Haploid induction in Mimulus aurantiacus Curtis obtained by pollination with gamma irradiated pollen

- A haploid induction protocol was developed for Mimulus aurantiacus.
- A new microsatellite marker was developed for M. aurantiacus.
- A new method for embryo rescue has proved successful.
- The nuclear DNA content of M. aurantiacus was measured for the first time.

In this study we present a successful approach to haploid induction in Mimulus aurantiacus Curtis. In Europe, it is a promising ornamental plant and is intensively studied for its adaptive differentiation and speciation processes due to high ecological and phenotypic diversity of natural populations. The small nuclear DNA content, determined in our study as 0.607 pg DNA (2C), indicated the suitability of this species for DNA sequencing approaches. Among several methods of haploid induction tested, pollination with gamma-irradiated pollen proved to be the only effective method and produced 4 haploid, 319 diploid, 5 triploid and 3 aneuploid plants. A new microsatellite marker was developed for determination of plantlets origin, from EST sequences deposited in NCBI GenBank. Together with the Mim-top6B microsatellite marker developed earlier, it revealed that 43 of the tested plants were homozygous and 171 heterozygous. The majority of heterozygotes were hybrids between female gamete and irradiated pollen, which retained its fertilization ability after irradiation. Genetic analysis of triploid and aneuploid regenerants provided novel insight into the mechanism of haploid embryo formation following pollination with irradiated pollen. On the basis of genetic analysis of a heterozygous aneuploid (with ploidy between haploid and diploid levels), we propose that paternal chromosome elimination might be the causative mechanism of haploid plant formation while the genetic structure of triploid regenerants provided evidence of regeneration from endospermal cells.