8. Cytogenetic and molecular characterization of an intergeneric hybrid between Oryza sativa L. and Porteresia coarctata (Roxb.)Tateoka

D.S. BRAR, R.M. ELLORAN, J.D. TALAG, F. ABBASI and G.S. KHUCH

International Rice Research Institute, P.O. Box 933, 1099 Manila, Philippines

The genus Oryza has two cultivated species and 22 wild species. A number of genes for resistance to biotic stresses have been transferred from wild species into the cultivated rice through embryo rescue and backcrossing procedures (Brar and Khush 1997). The wild perennial grass species Porteresia coarctata Tateoka (formerly O. coarctata Roxb., 2n = 48) is considered as a potential source of genes for salinity tolerance.

We report the successful production of intergeneric hybrid (O. sativa x P. coarctata) through embryo rescue. Panicles of O. sativa cv. IR56 were emasculated and pollinated with pollen of P. coarctata. Immature embryos after 10-14 days of pollinations were excised and cultured on the nutrient medium to obtain the hybrid seedling. Isozyme analysis was used to characterize the F1 hybrid. Meiotic chromosome analysis was carried out from anthers squashed in 2% acetocarmine. Total genomic DNA of O. sativa and P. coarctata was individually used as a probe. EcoRI digested DNA from both the species was used in DNA hybridization to determine the molecular divergence and nature of the F1 hybrid.

The hybrid is slow growing and is intermediate in morphological characteristics (Fig. 1 ). It is completely male sterile. Jena (1994) also reported a hybrid between (O. sativa and P. coarctata. Starch gel electrophoresis showed IR56 and P. coarctata to be highly polymorphic. Out of 20 loci, Amp1, Amp2, Amp3, Amp4, Pgi1, Pgi2, Pgd1, Pgd2, Adh1, Sdh1, Mal1, Got1, Got3, Mdh1, Mdh2, Mdh3, Est1, Est2, Est5, Est9 examined, 16 were polymorphic. The F1 showed isozyme bands from both parents indicating hybrid nature. Meiotic chromosome analysis of the hybrid showed 36 univalents at metaphase I indicating lack of pairing between chromosomes of O. sativa and P. coarctata (Fig. 2).

Hybridization of P. coarctata DNA when used as a probe showed strong hybridization with P. coarctata and limited cross hybridization with IR56 indicating strong diver-gence between the genomes of O. sativa and P. coarctata (Fig. 3). Similar divergence was observed when IR56 DNA was used as a probe. Total genomic DNA hybridization approach has been used to determine divergence between various species of Oryza (Aggarwal et al. 1997). Cytogenetic and molecular analysis confirmed true hybrid nature of the F1 and showed limited homoeology between (9. sativa and P. coarctata genomes.

We have vegetatively multiplied the F1 hybrid. These hybrid plants are being used in backcrossing with IR56 for the transfer of genes for salinity tolerance from P. coarctata into rice.

References

Aggarwal, R.K., D.S. Brar and G.S. Khush, 1997. Two new genomes in the Oryza complex, identified on the basis of molecular divergence analysis using total genomic DNA hybridization. Mol. Gen. Genet. 254:1-12.

Brar, D.S. and G.S. Khush, 1997. Alien introgression in rice. Plant Molecular Biology 35: 35-47.

Jena, K.K., 1994. Development of intergeneric hybrid between O. sativa and P. coarctata. RGN11 : 78-79.

Figs. 1-3: 1. Intergeneric hybrid between Oryza sativa (2n = 24) and Porteresia coarctata (2n = 48) produced through embryo rescue: 2. Meiotic metaphase showing 36 univalents in the F1 hybrid indicating lack of homoeology between chromosomes of O. sativa and P. coarctata; 3. Hybridization after probing with genomic DNA of P. coarctata (A) and IR56 (B). Lane 1-molecular marker, 2-O. sativa cv. IR56; 3- F1 hybrid; 4-P. coarctata. (A) showing strong hybridization signal after probing with genomic DNA of P. coarctata (lane 4) and very low cross hybridization with DNA of IR56 (lane 2). B-strong hybridization signal after probing with DNA of IR56 (lane 2) and limited hybridization with DNA of P. coarctata (lane 4).