Silencing Seed Dormancy Genes to Mitigate Risk of Transgene Flow to Weedy Rice.

Gene flow from transgenic crops to sexually compatible weedy relatives is inevitable, but the risk can be reduced by proposed transgene containment and mitigating strategies. This project aimed to develop a new transgene-mitigating (TM) approach to reduce the fitness of transgene-containing plants in weed populations by linking a primary transgene with multiple TM factors to silence major genes for seed dormancy in weedy rice. A built-in construct, which consists of the herbicide resistance (HR) gene, Bar and inverted repeat sequences (IRSs) from coding sequences of the seed dormancy genes SD7-1, SD7-2, and SD12 (Bar::IRS^SD7-1::IRS^SD7-2::IRS^SD12), was used to transform the janopica cultivar, Nipponbare. A total of 8 transgenic events were generated and 2 (25%) were found to have a single-copy of the construct. Transgenic T₀  lines from all derived plants were morphologically similar to Nipponbare, except for glufosinate resistance. To evaluate gene-silencing effects in segregating populations, T₀ plants with single copies of construct (heterozygous for the transgene or F1-like) were crossed with two lines of weedy red rice from temperate and tropical areas, respectively, to generate BC1F1-like and higher segregating generations. The two BC1F1-like populations segregated equally into herbicide resistance and susceptible (HS) groups. The HR and HS plants displayed 50% and 26% germination, respectively, for seeds stored at room temperature for 90 days. The significantly higher germination percentage (p < 0.01) for the HR group indicates that one or more of the 3 dormancy genes were silenced by small RNAs generated from IRSs on the built-in construct.  Research is being conducted: 1) to evaluate HR plants in higher generations for seed dormancy and longevity, and other fitness-related traits under green house and field conditions, and 2) to determine the silencing effects of the TM construct on genotypic and allelic frequencies of the 3 loci across generations.