Life histories and breeding systems strongly affect the genetic diversity of seed plants, but the genetic architectures that promote outcrossing in Oryza longistaminata, a perennial wild species in Africa, are not understood. We conducted a genetic analysis of the anther length of O. longistaminata accession W1508 using advanced backcross quantitative trait locus (QTL) analysis and chromosomal segment substitution lines (CSSLs) in the genetic background of O. sativa Taichung 65 (T65), with simple sequence repeat markers. QTL analysis of the BC3F1 population (n = 100) revealed that four main QTL regions on chromosomes 3, 5, and 6 were associated to anther length. We selected a minimum set of BC3F2 plants for the development of CSSLs to cover as much of the W1508 genome as possible. The additional minor QTLs were suggested in the regional QTL analysis, using 21 to 24 plants in each of the selected BC3F2 population. The main QTLs found on chromosomes 3, 5, and 6 were validated and designated qATL3, qATL5, qATL6.1, and qATL6.2, as novel QTLs identified in O. longistaminata in the mapping populations of 94, 88, 70, and 95 BC3F4 plants. qATL3, qATL5, and qATL6.1 likely contributed to anther length by cell elongation, whereas qATL6.2 likely contributed by cell multiplication. The QTLs were confirmed again in an evaluation of the W1508ILs. In several chromosome segment substitution lines without the four validated QTLs, the anthers were also longer than those of T65, suggesting that other QTLs also increase anther length in W1508. The cloning and diversity analyses of genes conferring anther length QTLs promotes utilization of the genetic resources of wild species, and the understanding of haplotype evolution on the differentiation of annuality and perenniality in the genus Oryza.
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