Use of A 3 Cytoplasm to Reduce Risk of Gene Flow through Sorghum Pollen

search at Ithaca, NE, involving sudangrass, a cultivated form of S. bicolor subsp. drummondii, showed natural A critical impediment to field testing and deployment of transgenic outcrossing averaging 39 and 57% in each of two years sorghum [Sorghum bicolor (L.) Moench] is the threat of gene flow to weedy relatives through pollen. A technique using sorghum with (Pedersen et al., 1998). If similar outcrossing is present A3 cytoplasmic male sterility to control transgene flow through pollen in shattercane, gene flow through pollen from cropped while using nontransgenic pollinators is described and an experiment sorghum to shattercane populations could be rapid. was designed to evaluate the risk of viable pollen flow using A3 hybrids The use of cytoplasmic male sterility to prevent the under field conditions. Seed set under pollinating bags (an indicator release of viable pollen from transgenic corn has been of fertile pollen) was evaluated at the University of Nebraska Field recently proposed by Feil et al. (in press). In their sysLaboratory at Ithaca, NE, in 2001 and 2002 on selfed F2 progeny tem, transgenic plants are male sterile and are grown grown from open pollinated seed of 12 F1 hybrids produced in A1 in a mixture with fertile nontransgenic pollen donors. and A3 cytoplasm. The F2 seed was produced in hybrid yield trials in Additionally, exploiting male sterile:male fertile polli1997 and 1998 at Ithaca, NE. In each evaluation year, the experimental design was a split-split plot with seed production year the main plot nator mixtures has already proven commercially sucfactor, hybrid as the subplot factor, and cytoplasm as the sub-subplot cessful for the production of high oil maize (Bergquist factor. Cytoplasm effects were highly significant, with percent seed et al., 1998a) and high grain quality maize (Bergquist set on A1 F2 individuals averaging 74%, and on A3 F2 individuals et al., 1998b). It appears that exploiting male sterile:male averaging 0.04%. Upper confidence limits (P 0.05) for percent seed fertile pollinator mixtures to reduce the threat of transset were 1.32% or less for the progeny from all A3 hybrids. Polymerase gene flow through pollen in sorghum could be commerchain reaction (PCR) analysis confirmed that four male fertile individcially viable. uals (from a population of 1007) were detected with A3 cytoplasm. Nearly all commercial sorghum hybrids are currently These results support the hypothesis that gene flow through pollen produced by means of A1 cytoplasmic male sterile seed can be severely restricted but not eliminated in sorghum by the use of A3 cytoplasmic male sterility. parents. This is at least partially due to the availability of many lines known to restore fertility (R-lines) when crossed to A1 cytoplasm (Bosques-Vega et al., 1989; Torres-Cardona et al., 1990). Since many lines are A impediment to field testing and deployknown to restore fertility in A1 cytoplasm, the probabilment of transgenic sorghum is the risk of gene ity of male fertile volunteer escapes following sorghum flow to weedy relatives through pollen. Crop-to-weed harvest is also high. In a system such as that proposed gene flow has been documented from sorghum to johnby Feil et al. (in press), the risk of transgene flow from songrass [Sorghum halepense (L.) Pers.], a noxious fertile volunteer escapes is unacceptable for sorghum weed, at distances of up to 100 m (Arriola and Ellstrand, hybrids based on A1 male sterile cytoplasm. Even if 1996). Although the triploid progeny of johnsongrass lines known to maintain A1 cytoplasmic male sterility sorghum hybrids would be expected to be sterile, viable (B1–lines) were to be used as pollinators in the mixture, seed production on johnsongrass sorghum hybrids stray pollen from shattercane or neighboring sorghum has been reported (Arriola and Ellstrand, 1997; Hoangfields with the capacity to restore fertility to A1 cytoTang and Liang, 1988) and fitness of hybrid progeny found to be equivalent to johnsongrass (Arriola and plasmic male steriles could be expected to pollinate Ellstrand, 1997), causing those authors to conclude that some of the transgenic sorghum females and introduce “a transgene that is either neutral or beneficial to johnmale fertile transgenic plants into the population of songrass would likely persist in populations growing in volunteer escaped plants. agricultural conditions under continued gene flow from Because of the near absence of lines capable of rethe crop.” storing fertility to A3 cytoplasmic male sterile lines Shattercane [S. bicolor subsp. drummondii (Nees ex (Bosques-Vega et al., 1989; Torres-Cardona et al., 1990), Steud.) de Wet & Harlan], a noxious weed of economic it should be possible to greatly reduce the risk of gene importance to commercial corn (Zea mays L.), soybean flow through pollen from commercial transgenic sor[Glycine max (L.) Merrill], and sorghum production, ghum to weedy relatives, or commercial nontransgenic crosses freely with cropped sorghum. Previous field resorghum, by adapting the technique proposed by Feil et al. (in press) to A3 cytoplasmic male sterile sorghum. J.F. Pedersen and D.L. Funnell, USDA, ARS, NPA Wheat, Sorghum Such an adaptation would include the following steps. and Forage Research, Dep. of Agronomy, University of NebraskaThe transgenic line is converted to an A3 male-sterile Lincoln, Lincoln, NE 68583-0937; D.B. Marx, Dep. of Biometry, Unicytoplasm line by backcrossing under strict isolation, versity of Nebraska-Lincoln, Lincoln, NE 68583-0712. Joint contribupreferably in a greenhouse or growth chamber to reduce tion of the USDA-ARS and the University of Nebraska Agric. Exp. Stn. as Paper no. 13619, Journal Series, Nebraska Agric. Exp. Stn. the risk of escape of viable transgenic pollen. Once Received 8 March 2002. *Corresponding author (jfp@unlserve. converted to an A3 line, seed increase would be perunl.edu). formed in the field following the normal procedure for A/B-lines (Quinby and Schertz, 1970) with intensive Published in Crop Sci. 43:1506–1509 (2003).