Non-random transmission of parental alleles into crop-wild and crop-weed hybrid lineages separated by a transgene and neutral identifiers in rice

Abstract

It is essential to assess environmental impact of transgene flow from genetically engineered crops to their wild or weedy relatives before commercialization. Measuring comparative trials of fitness in the transgene-flow-resulted hybrids plays the key role in the assessment, where the segregated isogenic hybrid lineages/subpopulations with or without a transgene of the same genomic background are involved. Here, we report substantial genomic differentiation between transgene-present and -absent lineages (F₂-F₃) divided by a glyphosate-resistance transgene from a crop-wild/weed hybrid population in rice. We further confirmed that such differentiation is attributed to increased frequencies of crop-parent alleles in transgenic hybrid lineages at multiple loci across the genome, as estimated by SSR (simple sequence repeat) markers. Such preferential transmission of parental alleles was also found in equally divided crop-wild/weed hybrid lineages with or without a particular neutral SSR identifier. We conclude that selecting either a transgene or neutral marker as an identifier to create hybrid lineages will result in different genomic background of the lineages due to non-random transmission of parental alleles. Non-random allele transmission may misrepresent the outcomes of fitness effects. We therefore propose seeking other means to evaluate fitness effects of transgenes for assessing environmental impact caused by crop-to-wild/weed gene flow.

Figure 1

A schematic pedigree to illustrate the production of F₂ (a) and BC₁ (b) crop-wild or crop-weed hybrid lineages for estimating the fitness effect of a transgene. “GE crop” indicates a genetically engineered crop parent; “Wild” indicates a wild or weedy parent; “++ and +−” indicate transgene-homozygous and -hemizygous GE lineages, and “− −” indicates non-GE lineages.

Figure 2

Phenotypic variation of seedlings in F₃ transgene-present (orange arrow) and transgene-absent (yellow arrow) hybrid lineages derived from an artificial cross between an epsps (5-enolpyruvoylshikimate-3-phosphate synthase) transgenic rice line and wild rice (Oryza rufipogon).

Figure 3

A genomic linkage map illustrating the physical location of the epsps (5-enolpyruvoylshikimate-3-phosphate synthase) transgene on chromosome-1 (green box), the six neutral SSR (simple sequence repeat) identifiers (pink boxes for crop-wild and blue boxes for crop-weed hybrids), and 52 (red letters, for crop-wild hybrids) and 32 (blue letters, for crop-weed hybrids) SSR markers located across the 12 rice chromosomes.

Figure 4

Average frequencies of crop-parent alleles in hybrid lineages (a) and groups (b) separated from an F₂ or F₃ hybrid population containing an epsps transgene. “W” represents wild rice (Oryza rufipogon); “WD” represents weedy rice. Dark-gray and white columns in (a) indicate lineages with and without a transgene, respectively. Light-gray columns in (b) indicate randomly formed groups. Bars indicate standard deviation.

Figure 5

Average frequencies of parental alleles in hybrid lineages (a,b) and ideal groups (c) separated from a transgene-free F₂ or F₃ crop-wild (W) /weed (WD) hybrid population. (a) CM lineages are grouped by sampling individuals with crop-parent markers; (b) WM or WDM lineages are grouped by sampling individuals with wild- or weedy-parent markers; (c) Ideal groups are formed randomly. Blue and light-blue columns indicate frequencies of crop-parent alleles in lineages with and without markers, respectively; orange and light-orange columns indicate frequencies of wild- or weedy-parent alleles in lineages with and without markers, respectively. Bars indicate standard deviation. *,**Significance at P < 0.05 or P < 0.01, based on the one-tail paired t test.

Figure 6

Hybrid lineages (a) and ideal groups (b) created from an experimental hybrid population (F₂ or F₃) for analyses. Identifier represents either a transgene or a neutral marker that are used to separate hybrid lineages.