The wild rice species Oryza rufipogon displays wide intraspecific variations. It is considered to be the progenitor of the cultivated rice species Oryza sativa with two ecotypes, japonica and indica. p-SINE1 is the first plant retroposon SINE identified in O. sativa and is present in strains of Oryza species with the AA genome in multiple copies. Some p-SINE1 members show interspecific insertion polymorphism among the rice strains and are thus assumed to be suitable markers for classifying the strains and to infer their relationships based on the presence or absence of the members at the corresponding loci. Subfamily members of p-SINE1 show insertion polymorphism in the O. sativa -O. rufipogon population. To determine the origin of cultivated rice, we used these p-SINE1 members to bar-code each of 101 cultivated and wild rice strains based on the insertion polymorphism at the respective loci. A phylogenetic tree constructed using the bar codes given to all the rice strains showed that O. sativa strains could be divided into two groups corresponding to japonica and indica, whereas O. rufipogon strains were divided into four groups, of which, one group consisted of annual O. rufipogon strains, while the other three of the perennial O. rufipogon strains. Japonica strains were closely related to the O. rufipogon perennial strains of one group, and the indica strains to the O. rufipogon annual strains, indicating that O. sativa had been derived polyphyletically from O. rufipogon. Furthermore, we identified new p-SINE1 members showing interspecific insertion polymorphism from representative strains of four wild rice species with the AA genome, O. barthii, O. glumaepatula, O. longistaminata and O. meridionalis. A phylogenetic tree constructed using the polymorphic p-SINE1 members showed that O. longistaminata and O. meridionalis had diverged early, whereas the other species including O. rufipogon diverged relatively recently in an evolutionary time. We also presented the results of mapping of all p-SINE1 members identified from databases of rice genome sequences, and showed that they were present in all the chromosomes, but not distributed in the regions near the centromere in each chromosome, unlike other retroelements.
The mechanism of SDR 2n-pollen formation was analyzed in two intra-sectional diploid (2n = 2x = 24) Lilium hybrids (Enchantment × L. pumilum). Variable frequencies of 2n-pollen were found. Meiotic analysis indicated that the intra-sectional hybrids showed perfect chromosome pairing in most cases at metaphase I and normal anaphase I movement of pollen mother cells (PMCs), but produced 2n-pollen by second division restitution (SDR). A high bivalent formation (11.9II and 11.8II, respectively) at metaphase I, irregular meiotic division such as unbalanced chromosome separation and chromatic fragmentation resulted yet in acceptable pollen fertility for cross-pollination. The hybrids were fertile, and when used as male parents, offspring could be generated. The significance of the occurrence of 2n-pollen for the breeding of lilies was analyzed.
Many Japanese upland rice varieties show a high level of resistance to rice stripe virus (RSV). This resistance is thought to be controlled by a pair of complementary dominant genes, Stva and Stvb. Quantitative trait locus (QTL) analysis for the rice stripe virus resistance was performed using 120 F2 plants/ F 3 lines derived from a cross between the susceptible variety Nipponbare, and the resistant line Upland Rice Kanto72 (URK72). As a result, two QTLs were detected on chromosomes 2 and 11. The chromosomal regions of the two QTLs were introgressed into two resistant paddy lines Chugoku40 and Chugoku41, which were developed by introducing resistance genes from URK72. In order to verify the two QTLs, we selected two informative plants NR8 (heterozygous at the QTL on chromosome 11) and NR23 (heterozygous at the QTL on chromosome 2), from F2 plants and we used the self-pollinated progenies of NR23 (99 plants) and NR8 (96 plants) for further QTL analyses. In these secondary analyses, one QTL was mapped near the RFLP marker G257 on chromosome 11 and the other QTL was mapped near SSR marker MS-11 on chromosome 2. Since the Stvb gene is allelic with the other RSV resistance gene Stvb-i, mapped on chromosome 11, the QTL detected on chromosome 11 was thought to correspond to the Stvb gene. However, this QTL seemed to have a larger effect on RSV resistance than previously reported and was found to contribute to the suppression of RSV infection. On the other hand, since the QTL detected on chromosome 2 did not suppress the RSV infection, it probably enhanced the QTL on chromosome 11 and suppressed the symptoms after infection by RSV.
The common cutworm (Spodoptera litura Fabricius; Lepidoptera: Noctuidae) is a menace to soybean (Glycine max (L.) Merr.) production in southwestern Japan. We have been evaluating soybean germplasm for resistance to common cutworm in order to develop resistant cultivars and have found a cultivar named ‘Himeshirazu’, which is distinguished by its high level of resistance. We compared the antibiosis of Himeshirazu with those of ‘Fukuyutaka’ (susceptible) and ‘Sodendaizu’ (resistant). Himeshirazu depressed the weights of individual common cutworm larvae and prolonged the duration of the instar stage, compared with the other two cultivars. We analyzed the inheritance of this antibiosis in an F2 population derived from a cross between Fukuyutaka and Himeshirazu. The broad-sense heritability of the antibiosis was estimated as 71.3 %. The segregation pattern in the F2 progeny suggested that a recessive factor controlled a considerable part of the antibiosis. From the relationship between the antibiosis and the genotype of a simple sequence repeat marker, Satt220, in the F2 progeny, we inferred that the locus of the putative recessive factor was located on linkage group M.
Fruit color of pungent pepper (Capsicum annuum L.) is a trait of economic importance in pepper breeding. To develop a system of molecular marker-assisted selection (MAS) for breeding pepper varieties with various fruit colors, we conducted molecular genetic analyses on six genes involved in the carotenoid biosynthesis pathway. Capsanthin-capsorubin synthase (CCS) gene showed a polymorphism in the PCR pattern in the segregation population derived from a cross between a pepper accession (cv. msGTY-1) with orange fruits and a pepper accession (cv. 277long) with red fruits. A deletion was found in the upstream region of the CCS gene in the plants with orange fruits. Southern hybridization analysis and sequencing analysis indicated that 211-bp of the downstream region of the gene was conserved in the plants with orange fruits, while no transcript of the CCS gene was detected by RT-PCR in the mature orange fruits. Carotenoid composition analysis using the Thin layer chromatography (TLC) method showed that one of the major pepper carotenoids, capsanthin, was present in the red fruits, but not in the orange fruits. The PCR polymorphism of the CCS gene and TLC pattern of carotenoid composition were completely cosegregated with the fruit color in the F2 population, suggesting that the CCS gene determines the fruit color by changing the carotenoid composition.
Polymorphisms of β-amylase among 19 species (27 taxa, 337 accessions) of wild barley, including Hordeum vulgare ssp. spontaneum (174 accessions), H. bulbosum (33), H. murinum (81), H. marinum (28), H. brachyantherum (4), H. jubatum (2), H. chilense (2) and H. roshevitzii (2) were studied using both isoelectric focusing (IEF) and thermostability analysis. Wide genetic variations were found. In general, the IEF patterns of H. vulgare ssp. spontaneum were markedly different from those of other wild species. Two new β-amylase IEF patterns (Ia and III) were observed in H. vulgare ssp. spontaneum in addition to the two patterns (I and II) found in cultivated barley (H. vulgare ssp. vulgare). In H. bulbosum, H. murinum, H. marinum and other wild species, 21 new IEF patterns were observed. Besides the A, B and C thermostability types reported in cultivated barley, new thermostability types (A+, A-B, B-C and C-) were frequently observed. Some accessions from H. arizonicum, H. jubatum, H. depressum and H. brachyantherum showed superior thermostability (A+). The genetic differentiation of β-amylase in relation to the phylogeny of genus Hordeum is also discussed.
We investigated the usefulness of the promoter of a gene for chrysanthemum chlorophyll-a/b-binding protein (Cab) for transgene expression in the chrysanthemum Dendranthema grandiflorum (Ramat.) Kitamura. We used the promoter region of a Cab gene isolated from the chrysanthemum-wild species D. japonicum Makino. The 35S promoter of cauliflower mosaic virus (CaMV) or the Cab promoter was fused to the β-glucuronidase gene (gus) and introduced into the chrysanthemum. We obtained 300 putative transformants (115 with 35S/gus and 185 with Cab/gus). GUS assay of the leaves of the in vitro plants revealed that 9.6 % (11/115) of the putative plants to which 35S/gus had been introduced and 24.3 % (45/185) of the putative plants to which Cab/gus had been introduced were GUS-positive. Southern blot analysis showed that even the GUS-negative plants harbored the gus gene in their genomes. The Cab promoter expressed the transgene more efficiently than the 35S promoter and could be used for transgene expression in chrysanthemum leaf tissues.
Hybrid seedlings from the cross Nicotiana tabacum ×N. suaveolens, obtained by test-tube pollination and ovule culture, expressed lethality at 28°C. Characteristic lethal symptoms in these hybrid seedlings consisted of browning of hypocotyls and roots. One hundred and seventeen hybrid seedlings were eventually obtained by the use of test-tube pollination and ovule culture. Hybrid seedlings maintained at 36°C did not express any lethal symptoms. Hybrid seedlings used for further experiments were transferred to 36°C immediately after germination at 28°C. When hybrid seedlings cultured at 36°C were transferred to 28°C, their growth stopped and lethal symptoms were expressed. During the progressive expression of lethality, apoptotic features such as chromatin condensation, nuclear fragmentation and DNA fragmentation were detected. On the other hand, there was no sign of apoptotic cell death in the hybrid seedlings at 36°C. Based on the observation that the same lethal symptoms and the same apoptotic features were observed in the reciprocal cross, N. suaveolens ×N. tabacum, we suggest that not only the underlying causes of hybrid lethality but also the underlying causes of apoptotic cell death are due to the interaction of coexisting heterogeneous genomes, rather than to the effect of cytoplasmic genes. Furthermore, the progression of apoptotic cell death in the cross N. tabacum × N. suaveolens began in stems and roots, followed by leaves.
After the completion of the genome sequencing project of common rice (Oryza sativa L.), comparative genomic studies between rice and related species became important to reveal the function of each gene. The rice genome contains two copies of the gene encoding zeta class glutathione S-transferase (GSTZ) that is reported to be the enzyme in the catabolic pathway of tyrosine and phenylalanine. Two GSTZ genes of O. sativa, OsGSTZ1 and OsGSTZ2, display a tandem arrangement. Upstream OsGSTZ1 gene is constitutively expressed, whereas the downstream OsGSTZ2 gene is inducible by stresses. We analyzed the expression of the GSTZ gene in the African cultivated species O. glaberrima and wild species O. longistaminata by using RT-PCR. The results showed that both GSTZ1 and GSTZ2 genes were expressed in O. longistaminata, whereas only the cDNA fragment of downstream GSTZ2 was detected in O. glaberrima. Thus, the genomic sequence of the GSTZ1 locus of O. glaberrima was determined by PCR genomic walking. Sequence comparison between O. sativa and O. glaberrima revealed that the genomic sequence upstream from the eighth intron of the OgGSTZ1 gene was highly homologous, in inverted orientation, to a BAC clone, over 1-Mb apart from the OsGSTZ1 gene in O. sativa. This result suggested that the OgGSTZ1 gene was disrupted by a large rearrangement or inversion. The genetic alteration was also observed in several lines of O. glaberrima and its ancestral wild species O. barthii. The loss of the OgGSTZ1 gene in O. glaberrima is important, as the expression of the gene is 12-fold higher than that of OsGSTZ2 in O. sativa and since O. sativa and O. glaberrima differ in several such as seed yield and annual/perennial habitat that might be related to the translocation of nitrogen metabolites from leaves to seeds.