Soybean androgenesis I: identification of pyramidal stressors in anther cultures that sustain cell divisions and putative embryo formation from isolated microspore cultures

Soybean [Glycine max (L.) Merr.] is one of the most complete food crops and has fed people and livestock since its domestication. Soybean androgenesis has been a challenging process yet to be advanced significantly. There have been minor advances, e.g., reports of a 2% induction rate in anther culture, formation of roots, rare shoots, but few plantlets and these did not survive to maturity. This research attempted to identify pyramidal stressors (the combination of temperature shock and incubation conditions) for soybean androgenesis using anther cultures. The results across genotypes Jack, Thorne, Williams 82 and IAS-5 indicated that androgenesis can be stimulated up to 9 to 12% induction frequency by the use of 10°C day/8°C night for 3 d then 4°C overnight dark pretreatment of donor plants, a series of incubation temperatures from 11°C to 18°C to 25°C, and nitrogen starvation medium. The adaptation of the anther culture protocol for isolated microspore cultures of IAS-5 and Embrapa-1 resulted in more than 90% of culture replicates (ave. 4 × 105 microspores mL−1, 500 μL per replicate) exhibiting sustained cell divisions with complex multicellular structures, including formation of one or more putative embryos. These results are a promising step towards the use of microspore cultures for soybean androgenesis, and a possible route to doubled haploid breeding for soybean.