Analysis of Genes for Stigma Coloration in Rice - IRRI books

advantageous than wing somatic cells, since the inserted gene could be duplicated, thus
eliminating segregation in future generations.
Rice plants were obtained from isolated microspore culture as early as 1980 (Chen
et al 1980). However, the low induction frequencies of callus and plants were
attributable to the lack of understanding of factors influencing the initiation of
androgenesis and further development of microspores in vitro.
We have tried to overcome the problem of low efficiency by treatment of the donor
plants and by manipulation and evaluation of media and media components.
Callus formation and plant regeneration in isolated microspore
culture of three rice varieties
A number of methods have been employed to produce pollen-derived rice embryos:
shedding culture, cold treatment, and preculture of anthers before isolation (Nitsch
1974). Isolation and culture in this experiment followed Nitsch (1974).
Japonica variety Taipei 309
Our initial attempt to study isolated microspore culture in rice used Taipei 309, a
responsive variety. Isolated microspores from 7-d precultured anthers were cultured in
E10 [modified Gamborg’s (1968) B5] liquid medium, E10A (with 1 mM alanine),
E10G (with 1 mM glutamine), and El0P (with 1 mM proline). Only large microspores
50–58 µm in diameter with thin, light pink-colored cell walls (p-grains) seemed to
develop into pollen embryos. Eight-cell division frequency increased rapidly during
the first 10 d of culture in the 4 media tested (Fig. 1). E10P yielded the highest division
frequency of 3.9% and a calli plating efficiency of 3.5% (Table 1). After a 14-d culture
1. Effect of media containing various amino acids on division frequency of Taipei 309 pollen at different
durations in culture (av of 4 replications ±SE).
312 Zapata et al