The chemical energy synthesized during photosynthesis as carbohydrates, lipids and proteins accumulates in seeds and provides a food source for animals. Seeds are also important genetic delivery systems essential for sustainable agriculture and environmental control. Genetic information in elite cultivars of crop species accumulated during breeding programs is distributed in the form of seeds. Ensuring successful germination and seedling establishment is a significant first step in agricultural production. The molecular and biochemical mechanisms of seed germination are not fully understood. Our knowledge of the interactions between the embryo, endosperm and testa has been advanced through tomato seed research, a model system for seed germination research. Recent discoveries using Arabidopsis thaliana have provided additional information about the molecular and genetic mechanisms of seed dormancy and germination. Genes expressed during seed development determine the size, shape and chemical properties of mature seeds and affect seed dormancy. In imbibed seeds, genes associated with hormone biosynthesis and degradation play critical roles in radicle emergence. The physical, chemical and physiological changes in the embryo, endosperm and testa, as well as the interactions between these tissues all contribute to successful germination. Recent literature on seed science research needs to be compiled to provide a clear picture for seed germination. Hypotheses to explain global mechanisms of seed germination are examined in this review.
Species of Zingiberaceae display a diversity in habitat, ethnobotanical use and morphology. However, little is known about the genetic relationships among taxa and genetic diversity, primarily due to the lack of suitable molecular markers. We tested the cross-amplification potential of microsatellite markers among taxa to identify a larger number of genetic markers. To assess the applicability of rice microsatellite markers to the Zingiberaceae, we tested 12 microsatellite markers for 14 genotypes from three genera of this family: Zingiber, Alpinia and Curcuma. The origin of the genotypes was diverse, covering eight Asian countries. Four microsatellite primer sets failed to amplify fragments in all genotypes studied, whereas the other primer sets amplified all the genotypes. Among the 141 bands, that could be scored, 140 (99.5%) were polymorphic. On the average, each microsatellite primer set amplified 17.6 DNA fragments. In general, amplified fragments were larger than the original rice fragments including the microsatellite region, although in some cases, the amplified bands were similar in size. Though sequence analysis of these bands confirmed the absence of target repeat motif, amplification of a large number of polymorphic bands provided the basis to perform an analysis of genetic diversity. Primers could generate enough polymorphism for possible use in diversity studies, based on provisional multivariate analyses such as cluster analysis and principal component analysis (PCA). The whole set of genotypes based on molecular data was classified into four clusters after cluster analysis. Genotypes from the Curcuma and Alpinia genera were grouped into clusters I and II, respectively. Clusters III and IV comprised genotypes from the genus Zingiber. PCA led to a similar classification. The high polymorphism documented in the present study indicated that the rice microsatellite primers were useful for genetic diversity studies among genera in the family Zingiberaceae.
Seed storage protein β-conglycinin of soybean contains major allergens in its subunits, and displays a lower amino acid score and lower functional gelling properties than glycinin. Therefore, decrease in the content of β-conglycinin is one of the objectives of soybean breeding programs. A β-conglycinin-deficient mutant QT2 was identified from a wild soybean in Kumamoto prefecture, and the phenotype was found to be controlled by a single dominant gene Scg-1 (Suppressor of β-conglycinin). Fukuyutaka and a near-isogenic line of Fukuyutaka, QY7-25 harboring the Scg-1 gene were used for development of DNA markers associated with β-conglycinin deficiency. Ten single nucleotide polymorphisms (SNPs) in the β subunit genes were detected between Fukuyutaka and QY7-25. Two β subunit gene loci were found to cosegregate with β-conglycinin deficiency with the DNA marker based on SNPs in a F2 population derived from a cross between Fukuyutaka and QY7-25. The DNA marker also enabled to detect polymorphisms between QY7-25 and major soybean cultivars and could be used as a practical tool for the introduction of Scg-1 gene into soybean. The chromosome region associated with β-conglycinin deficiency was located on linkage group I of a soybean genetic linkage map with the developed marker using a F2 population from the parents, Misuzudaizu and Moshidou Gong 503.
Somatic embryos of Glycine max (L.) Merrill cultivar ‘Jack’ were co-transformed with coat protein (CP) gene of attenuated isolates of soybean mosaic virus (SMV) and hygromycin phosphotransferase (hpt) gene by means of microprojectile bombardment. These transformed embryogenic tissues were selected in hygromycin-containing liquid medium. The hygromycin-resistant embryogenic tissues obtained through the selection were regenerated, and CP gene was detected in the 11 transgenic plants out of them. In order to assess their resistance to SMV, mechanical inoculation was performed in T1 generation. The disease symptom was examined visually and confirmed by the enzyme-linked immunosorbent assay (ELISA). Finally we obtained three independent lines highly resistant to SMV. This is the first report of the soybean plants that were conferred a high resistance to SMV by the transformation with CP gene of the SMV attenuated isolates. In these three lines, the presence of transgene transcript was confirmed by Northern blot analysis, and the transgene product was detected in two of them by Western blot analysis.
Bambara groundnut (Vigna subterranea (L.) Verdc. or Voandzeia subterranea (L.) Thouars.) is one of the most important food crops in Africa and other parts of the world. Although abundant genetic resources have been identified, genetic inheritance studies and varietal improvement could not be initiated due to unsuccessful hybridization. Since the attempts we made at applying the hybridization method proposed by Schenkel (2002) and Massawe et al. (2003) were not successful, we eventually developed a more efficient method. We report herewith a hybridization technique that yielded 23 F1 seeds from 4 crosses of morphologically distinct parental lines, TVsu 11, TVsu 870, TVsu 1061 and a Thai local variety. Emasculation method and pollination time were found to be the two critical factors that affected seed set. Emasculation by petal cutting could be performed at any time between 3 to 10 PM. However, hybridization was achieved only when pollination was conducted during the one hour period after the initiation of pollen shedding, namely at around 2:30 to 3:30 AM in Thailand. More successful crosses were obtained when the development stage of the female plants corresponded to peduncle elongation. Successful hybridization was confirmed by growing out the F1 seeds and observing their phenotypic hybrid traits.
To elucidate the effects of different glutenin subunit alleles on the parameters of wheat flour quality, we analyzed the dough and gluten properties using a doubled-haploid population derived from a cross between two wheat cultivars differing in the glutenin subunit loci except for Glu-D3, namely Grandin (Glu-A1b, Glu-B1c, Glu-D1d, Glu-A3f, Glu-B3h and Glu-D3a) and Kitamiharu 57 (Glu-A1a, Glu-B1i, Glu-D1a, Glu-A3c, Glu-B3b and Glu-D3a). Based on the analysis of the DH lines, no significant differences in the flour protein content were detected among the glutenin alleles. However, the Glu-D1 and Glu-B3 alleles exerted significant effects on the dough and gluten properties. The DH lines with the Glu-D1d or Glu-B3b alleles showed a significantly higher gluten index, determined using the Glutomatic system and longer dough development time determined using a mixograph. The Glu-D1d and Glu-B3b alleles additively increased the dough and gluten strength. The DH lines with the Glu-A1a allele also showed a significantly higher gluten index and longer dough development time than those with the Glu-A1b allele, though to a lesser extent. These results indicated that the Glu-B3b allele exerted similar effects on the dough and gluten strength as the Glu-D1d allele. Since the DH lines harboring the Glu-D1d and Glu-B3b alleles showed excessive values for the strength of dough and gluten, it was suggested that both the Glu-1 and Glu-3 allelic composition should be considered to improve the wheat flour quality.
A doubled-haploid (DH) population (n = 176) obtained by anther culture of an F1 hybrid between a line susceptible to Phytophthora capsici ‘K9-11’ (Capsicum annuum L.) and a line resistant to P. capsici ‘AC2258’ (C. annuum L.) was inoculated with P. capsici. QTL analysis of the resistance was performed using a linkage map consisting of 16 linkage groups (LGs), covering a total distance of 1100.5 cM. Three QTLs were detected on LG1, LG6 and LG7. The QTL with the highest LOD score, detected on LG7, explained 82.7% of the phenotypic variance with a LOD score of 67.02. This QTL was designated as Phyt-1. The nearest marker was an AFLP marker, M10E3-6. The second QTL, designated as Phyt-2, was found on LG1. It explained 6.4% of the phenotypic variance with a LOD score of 2.54. The nearest RAPD marker was RP13-1. The other QTL, designated as Phyt-3, which was found on LG6, explained 5.6% of the phenotypic variance with a LOD score of 2.20. The nearest AFLP marker was M9E3-11. It was confirmed that the lines with a high resistance could be efficiently selected by using two markers, M10E3-6 and RP13-1, simultaneously. The presence of both Phyt-1 and Phyt-2 under homozygous conditions may enable to breed resistant cultivars of sweet pepper. The molecular markers identified in the present study could be useful for marker-assisted selection (MAS) in order to breed sweet pepper cultivars with a high resistance to P. capsici using ‘AC2258’ as a source of resistance genes.
To clarify the effect of heterosis for the earliness of head formation in cabbage, we applied the diallel analysis to four developmental characteristics that were highly correlated with the earliness of head formation: leaf position at which the head formation started (LPH), length of the 5th wrapper leaf (L5), width of the 15th wrapper leaf (W15) and leaf shape index (width/length) of the 15th wrapper leaf (LSI15). LPH showed large additive and average dominant effects, high broad and narrow sense heritabilities, and incomplete dominance directed to low. Both L5 and W15 showed remarkably large dominance effect, low narrow sense heritability, and overdominance directed to long and wide. LSI15 showed large additive effect, comparatively high broad and narrow sense heritabilities, and incomplete dominance. From these results, we discussed how to raise the early head forming F1 hybrid cabbage by taking into account the mode of inheritance of these developmental characteristics.
Soybean cyst nematode (SCN) (Heterodera glycines Ichinohe) is one of the most economically damaging pests of soybean. The damage can only be effectively controlled through the use of resistant soybean [Glycine max (L.) Merr.] cultivars. Since QTL mapping is a highly effective approach for studying genetically complex forms of plant resistance to pests, it should be used to identify and map QTLs for resistance to SCN from various genetic backgrounds. The objective of the present study was to identify closely linked DNA markers to QTLs for resistance to SCN race 3 in the cultivar Toyomusume derived from a resistant cultivar Gedenshirazu, using the AFLP (amplified fragment length polymorphism) method combined with bulked segregant analysis (BSA). A population of 115 recombinant inbred lines (RILs) derived from a cross of Toyomusume (resistant) and Tsurukogane (susceptible) was used to map QTLs related to the resistance to SCN race 3. After genotyping of all the RILs with molecular markers including AFLP, RFLP, SSR and RAPD markers, composite interval mapping was performed for QTL analysis. In addition to known rhg1 on the linkage group (LG) G, two significant QTLs, rhg-t1 and rhg-t2, on the linkage group B1 near the AFLP markers CAT_CTC1 and GTC_ATT, respectively, were identified using two experimental assays. Epistatic analysis suggested that the combination of rhg1 and rhg-t1 could provide a higher level of resistance to SCN race 3, compared to any single QTL. The AFLP markers surrounding rhg1 and rhg-t1 could be used for marker-assisted selection of SCN resistance derived from Gedenshirazu, when converted to PCR-based markers.
A DNA profiling method was developed for fresh and processed fruits in pear. Four different DNA extraction methods were examined to isolate genomic DNAs from fresh pear fruits, i.e., CTAB-based method and methods based on 3 kinds of DNA extraction kits (DNeasy Plant Mini Kit, G2 buffer & Genomic-tip20/G, Nucleon Phytopure Plant DNA Extraction Kit). All the methods enabled to recover genomic DNAs from samples of fresh fruits with such a high quality that cultivar identification could be performed based on SSR markers. Among them, the G2 buffer & Genomic-tip20/G method gave the best results for samples of fresh as well as processed fruits. Partially degraded genomic DNAs that were isolated from samples of dried fruits could be amplified by all the tested SSR markers. SSR analysis revealed that genotypes from dried fruits were identical with those of the European pear cultivar ‘Bartlett’, indicating that cultivar identification could be successfully performed. Severely degraded genomic DNAs less than 500 bp in size were recovered from samples of canned fruits and fruit juice. The amount and quality of the extracted DNAs were sufficient to enable amplification by primers corresponding to high copy rDNA sequences, whereas no bands were produced by primers of chloroplast DNA sequences. Out of 15 SSR loci, 9 SSRs with target sequences less than 150–160 bp could successfully amplify fragments for genomic DNAs from samples of canned fruits and fruit juice. In contrast, no amplified fragments were observed for the remaining 6 SSRs with longer target sequences. DNA profiling and cultivar identification were successfully performed by using SSR markers to amplify short target sequences less than 150 bp.
A collection of 418 Spanish barley accessions was screened for resistance to leaf rust in the field at Córdoba, Spain during the 2002–2003 season. Six accessions displaying the lowest disease severity with no macroscopically visible necrosis were selected for further studies on components of resistance. Five of them showed a significantly higher relative latency period than the susceptible line L94 and were similar to the partially resistant Vada. All of them showed lower relative infection frequency and smaller colony size than the susceptible L94 under controlled conditions. Histological studies indicated that the resistance in four of these accessions was based on a higher percentage of early aborted colonies not associated with host cell necrosis. In the remaining two accessions, resistance was based on hypersensitivity.
Selection of mutants by the reverse genetic approach is a useful tool for gene function analysis and crop improvement. TILLING (Targeting Induced Local Lesions in Genomes), in which mutants are selected from the progeny of EMS-treated plants by the SNP analysis using heteroduplex cleavage enzyme from celery, has been applied to various plant species. Since gamma-ray-induced point mutations have not been identified by the reverse genetic approach, we attempted to select mutants from a population of the gamma-ray-irradiated rice. A heteroduplex between wild-type DNA and mutant DNA was cleaved by an endonuclease extracted from petioles of Brassica rapa, and cleaved DNA was detected using agarose gel electrophoresis. M2 plants derived from 2,130 M1 plants irradiated by gamma rays were used for the screening of mutants having nucleotide changes in 25 regions from 1.0 kb to 1.5 kb. Six mutations were identified and the rate of mutation induced by gamma rays was estimated to be one mutation per 6,190 kb. Four of the mutations were single nucleotide substitutions and two were 2-bp and 4-bp deletions. These results suggest that the point mutation rate with gamma rays was quite lower than that with EMS, but that the rate of knockout mutations among detected mutations generated by the gamma-ray irradiation was higher than that by the EMS treatment.