants, and 61% of the species belong to the grass family. C 4 plants are usually found between 30° N and 30° S latitudes; however, some species extend beyond this range (Moser et al., 2004). The C 4 pathway gives the warm-season grasses an advantage for performing in hot and dry climates and is one reason why these groups of grasses are found mainly in the tropics and subtropics (Clayton and Renvoize, 1986). Developing warm-season grasses for turf is a relatively new concept that began about 60 yr ago. Most of these grasses were used in their native habitat or introduced for use as forages because they could survive under low fertility and in extreme environments (such as drought). Many stoloniferous grasses are somewhat "plastic" and can be changed by management conditions to perform well for diff erent uses (e.g., forage, turf, water conservation, etc.). Cultivar Development Many of the warm-season turfgrass have genetically controlled self-incompatibility systems (see below), which aid in making crosses. If these systems are not present, then marker genes controlling stem, fl ower, and anther color can often be eff ectively used to distinguish self-pollinated plants from crosses. Plant material for individual crosses should be identifi ed and managed properly before pollination. Crossing can be accomplished in most of these turfgrass species by placing

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.

BackgroundOne theoretical explanation for the relatively poor performance of Brassica rapa (weed) × Brassica napus (crop) transgenic hybrids suggests that hybridization imparts a negative genetic load. Consequently, in hybrids genetic load could overshadow any benefits of fitness enhancing transgenes and become the limiting factor in transgenic hybrid persistence. Two types of genetic load were analyzed in this study: random/linkage-derived genetic load, and directly incorporated genetic load using a transgenic mitigation (TM) strategy. In order to measure the effects of random genetic load, hybrid productivity (seed yield and biomass) was correlated with crop- and weed-specific AFLP genomic markers. This portion of the study was designed to answer whether or not weed × transgenic crop hybrids possessing more crop genes were less competitive than hybrids containing fewer crop genes. The effects of directly incorporated genetic load (TM) were analyzed through transgene persistence data. TM strategies are proposed to decrease transgene persistence if gene flow and subsequent transgene introgression to a wild host were to occur.ResultsIn the absence of interspecific competition, transgenic weed × crop hybrids benefited from having more crop-specific alleles. There was a positive correlation between performance and number of B. napus crop-specific AFLP markers [seed yield vs. marker number (r = 0.54, P = 0.0003) and vegetative dry biomass vs. marker number (r = 0.44, P = 0.005)]. However under interspecific competition with wheat or more weed-like conditions (i.e. representing a situation where hybrid plants emerge as volunteer weeds in subsequent cropping systems), there was a positive correlation between the number of B. rapa weed-specific AFLP markers and seed yield (r = 0.70, P = 0.0001), although no such correlation was detected for vegetative biomass. When genetic load was directly incorporated into the hybrid genome, by inserting a fitness-mitigating dwarfing gene that that is beneficial for crops but deleterious for weeds (a transgene mitigation measure), there was a dramatic decrease in the number of transgenic hybrid progeny persisting in the population.ConclusionThe effects of genetic load of crop and in some situations, weed alleles might be beneficial under certain environmental conditions. However, when genetic load was directly incorporated into transgenic events, e.g., using a TM construct, the number of transgenic hybrids and persistence in weedy genomic backgrounds was significantly decreased.