First stages of microspore reprogramming to embryogenesis through anther culture in Prunus armeniaca L.

Abstract

Prunus armeniaca L. is a worldwide known species, very important particularly in the Mediterranean basin. Microspore embryogenesis through in vitro anther culture is a widely used method to obtain haploid and doubled haploid (DHs) plants which are being routinely used in breeding programmes for new superior cultivar development in many crops. Haploid–diploidization through gametic embryogenesis allows single-step development of complete homozygous lines from heterozygous parents. In the case of fruit crops, with long reproductive cycle, a high degree of heterozygosity, large size, and, often, self-incompatibility, there is no way to obtain haploidization through conventional methods. Induction of microspore embryogenesis in vitro is switched by a stress treatment. In many species, heat or cold stress has been reported to trigger pollen embryogenesis, the response being genotype dependent. In the present work we analyzed whether microspore reprogramming could be induced in apricot cultivars by cold stress through anther culture. We report the development of an in vitro anther culture protocol in P. armeniaca L. and analyse the response of several cultivars to stress treatments and culture media for inducing pollen embryogenesis. Results showed the formation of multicellular pollen and proembryos. The effect of two culture media in the embryogenic response was also analyzed, being the responses genotype-dependent. Monitoring of the cellular changes on the microspores was performed by structural and confocal microscopy analyses. Results indicated that the reprogramming of the microspore and the first steps of the embryogenic pathway have been achieved in different varieties of P. armeniaca, which constitutes a crucial step in the design of protocols for the regeneration of microspore-derived embryos and DH plants, for future potential applications in breeding programmes of this economically important fruit tree