The QTLs, qBFR4-1 and qBFR4-2 confer field resistance to rice blast. The genetic locus controlling the resistance was mapped in a long arm region on chromosome 4 in the upland rice variety Kahei. The relationship between the level of blast field resistance and putative alleles at the qBFR4-1 and qBFR4-2 loci was analyzed using RFLP markers C600 and G271 that are the closest to each of the QTLs and by a field test for blast resistance among Japanese upland rice varieties. As a result, the upland rice varieties were divided into six varietal groups. Three major putative RFLP alleles (C600-a, C600-b and C600-c) were postulated at the C600 locus based on their fragment length, and the mean scores for blast field resistance of varieties with each allele were 2.81, 2.20 and 0.69, respectively. The C600-b allele was the most frequently observed in the Japanese upland varieties (46 %). Two major putative RFLP alleles (G271-a and G271-b) were postulated based on their fragment length, and the mean score for the resistance of varieties with each allele was 2.37 and 1.68, respectively. The varieties with the allele C600-c were the most resistant and the varieties with the alleles C600-c and G271-b showed the highest level of blast field resistance (0.58). On the other hand, the varieties with the allele C600-a and G271-a represented by the lowland variety Nipponbare showed the lowest level (2.98). These results suggest that the two QTLs for blast field resistance harbor two to three major alleles and that the levels of field resistance in upland rice varieties of Japan can be mainly assessed by the RFLP alleles tightly linked to the QTLs on chromosome 4. The RFLP markers used in this study seemed to be suitable for DNA marker-assisted selection of blast field resistance in rice breeding.

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A sweet pea (Lathyrus odoratus L.) cultivar, ‘Grace’, which has tendrils, was crossed with ‘Snoopea purple’ lacking tendrils. The resultant F₂ population was used to identify RAPD (random amplified polymorphic DNA) markers linked to a tendril gene. The presence of tendrils was found to segregate in a dominant fashion. A total of 302 random primers were used to screen a pair of bulked DNA samples of the F₂ plants. Only two primers, WB32 and WB67, showed polymorphism between the bulked samples. The former generated a DNA fragment specific to the bulked sample with tendrils, while the latter amplified a fragment in the bulked sample without tendrils. Segregation of these RAPD markers was examined in the F₂ population. One of them, WB32a was found to be linked to the tendril gene. The marker was then cloned and sequenced. A pair of primers was designed for specific amplification of this marker. The primer pairs amplified a clear and dominant band, SWB32a, and the band was specific to individuals with tendrils. The linkage between the marker, SWB32a and the gene for tendrils was demonstrated in the F₂ population in a distance of 7.7 cM. Use of this genetic marker in the breeding of sweet pea cultivars without tendrils was discussed.

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Multi-step yield screening for a population of test varieties can be performed using three different patterns; a pre-assigned upper fraction of test varieties is selected in each step (pattern I), or, test varieties that showed yield scores superior to one or a few check varieties are selected in each step (II), or, test varieties superior to a critical fixed standard are selected in each step (III). Pattern II has been commonly used in actual screening trials. By comparing the efficiencies (probability of success) of these patterns based on the Monte Carlo calculations for finite populations composed of 20, 40 or 60 test varieties, we concluded that pattern I is the most efficient. Supplementary procedures should be used to minimize the loss due to investing for undesirable populations that lack superior varieties, such as totally discarding the population if none of the test varieties showed a promising score in the first step, and continue screening in pattern I if one or more varieties showed a promising score. Screening in three steps will be efficient when the variety × year interaction is absent, but more steps may be used when a large variety × year interaction is anticipated to occur or is observed in the data in earlier steps. We conclude that patterns II and III are inefficient because they are subject to a large error due to the variety × year interaction or yearly effect.

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We developed 31 sets of melon (Cucumis melo L.) microsatellite markers in this study. Primers for these markers were designed from clones containing simple sequence repeats (SSRs) of (GA/CT)n or (GT/CA)n isolated from a melon genomic library. These microsatellite loci were examined for the detection of alleles among twelve lines and cultivars from six varieties of C. melo. A total of 28 marker loci showed polymorphism by 2.0 % agarose TBE gel separation. And these markers were also examined for cross-species amplification applicable to nine other species in Cucurbitaceae. Among them, 13 marker loci were commonly detected within the major cucurbit crops, cucumber, pumpkin and watermelon using the present primer sets. The species to which the melon microsatellite markers would be most applicable was Momordica charantia (24 markers), followed by Cucumis sativus (20 markers) and Cucurbita maxima (18 markers). These conserved microsatellite loci could be used as anchor markers in studies on synteny in Cucurbitaceae.

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Maize transcription factors for the genes of anthocyanin synthesis pathway, C1/B-peru, were delivered into developing wheat coleoptiles by a particle bombardment method. Anthocyanin pigments were induced as discrete red spots and their number reached about 60 spots per coleoptile, compared to about 20 diffused blue GUS spots, which were induced by the GusA gene transferred concomitantly. Coleoptiles of seedlings collected 36 h after germination were most suitable tissue for the expression of delivered foreign genes. One to five μg of plasmid DNA for coating gold particles was sufficient for induction of anthocyanin and GUS, indicating that the transferred gene was expressed efficiently in coleoptile cells. Helium gas pressure (900, 1100, 1300 or 1500 pounds per square inch) and distances (10, 15, or 20 cm) between the stopping screen and coleoptile did not affect significantly the efficiency of gene transfer. Seedlings arranged in a circle with a 1-cm radius on a MS medium plate were targeted well by gold particles. The results showed that the wheat coleoptile was a good target tissue for transient assay of wheat genes and that C1/B-peru can be used as a reporter gene.

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Parentage of 16 peach cultivars, including 9 cultivars bred by controlled hybridization, 2 from bud sport mutations and 5 from chance seedlings, was analyzed using 17 SSR markers. The parent-offspring relationships of 9 crossbreeding cultivars were confirmed because SSR alleles were transmitted to offspring cultivars from their parents without discrepancy at all the used loci. ‘Akatsuki’ and ‘Gyousei’ showed identical SSR genotypes, which suggested that ‘Gyousei’ was derived from a bud sport mutation. In contrast, another bud sport cultivar, ‘Hikawa Hakuhou’, showed different SSR genotypes from those of the original cultivar ‘Hakuhou’ at 12 SSR loci, indicating that ‘Hikawa Hakuhou’ was not a bud sport mutant of ‘Hakuhou’. Four cultivars that presumably originated from chance seedlings, i.e., ‘Abe Hakutou’, ‘Kawanakajima Hakutou’, ‘Kouyou Hakutou’ and ‘Shimizu Hakutou’, shared one allele with their putative parent ‘Hakutou’ at each SSR locus. SSR analysis suggested that these 4 cultivars were not bud sport mutants, but offsprings of ‘Hakutou’. It was concluded that SSR markers could be effectively utilized for parentage analysis in Japanese cultivated peaches.

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To determine the direct cause of the low seed set in sweet potato, 14 cross-compatible combinations among 11 varieties were histologically examined for the process from fertilization to embryogenesis at 4, 7 and 48 hours after pollination. Pollen fertility on the flowering day exceeded 80 % in most of the varieties examined and pollen germination 4 hours after pollination reached 10~20 % in most of the crosses. These results show that neither the fertility of mature pollen nor the pollen germination on stigma is the principal cause of the low seed set. The mean number of pollen tubes passing through the conducting tissue of the ovary 7 hours after pollination largely differed among the cross combinations. However, it did not show a close relationship with the number of seeds per capsule. Among the embryosacs examined 48 hours after pollination, those that had not been penetrated by pollen tubes were mostly abnormal, in which the development of the egg apparatus had been interrupted halfway. On the other hand, the number of normal young embryos was closely correlated with the number of seeds per capsule. These results suggest that a large number of such abnormal embryosacs is the principal cause of the low seed set of sweet potato.

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Fine mapping of Hd4 and Hd5, quantitative trait loci (QTLs) for heading date in rice, was performed by using advanced backcross progeny derived from a cross between a japonica rice variety, Nipponbare, and an indica variety, Kasalath. Hd4 was mapped between restriction fragment length polymorphism (RFLP) markers R46 and C39 in the proximal region of chromosome 7, and Hd5 was mapped between C166 and R902 on the short arm of chromosome 8; both QTLs mapped as single Mendelian factors. We used marker-assisted selection to develop two nearly isogenic lines (NIL), designated NIL(Hd4) and NIL(Hd5), in which small chromosomal segments of Kasalath including Hd4 and Hd5, respectively, each were substituted into the genetic background of Nipponbare. Compared with that of Nipponbare, days-to-heading of NIL(Hd4) and NIL(Hd5) increased under long-day and natural-field conditions, but no differences were observed between those of the two NILs and Nipponbare under short-day conditions. Epistatic interaction was detected between Hd5 and Hd1, a key photoperiod sensitivity QTL, on the basis of an analysis of the F₂ population derived from a cross between the NIL(Hd5) and NIL(Hd1). This result suggests that Hd5 is involved in photoperiod sensitivity and may act downstream or upstream of Hd1 in the same photoperiodic pathway. In comparison, the genetic effect of Hd5 was additive to that of Hd2, another key photoperiod-sensitivity QTL, indicating that Hd2 acts in a different photoperiodic pathway other than that of Hd1 and Hd5. The genetic effect of Hd4 was additive to those of Hd1 and Hd2; thus, epistatic interaction between these loci was not detected.

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An antisense gene strategy was used to suppress the expression of rice allergen (RA) genes that comprise a multigene family. We produced transgenic rice plants harboring antisense genes that consisted of cDNA for a major RA driven by promoters of the RA, rice starch branching enzyme, rice prolamine and rice glutelin genes, which are specifically expressed in developing seeds. Among more than 100 transformants, the RA content in some transformants was reduced to no more than 20 % of that in non-transformant. However, an allergen-free transformant was not obtained. Since the transcribed antisense mRNAs are not fully used for suppression of the RA genes in transformants, we examined here the possibility of a spatial or temporal difference in the expression of the introduced antisense and endogenous RA genes. However, we found no remarkable difference in the place or timing of their expression. We also examined the effect of antisense genes on each of the genes in the RA family in transformants by two-dimensional electrophoresis and immunoblotting. The results showed that the contents of the major RA and other RAs with nucleotide sequences that are highly homologous to the antisense sequence were markedly lowered, while the contents of other RAs with sequences showing relatively low homology to the major RA were hardly affected. These results suggest that the effect of the antisense gene on the RA gene family depends on the homology between the antisense sequence used and the target sequence.

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Cucumber mosaic virus (CMV) is an important plant pathogen worldwide, which infects and causes yield losses to many solanaceous crops but rarely to potatoes. In the study reported here, we have tested the susceptibility of various potato genotypes to three different CMV strains, Pf-CMV and Fny-CMV, which belong to subgroup I, and A9-CMV, a member of subgroup II. Eight potato genotypes were found that could be systemically infected by at least one of the three CMV strains. Furthermore, although most potato cultivars were resistant to systemic infection at 24°C, all became infected systemically when inoculated plants were grown at 30°C. These results suggested that the natural resistance that most potato crops express to CMV might be overcome under high-temperature growing conditions following infection, and that CMV resistance in potato showed virus strain specificity.

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A small insert genomic library of eggplant (Solanum melongena L.) was screened with eight trinucleotide probes. Based on the screening of approximately 108,000 plaques, the occurrence of (AAC/TTG) _n, (AAG/TTC)_n, (ACC/TGG)_n, (ACG/TGC)_n, (ACT/TGA)_n, (AGC/TCG) _n, (AGG/TCC)_n and (ATC/TAG) _n repeats was estimated to be, respectively, 1.2 × 10 ³, 1.3 × 10², 4.0 × 10², 7.6 × 10, 1.1 × 10², 5.7 × 10, 1.3 × 10² and 3.8 × 10, i. e. once every 951 kbp, 8.3 Mbp, 2.8 Mbp, 14.5 Mbp, 9.7 Mbp, 19.3 Mbp, 8.3 Mbp and 29 Mbp in the eggplant genome, respectively. Two trinucleotide repeats, (AAC/TTG)_n and (ACC/TGG) _n, predominated, accounting for 84.5 % of the isolated trinucleotide microsatellites. A total of 83.5 % of the identified trinucleotide repeats contained seven repeat units or less. Inserts of the positive clones were sequenced and 85 PCR primer sequences bordering the microsatellite repeats were designed. In order to assess the polymorphism, 11 S. melongena lines and 11 Solanum relatives were used. The mean number of alleles per polymorphic locus was 2.1 for S. melongena lines and 2.9 for Solanum relatives. Low levels of polymorphism were observed in eggplant, with the average heterozygosity being 0.31 between S. melongena lines and 0.32 between Solanum relatives. Most of the markers that detected polymorphism between S. melongena lines contained eight or more repeat units. The motif most frequently found in Solanum was (AAC/TTG)_n in this study. All the primer sets which detected PCR products in S. melongena yielded PCR products in S. incanum, which is in agreement with its very close relationship with S. melongena. Further transferability, however, was rather low, since only half of the primer sets gave products in Solanum relatives except for S. incanum. Analysis of the polymorphic loci in an eggplant F₂ population revealed a codominant mendelian segregation.

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