An efficient procedure for embryogenic callus induction and double haploid plant regeneration through anther culture of Thai aromatic rice (Oryza sativa L. subsp. indica)

Abstract

Anther culture is a biotechnology technique that can be used for the production of pure lines. The aims of this investigation were to induce embryogenic callus from major and minor culms of Thai aromatic rice cultivars and to subsequently regenerate double-haploid green plantlets by the application of exogenous polyamines. Embryogenic callus derived from anther culture was successfully induced in varieties KDML105, Homjan (HJ), and Pathumthani 1 (PT1). Production of embryogenic callus from anthers collected from the major culms was greater than those collected from the minor culms, especially in cultivar HJ. Plantlet regeneration in the three rice cultivars was observed from embryogenic callus and was highest, at 12.1%, from variety HJ treated with 0.5 mM spermidine. Plantlet regeneration from anther-derived embryogenic callus was dependent on the plant genotype, the types of exogenous polyamines, and the interactions of these factors. The percentage of haploid plantlets regenerated in PT1, KDML105, and HJ were 68.1%, 70.7%, and 78.5%, respectively. Only haploid plantlets were treated with colchicine for double-haploid production. This investigation has increased the knowledge of both embryogenic callus induction and plantlet regeneration in aromatic rice and has lead to the development of a pure, double-haploid line for the use in rice breeding programs in Thailand.

Exploring factors affecting anther culture in rice (Oryza sativa L.)

“…Culturing of anthers of PT-1, KDML-105, and HJ rice variety at 25 ± 2°C in darkness for 6 weeks successfully induced embryogenic calli [77]. Highest callus induction frequency was obtained in anther culture of japonica/indica and indica/japonica hybrids [78] at 25 ± 2°C for 8 weeks.…”

Scope of androgenesis for genetic improvement in rice

“…Either osmotic stress (mannitol), mild heat (30-40 o C) or chemical or more specifically, the cold pretreatment of anthers is needed to initiate sporophytic growth [28,[32][33]. Mishra and Rao have listed various studies on stress pre-treatment of anthers to induce callusing response [23].…”

“…Nitrogen promotes development of superior microspores with high embryogenic activity [39].The most common medium for development of androgenic callus is N6 and MS-medium + 2,4-D(2-3mg/l) with some variations (higher nitrogen and iron concentration) as needed [40], while MS medium, supplemented with BAP/Kinetin with lower concentration of NAA, is generally used for plant regeneration. Various experiments conducted by different researchers have also shown that application of polyamines promotes the plantlet regeneration [33]. It was observed that IAA reduced the shoot formation, but increased the root development [41].…”

Anther Culture as a Supplementary Tool for Rice Breeding

“…In rice anther culture, the morphological trait that has been used is the measured distance between the nodes of the last two leaves: the flag leaf, and the penultimate leaf [27,29]. In some cases the panicle length at the time of harvest has also been used as a visually identifiable guide [30,31]. The use of a direct cytological marker such as the degree of starch accumulation in microspores has been identified as more accurate than the internode distance and even more convenient than the laborious nuclear staining process to rapidly assess microspore maturity.…”