Production of Haploid Plant of ‘Banpeiyu’ Pummelo [Citrus maxima (Burm.) Merr.] by Pollination with Soft X-Ray-Irradiated Pollen

Abstract

To induce haploid plants in Citrus maxima (Burm.) Merr. 'Banpeiyu', we evaluated the effect of pollination with soft X-ray-irradiated pollen on fruit set and seed development, and carried out ovule culture. When 'Banpeiyu' pummelo pistils were pollinated with X-ray-irradiated pollen of 'Fukuhara' sweet orange [C. sinensis (L.) Osbeck], the exposure doses affected the fruit set. The number of seeds per fruit was also affected by the exposure dose, and tended to decrease as the dose increased; however, all developed seeds obtained from these crosses were diploid. In the ovule culture of 'Banpeiyu' pummelo, six embryoids shown haploidy were obtained in all treatments. One haploid plantlet with 9 chromosomes was regenerated from an embryoid in a culture of ovules established 40 days after pollination with 400 Gray (Gy)-irradiated pollen of 'Tosa-buntan' pummelo (C. maxima). This haploid was suggested to be derived from 'Banpeiyu' pummelo by random amplified polymorphic DNA (RAPD) and cleaved amplified polymorphic sequence (CAPS) analysis.

Induction of haploid plants in citrus through gamma-irradiated pollen and ascertainment of ovule age for maximum recovery of haploid plantlets

“…From the earlier findings by different researchers on induction of haploid plants through irradiated pollen techniques it was confirmed that the best irradiation dose of pollen for induced parthenogenesis varies from crop to crop and species to species; Froelicher et al (2007) reported that 300 Gy was best for induction of haploid plants in mandarin, whereas doses between 25 and 50 Gy were found optimum for squash (Kurtar et al, 2002), and a dose of 1500 Gy gave the best results for haploid induction in kiwifruit (Chalak and Legave, 1997). Another interesting result obtained from this experiment was that haploid plants were obtained only from ovule cultures at 50 DAP; however, Yahata et al (2010) reported that the ovule culture at approximately 40 DAP seemed to be appropriate for haploid induction in Citrus. On the other hand, all the plantlets regenerated from in vitro culture of embryos excised from mature seeds were diploid.…”

“…al., 1990;Chalak and Legave, 1997;Musial and Przywara, 1998;1999), apple (Zhang and Lespinasse, 1991;De Witte and Keulemans, 1994;Höfer and Lespinasse, 1996), pear (Bouvier et al, 1993), European plum (Peixe et al, 2000), blackberry (Naess et al, 1998), sweet cherry (Höfer and Grafe, 2003), and Citrus (Froelicher et al, 2007;Aleza et al, 2009;Yahata et al, 2010), the induction mechanism of those haploid plants is not yet clear. It is therefore imperative to ascertain the hybridity of those in vitro-raised plantlets to confirm their genetic origin using a precise system.…”

“…Recent biological studies of various plants suggest that parthenogenesis induced by pollination with inactivated (irradiated) pollen followed by pseudofertilized ovule culture might represent the appropriate technique for the induction of haploid plants as this technique has a higher frequency of haploid induction (Raquin, 1985;Rode and Dumas de Vaulx, 1987;Sauton and Dumas de Vaulx, 1987;Pandey et al, 1990;Zhang and Lespinasse, 1991;Bouvier et al, 1993;Katoh et al, 1993;Sari et al, 1994;Grouh et al, 2011). In Citrus, however, there is only scanty information available on parthenogenesis induced by irradiated pollen followed by in vitro culture of pseudofertilized ovules (Froelicher et al, 2007;Yahata et al, 2010) and it is the need of the hour to optimize the dose of irradiation for induction of haploid plants in citrus as conventional mutational damage may occur at low doses (Engvild, 1985;Sanders et al, 1991), whereas higher doses may lead to lethality problems for pollen grains. Moreover, no studies have been encountered to date concerning the age of citrus ovules for maximum recovery of haploid plants through the gamma-irradiated pollen technique.…”

“…Gamma rays are commonly used in haploidy programs because of their simple application, good penetration, reproducibility, high mutation frequency, and fewer disposal problems (Chahal and Gosal, 2002). However, the major obstacle for adapting this technique is the degeneration of the induced haploid embryo at the early stage of seed development, which causes seed abortion (Froelicher et al, 2007;Yahata et al, 2010), resulting in little or no recovery of haploid plants from the harvested seeds following pollination with gamma-irradiated pollen. Recent biological studies of various plants suggest that parthenogenesis induced by pollination with inactivated (irradiated) pollen followed by pseudofertilized ovule culture might represent the appropriate technique for the induction of haploid plants as this technique has a higher frequency of haploid induction (Raquin, 1985;Rode and Dumas de Vaulx, 1987;Sauton and Dumas de Vaulx, 1987;Pandey et al, 1990;Zhang and Lespinasse, 1991;Bouvier et al, 1993;Katoh et al, 1993;Sari et al, 1994;Grouh et al, 2011).…”

Morphological Characterization and Evaluation of Reproductive Function in a Haploid Pummelo [<i>Citrus maxima</i> (Burm.) Merr.]

“…Since then, haploids have been produced by anther culture (Germanà 2011, Hidaka et al 1979, Wang et al 2015, interploid hybridization (Oiyama & Kobayashi 1993), and pollination of irradiated pollen (Froelicher et al 2007, Yahata et al 2010. However, these haploids were very weak and grew more slowly than the original diploid plants.…”