12. Mapping of the ABERRANT REGIONALIZATION OF EMBRYO 1 gene, ARE1, by CAPS analysis

1)BioScience Center, Nagoya University, Nagoya, 464-8601 Japan
2)Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya, 464-8601 Japan
3)Graduate School of Agricultural and Life Sciences, University of Tokyo, Tokyo, 113-8657 Japan

In plants, most morphogenetic events occur in the post-embryonic phase, and almost all tissues and organs are derived from two meristematic cell masses, the shoot and root apical meristems (SAM and RAM). The SAM and RAM are differentiated during embryogenesis. Thus the most important objective in embryogenesis is considered to be the differentiation of these two apical meristems. We are interested in the mechanism of establishment of the shoot region during rice embryogenesis, since the elucidation of the mechanism of the SAM differentiation in embryogenesis should be one of the most important subjects to be explored in plant developmental biology. For analyzing its molecular mechanism, we took a molecular genetic approach and have screened for embryonic mutants with defects in the establishment of shoot regions.

A rice homeobox gene, OSH1, is thought to have important roles in shoot formation and maintenance. Its expression is localized in the shoot region throughout embryogenesis, and the earliest expression of OSH1 starts at the corresponding region where the SAM will develop by the late globular stage, which is earlier than morphological shoot differentiation (Sato et al., 1996). To date, many embryonic mutants have been identified (Nagato et al., 1989; Hong et al., 1995). Using these mutant stocks, we have rescreened for mutants with abnormal OSH1 expression in late globular stage embryos and obtained some mutants. One of these mutants, aberrant regionalization of embryo 1 (are1), shows broader OSH1 expression than that of WT in the aberrant position at the globular stage. As a result, are1 embryo simultaneously develops two shoots during embryogenesis. These two shoots can normally germinate and develop with some morphologic defects. To isolate the ARE1 gene, we have mapped the ARE1 locus by the linkage analysis using CAPS markers as the first step.

For mapping, we crossed japonica rice cv. Taichung 65 homozygous plants for are1 with an indica rice, cv. Kasalath, homozygous for the wild-type allele (ARE1/ARE1). Then, the F1 plants were cultivated and self-pollinated for obtaining F2 seeds. In the F2 seedlings (ARE1/ARE1, ARE1/are1, and are1/ are1), we screened homozygous plants for are1 by their phenotype and isolated the genomic DNAs for PCR analysis. We used two hundred F2 are1 homozygous plants, and the ARE1 locus was roughly mapped on the long arm of the chromosome 9. For further analysis, we have selected four markers, C670, S1057, C609, and S2655, located near the ARE1 locus. Using these markers, ARE1 was mapped between C609 and S2655 with distance of 1.8cM and 2.2cM, respectively (Fig. 1). For further analysis, about four thousand F2 plants were cultivated, and we are now analyzing these plants with several new markers located between C609 and S2655.


Harushima Y, Yano M, Shomura A, Sato M, Shimano T, Kuboki Y, Yamamoto T, Lin SY, Antonio BA, Parco A, Kajiya H, Huang N, Yamamoto K, Nagamura Y, Kurata N, Khush GS, Sasaki T, 1998. A high-density rice genetic linkage map with 2275 markers using a single F2 population. Genetics 148: 479-94.

Hong S-K, Aoki T, Kitano H, Satoh H, Nagato Y, 1995. Phenotypic diversity of 188 rice embryo mutants. Dev. Genet.16: 298-310.

Nagato Y, Kitano H, Kamijima O, Kikuchi S, Satoh H, 1989. Developmental mutants showing abnormal organ differentiation in rice embryos. Theor. Appl. Genet.78: 11-15.

Sato Y, Hong SK, Tagiri A, Kitano H, Yamamoto N, Nagato Y, Matsuoka M, 1996. A rice homeobox gene, OSH1, is expressed before organ differentiation in a specific region during early embryogenesis. Proc Natl Acad Sci USA 93: 8117-22.