Abstract
SummaryWide crosses have been used for decades as a method for transferring novel genetic material and traits in plant breeding. Historically, many products of wide crosses require tedious and inefficient surgical embryo rescue prior to embryo abortion to recover single plantlets. We have utilized transgenic switchgrass (Panicum virgatum L. cv Alamo) as a pollen donor in conjunction with antibiotic or herbicide selection for recovery of intra‐and interspecific F1 crosses by using developing ovules from the female parent and selecting for embryogenic cultures derived from the in situ immature embryo. Using this approach, several intravarietial crosses were generated between transgenic Alamo and the switchgrass varieties Kanlow, Blackwell and Cave‐in‐Rock as well as an interspecific cross with Atlantic coastal panicgrass. This procedure selected F1 embryogenic callus produced from the developing embryo contained within isolated immature ovules. Several clonal plants were successfully regenerated from each cross. Southern blot, PCR, phenotypic analyses and genomic analysis confirmed F1 hybrids. Using genotyping‐by‐sequencing shows the hybridization of the recovered plants by determining the ratio of transgressive markers to total compared markers between parents and their potential offspring. The ratio of transgressive markers to total compared markers was significantly lower between parents and their predicted offspring than between parents and offspring unrelated to them. This approach provides the possibility to move useful transgenes into varieties that are recalcitrant to direct transformation which can be optionally segregated thus useful to create new hybrids, as well as recovery of wide crosses that are either difficult or impossible using traditional techniques.
Highlights
Switchgrass (Panicum virgatum L.) is a semi-domesticated plant which is highly heterozygous and an anemophilous obligate outcrosser with both prefertilization and postfertilization incompatibility systems present (Martinez-Reyna and Vogel, 1998, 2002)
Selection of resistant embryogenic colonies occurred in Panicum virgatum cv Alamo over a 6- to 8-week period under either hygromycin and bialaphos selection
Leaves of WT Alamo swabbed with 3% (v/v) Finale showed sensitivity (Figure 1f), whereas those of the T0 p35S-bar transgenic regenerated plants scored for resistance to the herbicide (Figure 1g)
Summary
Switchgrass (Panicum virgatum L.) is a semi-domesticated plant which is highly heterozygous and an anemophilous obligate outcrosser with both prefertilization and postfertilization incompatibility systems present (Martinez-Reyna and Vogel, 1998, 2002). Two main ecotypes (‘lowland’ and ‘upland’ types) have been characterized and described by geographical adaptation (Casler et al, 2011). These two ecotypes show distinct morphological and physiological characteristics (Casler et al, 2011; Ersoz et al, 2012; Narasimhamoorthy et al, 2008), and the genetic distinctiveness of their nuclear genomes has been established (Eberhardt and Newell, 1959; Hopkins et al, 1996; McMillan and Weiler, 1959). With ample phenotypic and genetic diversity characterized within and among switchgrass cultivars and populations, data on hybrid plant development are somewhat limited by the time-consuming and laborious process to recover hybrid plants and fertile alloploids (Hultquist et al, 1996; Martinez-Reyna and Vogel, 2002)
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