Haplodiploid androgenetic breeding in oat: genotypic variation in anther size and microspore development stage

Abstract

Oat (Avena spp.) is poorly responsive to the haplodiploidization process, which leads to the production of homozygous lines in one step, increasing breeding efficiency. Androgenetic haploids in small grain cereal crops are obtained from microspores cultured at the mononucleate stage, which can be identified by the size of anthers. In order to identify the appropriate anther size for in vitro culture, microspore cytological analyses were made in Avena sativa cultivars UPF 7, UPF 18, UFRGS 14, Stout and Avena sterilis CAV 3361, cultivated in growth chamber under controlled light and temperature conditions. Variation was observed within and among genotypes for anther size at each microspore developmental stage and according to the position of spikelets in the panicle. Architecture variation in panicle shape and non-linear microsporogenesis maturation increased the challenge of identifying potentially androgenetic oat anthers. Cytological screening before culture is critical in identifying microspores at the right stage for oat androgenesis. Key words: androgenesis, anther culture, biotechnology, haploid breeding, microspores MELHORAMENTO POR HAPLODIPLOIDIZAÇÃO ANDROGENÉTICA:VARIAÇÃO GENOTÍPICA NO TAMANHO DAS ANTERAS E NO ESTÁGIO DE DESENVOLVIMENTO DOS MICRÓSPOROS EM AVEIARESUMO: A aveia (Avena spp.) tem sido pouco responsiva à haplodiploidização, um processo que aumenta a eficiência da seleção no melhoramento por gerar, em uma etapa, linhas puras homozigóticas. A fase mononucleada do micrósporo é critica para o sucesso da androgênese in vitro nos cereais de inverno e, em geral, pode ser inferida pelo tamanho da antera. Foram medidas anteras e analisados citológicamente micrósporos das cultivares de Avena sativa UPF 7, UPF 18, UFRGS 14, Stout e da linhagem CAV 3361 de Avena sterilis, cultivadas em câmaras de crescimento sob temperaturas dianoite variando de 16ºC a 9ºC e 12 horas de intensidade luminosa de 300 mol m -2 s -1. O tamanho das anteras em cada fase de desenvolvimento dos micrósporos variou significativamente entre genótipos e de acordo com a região de inserção das espiguetas na panícula. A variação na arquitetura da panícula e a maturação não linear das espiguetas aumentam as dificuldades para a identificação das anteras potencialmente androgenéticas e podem explicar, em parte, os baixos resultados da androgênese na aveia. Os dados mostram a necessidade de uma análise citológica prévia para auxiliar a determinar a fase ideal de desenvolvimento dos micrósporos potencialmente responsivos à cultura de anteras, para o uso da androgenese na aveia. Palavras-chave: biotecnologia, cultura de anteras, haplóides, melhoramento vegetal

The Effect of Zinc, Copper, and Silver Ions on Oat (Avena sativa L.) Androgenesis

“…The application of suitable physiochemical factors promotes a stress response that arrests microspores or young pollen grains in their gametophytic pathway. The switch from gametophytic to sporophytic routes, named the “development window” by Smykal and Pechan [ 17 ], is relatively limited and happens only between the mononucleate and median binucleate stages of microspores [ 18 ]. However, modifications of the stress pretreatment [ 4 , 19 ], carbohydrate source [ 20 , 21 ], or growth regulators of culture media [ 6 , 22 , 23 ] have led to a significant efficiency improvement of both the androgenesis induction and regeneration of haploid plants (HP).…”

Factors inducing regeneration response in oat (Avena sativa L.) anther culture

“…In most cases, the uninucleate stage of the microspore or just after the first pollen mitosis, i.e., the early binucleate stage, is the most suitable for androgenic response in cereal crops. The most appropriate developmental stage of microspores can be identified by the size of anthers or by staining anthers with acetocarmine to observe the nuclei under a light microscope (De Cesaro et al 2009;Sood and Dwivedi 2015). Also, the degree of starch deposition in the microspores was identified as an efficient cytological indicator to determine the microspore maturity (Mayakaduwa and Silva 2017).…”

“…Such pretests are useful to establish a correlation between easily observable morphological traits and the microspore maturity within the anthers, e.g., if the tip of the panicle is inside the leaf sheath at the level of the second leaf base, as reported by , it means that floret buds contain anthers or microspores suitable for androgenesis. The androgenesis competence of the microspore development stage varies with the species or cultivars but is relatively limited and was named by Smykal and Pechan 2000as the Bdevelopment window.^In this restricted period, the switch from gametophytic to sporophytic routes happens only between the mononucleate and median binucleate stages of microspores (De Cesaro et al 2009). The structural variation of oat panicle shape and morphology accompanied by non-linear microsporogenesis maturation in the panicles makes it difficult to identify anther size with microspores at the appropriate stage.…”

Isolated microspore culture of oat (Avena sativa L.) for the production of doubled haploids: effect of pre-culture and post-culture conditions

“…Oat (Avena sativa L.) is considered one of the more recalcitrant cereal crops with respect to doubled haploidy. Wide hybridization with maize pollen (Rines 2003;Rines and Daheen 1990;Sidhu et al 2006) generates doubled haploids as has anther culture (De Cesaro et al 2009;Kiviharju et al 2000;Kiviharju et al 2005;Ponitka and Slusarkiewicz-Jarzina 2009), but methods are inefficient. Haploid embryo production using wide crosses has an efficiency of 0.8 -6.7% (Sidhu et al 2006) and for anther culture up to 30 green plants/100 anthers has been reported (Kiviharju et al 2005).…”