Characterization of Hexose Transporter for Facilitated Diffusion of the Tonoplast Vesicles from Pear Fruit

Tonoplast vesicles were prepared from the flesh tissue of maturepear fruit. Sugar uptakes into the vesicles determined by twodifferent methods, the membrane and the gel filtration methods,were quite similar. The uptake was highest for glucose and subsequently,in order, for fructose, sucrose and sorbitol. It was not stimulatedby addition of ATP, although the vesicles could create a protongradient. However, the uptakes were significantly inhibitedby p-chloromercuribenzene sulphonate (PCMBS, SH-reagent andinhibitor of sugar transporter). Further, the PCMBS-sensitiveuptakes of glucose and fructose saturated with their increasedconcentrations. Thus, these PCMBS-sensitive uptakes are mediatedby the transporter of facilitated diffusion. The uptakes ofglucose or fructose each had two K_m values. K_m values for glucosewere 0.35 and 18 mM, and those for fructose were 1.6 and 25raM. The uptake of 0.2 mM glucose was inhibited by 2 mM fructoseand that of 2 mM fructose was inhibited by 2 mM glucose, butneither was inhibited by sucrose or sorbitol. O-methyl-glucose(OMG) also inhibited both the glucose and fructose uptakes.Therefore, the same transporter may mediate both glucose andfructose uptakes at lower concentrations; this hexose transportsystem differed from the sucrose and sorbitol transport systems. ¹Research Fellow of the Japan Society for the Promotion of Science. ²Present address: Faculty of Agriculture, Tohoku University,1-1 Tsutsumidori-Amamiyamachi, Aoba-ku, Sendai, 981 Japan.     

Ultrastructure of the Mesocarp of Mature Avocado Fruit and Changes Associated with Ripening

The mesocarp tissue of ripening avocado fruits was studied byfreeze fracture, thin section and scanning electron microscopy.Carbon dioxide and ethylene production by individual fruit weremonitored, and samples were analysed at several stages of theripening process. The tissue is composed primarily of large, isodiametric, lipid-containingparenchyma cells. At maturity these cells contain the normalcomplement of cell organelles, and all membranes appear intact.When ripening begins, several changes in the ultrastructureoccur. The most obvious changes are a loosening and eventualbreakdown of the cell wall, and swelling and vesiculation ofthe rough endoplasmic reticulum. In freeze fracture replicasa significant increase in the number of intramembranous particlesin the EF face of the plasmamembrane was observed at the climactericpeak. In post-climacteric, soft fruit the particle density of theEF face of the plasmamembrane decreased to the density observedin the membrane of pre-climacteric cells. All of the organellesand membranes appear whole and intact whether examined by thinsection, freeze fracture or scanning electron microscopy. However,the cell walls in post-climacteric fruit have almost completelydisappeared. These results indicated that the ripening process per se inavocados does not involve a complete loss of compartmentationnor a breakdown of organelle and membrane integrity. It may,however, lead to these or similar senescence changes as a resultof the loss of the cell walls. The variations in particle densityof the plasmamembrane during ripening may reflect one or moreof several structural, compositional, or functional membranephenomena, and this aspect of ripening warrants further study. Persea americana Mill., avocado pear, freeze fracture, fruit ripening, scanning electron microscopy, senescence, ultrastructure     

Bioinformatics-Based Identification of Candidate Genes from QTLs Associated with Cell Wall Traits in Populus

Quantitative trait locus (QTL) studies are an integral part of plant research and are used to characterize the genetic basis of phenotypic variation observed in structured populations and inform marker-assisted breeding efforts. These QTL intervals can span large physical regions on a chromosome comprising hundreds of genes, thereby hampering candidate gene identification. Genome history, evolution, and expression evidence can be used to narrow the genes in the interval to a smaller list that is manageable for detailed downstream functional genomics characterization. Our primary motivation for the present study was to address the need for a research methodology that identifies candidate genes within a broad QTL interval. Here we present a bioinformatics-based approach for subdividing candidate genes within QTL intervals into alternate groups of high probability candidates. Application of this approach in the context of studying cell wall traits, specifically lignin content and S/G ratios of stem and root in Populus plants, resulted in manageable sets of genes of both known and putative cell wall biosynthetic function. These results provide a roadmap for future experimental work leading to identification of new genes controlling cell wall recalcitrance and, ultimately, in the utility of plant biomass as an energy feedstock.     

Rapid Identification of Major QTLs Associated with Rice Grain Weight and Their Utilization

To uncover the genetics of rice grain weight, we constructed an RIL population derived from a cross between a large grain accession M201 and a small size variety JY293. Specific Locus Amplified Fragment Sequencing (SLAF-Seq) technology was used to genotype two bulked DNA pools made from individual DNA of the heaviest 30 lines and the lightest 30 lines according to the 1000 grain weight (TGW). Bulked segregant analysis (BSA) was used to identify SLAFs strongly associated with TGW. Two marker-intensive regions at 24,600,000–24,850,000 bp and 25,000,000–25,350,000 bp on chromosome 3 were identified tightly related to the TGW. Then a linkage map of chromosome 3 was constructed with SSR markers and some SLAF derived single nucleotide polymorphisms (SNPs). Quantitative trait locus (QTL) mapping for TGW, grain length, grain width, and grain thickness revealed one major QTL in the second hot-region and two other minor QTLs for grain weight. These three QTLs displayed hierarchical effects on grain length and grain weight in order of qTGW3.2 (qGL3) qTGW3.1 (GS3) qTGW3.3. Multiple comparisons of means among the eight combinations of 3 QTLs revealed that the lines with two of three QTLs deriving from M201 displayed a large grain weight phenotype (TGW 40.2g, average data of three years) and lines with both qTGW3.1 and qTGW3.3 alleles from M201 (42.5g) had similar grain weight to the qTGW3.2 (40.8g) alone. Two strategies with similar effectiveness were proposed to improve grain weight by marker-assisted selection (MAS). One is to introduce the novel qTGW3.2 allele alone, and the other is to pyramid qTGW3.1 and qTGW3.3 alleles together. One new allele of GS3 (39 bp deletion in intron 1) and two SNPs in coding sequence of qGL3 identified in this study from M201 are useful in pyramiding elite alleles for molecular breeding for improvement of rice yield.     

An AFLP, SRAP, and SSR Genetic Linkage Map and Identification of QTLs for Fruit Traits in Pear (Pyrus L.)

In this study, a population of 97 F₁ seedlings from a cross between the interspecific hybrid (European × Chinese species) pear ‘Bayuehong’ (BYH) and the Chinese pear ‘Dangshansuli’ (DS) was used for establishing linkage maps and for quantitative trait loci (QTL) discovery. Using amplified length polymorphism (AFLP), simple sequence repeat (SSR), and sequence-related amplified polymorphism (SRAP) markers, along with the S locus for self-incompatibility, two parental linkage maps were constructed. The map of BYH consisted of 214 markers (143 AFLPs, 64 SRAPs, 6 SSRs, and S) mapped on all 17 linkage groups of the pear genome with a total length of 1,352.7 cM. The map of DS was comprised of 122 markers (83 AFLPs, 37 SRAPs, 1 SSR, and S) distributed along all 17 linkage groups and covering 1,044.3 cM. Based on phenotypic data from two successive years (2007 and 2008) for six fruit traits, including fruit weight (in grams), fruit diameter (in centimeters), fruit length (in centimeters), soluble solids content, fruit shape index, and maturity date, 19 QTLs were detected. These QTLs were mapped on LG 01, LG 02, LG 05, LG 07, LG 08, LG 10 of the BYH map and LG 02, LG 06, LG 15 of the DS map and accounting for 7.1 to 22.0 % of the observed phenotypic variance. Four QTLs, Pfi-8-1 for fruit shape index, Pfm-8-1 for fruit maturity date, Pfw-7-1 and Pfw-8-1 for fruit weight (in grams), with LOD scores ≥3.5, were deemed as major genes. QTLs Pfi-8-1, Pfm-8-1, and Pfw-8-1 were co-localized on LG 08 of the BYH map, along with Pfl-8-1 for fruit length. It was observed that on LG 07 of the BYH map, QTLs for fruit length, fruit shape index, and fruit weight were clustered. When QTL locations from both years were compared, Pfl-7-1 and Pfl-7-2 for fruit length, Pfi-2-1 and Pfi-2-2 for fruit shape index, and Pfm-8-1 and Pfm-8-2 for fruit maturity date were stably mapped onto the same linkage groups, respectively. Moreover, Pfm-8-1 and Pfm-8-2 were also located within the same region of LG 08 of the BYH map.     

Identification of Fungal Species Associated with Cladode Spot of Prickly Pear and Their Sensitivity to Chitosan

México is the most important producer of prickly pear (Opuntia ficus‐indica) in the world. There are several fungal diseases that can have a negative impact on their yields. In this study, there was a widespread fungal richness on cladodes spot of prickly pears from México. A total of 41 fungi isolates were obtained from cladodes spot; 11 of them were morphologically different. According to the pathogenicity test, seven isolates caused lesions on cladodes. The morphological and molecular identification evidenced the isolation of Colletotrichum gloeosporioides, Alternaria alternata, Fusarium lunatum, Curvularia lunata. All these species caused similar symptoms of circular cladodes spot. However, it is noticeable that some lesions showed perforation and detachment of affected tissues by Fusarium lunatum. To our knowledge, this is the first report of the Fusarium lunatum as phytopathogenic fungus of cladodes of prickly pear. The chitosan inhibited the mycelium growth in the seven isolates of phytopathogenic fungi. Chitosan applications diminished the disease incidence caused by C. gloeosporioies and F. lunatum in 40 and 100%, respectively. Likewise, the lesion severity index in cladodes decreased. There are no previous reports about the application of chitosan on cladodes of prickly pears for the control of phytopathogenic fungi. Therefore, this research could contribute to improve the strategies for the management of diseases in prickly pear.     

The Conversion of Sucrose to Starch in Developing Fruits

Although the flow of carbon from sites where it is assimilated to its eventual destination as starch is, for convenience, investigated as a large number of partial processes, it is nevertheless possible to conceive of a simple unifying model: the plant as a set of conductors for reduced carbon compounds, the direction of flow and the flux being determined by gradients of concentration and resistance to movement (Beevers 1969, Jenner 1974 a). This simple idea can explain what is known about the overall process, and one attractive feature of a recent hypothesis on the nature of the regulatory mechanism (Herold and Walker 1979) is that it spans the gap between origin and destination.     

Identification of Quantitative Trait Loci (QTLs) for Fruit Quality Traits in Apple

A segregating mapping population of “Co-op 17” × “Co-op 16” was used to identify quantitative trait loci (QTLs) associated with various fruit quality traits in apple. Phenotypic data were collected over 2 years for fruit circumference (in centimeter), diameter at midpoint (in centimeter), length (in centimeter), weight (in gram), total soluble solids (in degree Brix), and total titratable acids (in percent) for the segregating population. The phenotypic data along with a previously constructed genetic map, based on simple sequence repeat markers derived from expressed sequence tag and bacterial artificial chromosome end sequence databases, were used in marker–trait association analysis. Interval mapping identified two QTLs linked to fruit size components on linkage groups 03 and 05 with limit of detection scores of 3.27–4.06 and 3.29–4.02 along with phenotypic variation accounting for 15.4–46.4 and 18.3–21.9 %, respectively.     

A genome scan with AFLP markers to detect fearfulness-related QTLs in Japanese quail

A quantitative trait loci (QTL) study was undertaken to identify genome regions involved in the control of fearfulness in Japanese quail (Coturnix japonica). An F2 cross was made between two quail lines divergently selected over 29 generations on duration of tonic immobility (DTI), a catatonic-like state of reduced responsiveness to a stressful stimulation. A total of 1065 animals were measured for the logarithm of DTI (LOGTI), the number of inductions (NI) necessary to induce the immobility reaction, open-field behaviour including locomotor activity (MOVE), latency before first movement (LAT), number of jumps (JUMP), dejections (DEJ) and shouts (SHOUT), corticosterone level after a contention stress (LOGCORT) and body weight at 2 weeks of age (BW2). A total of 310 animals were included in a genome scan using selective genotyping with 248 AFLP markers. A total of 21 suggestive or genome-wide significant QTL were observed. Two highly significant QTL were identified on linkage group 1 (GL1), one for LOGTI and one for NI. In the vicinity of the QTL for LOGTI, a nearly significant QTL for SHOUT and a suggestive QTL for LAT were also identified. On GL3, genome-wide significant QTL were observed for JUMP and DEJ as well as suggestive QTL for LOGTI, MOVE, SHOUT and LAT. A significant QTL for BW2 was observed on GL2 and a nearly significant one on GL1. These results may be useful in the understanding of fearfulness in quail and related species provided that fearfulness has the same genetic basis.     

Characterization of a Phytoplasma Associated with Pear Decline in Iran

Symptoms of pear decline (PD) were observed in several pear growing regions of Iran. Pear trees with typical symptoms of PD from Estahban (Fars Province) were examined for phytoplasma infection using polymerase chain reaction (PCR) assay. Graft inoculation of healthy pear trees with scions from diseased trees resulted in production of PD symptoms and transmission of phytoplasma as verified by PCR. Target DNA was amplified from symptomatic pear trees with fO1/rO1, an apple proliferation (AP) group-specific primer pair. Physical and putative restriction fragment length polymorphism (RFLP) analyses of fO1/rO1 primed PCR products showed profiles corresponding to AP group, 16SrX-C subgroup ( Candidatus Phytoplasma pyri). Percent similarity values and phylogenetic analysis of fO1/rO1 primed sequences confirmed that, as a member of AP subclade, Estahban PD phytoplasma has a closer relationship to PD and peach yellow leaf roll phytoplasmas than to AP ( Ca . Phytoplasma mali) and European stone fruit yellows ( Ca . Phytoplasma prunorum) phytoplasmas. This is the first report of PD phytoplasma in the eastern Mediterranean.     

梨品种资源果实轮纹病抗性的评价

采用田间人工接种的方法,2009-2010年连续两年对保存于国家梨种质资源更新圃的182个品种进行果实轮纹病抗性评价。结果表明：砂梨、白梨、秋子梨、种间杂交梨、西洋梨和新疆梨的两年平均发病率分别为6.15%、7.20%、7.43%、12.66%、17.00%、18.93%,梨品种间对轮纹病抗性存在差异,不同条件下多数品种轮纹病发病率有差异。依据两年平均发病率进行抗性评价分级,72个品种为高抗、63个品种为抗、25个品种为中抗、14个品种为低抗、8个品种为不抗。     

Identification of QTLs Associated with Oil Content in a High-Oil Brassica napus Cultivar and Construction of a High-Density Consensus Map for QTLs Comparison in B. napus

Increasing seed oil content is one of the most important goals in breeding of rapeseed (B. napus L.). To dissect the genetic basis of oil content in B. napus, a large and new double haploid (DH) population containing 348 lines was obtained from a cross between ‘KenC-8’ and ‘N53-2’, two varieties with >10% difference in seed oil content, and this population was named the KN DH population. A genetic linkage map consisting of 403 markers was constructed, which covered a total length of 1783.9 cM with an average marker interval of 4.4 cM. The KN DH population was phenotyped in eight natural environments and subjected to quantitative trait loci (QTL) analysis for oil content. A total of 63 identified QTLs explaining 2.64–17.88% of the phenotypic variation were identified, and these QTLs were further integrated into 24 consensus QTLs located on 11 chromosomes using meta-analysis. A high-density consensus map with 1335 marker loci was constructed by combining the KN DH map with seven other published maps based on the common markers. Of the 24 consensus QTLs in the KN DH population, 14 were new QTLs including five new QTLs in A genome and nine in C genome. The analysis revealed that a larger population with significant differences in oil content gave a higher power detecting new QTLs for oil content, and the construction of the consensus map provided a new clue for comparing the QTLs detected in different populations. These findings enriched our knowledge of QTLs for oil content and should be a potential in marker-assisted breeding of B. napus.     

梨种质对梨火疫病的抗性研究

针对未来梨火疫病侵入我国的风险,采用幼果人工接种鉴定技术,对保存的54份梨种质资源进行梨火疫病抗性评价。通过接种后的症状鉴别、再分离细菌的分子检测,证明有16份种质对梨火疫病表现感病,其中9份为中国种质,7份为引进种质。中国的37份（80．4％）梨种质表现抗梨火疫病,而引进种质中表现抗病的仅有1份（12．5％）。结果初步表明,在中国梨种质中可能存在较丰富的抗梨火疫病资源;东方梨的抗性较西方梨更强;而在梨的地方品种中蕴藏着良好的抗病性。     

Identification of QTLs associated with citrus resistance toPhytophthora gummosis

Citrus gummosis, caused by Phytophthora spp., is an important citrus disease in Brazil. Almost all citrus rootstock varieties are susceptible to it to some degree, whereas resistance is present in Poncirus trifoliata, a closely related species. The objective of this study was to detect QTLs linked to citrus Phytophthora gummosis resistance. Eighty individuals of the F1 progeny, obtained by controlled crosses between Sunki mandarin Citrus sunki (susceptible) and Poncirus trifoliata cv. Rubidoux (resistant), were evaluated. Resistance to Phytophthora parasitica was evaluated by inoculating stems of young plants with a disc of fungal mycelia and measuring lesion lengths a month later. Two QTLs linked to gummosis resistance were detected in linkage groups 1 and 5 of the P. trifoliata map, and one QTL in linkage group 2 of the C. sunki map. The phenotypic variation explained by individual QTLs was 14% for C. sunki and ranged from 16 to 24% for P. trifoliata. The low character heritability (h2 = 18.7%) and the detection of more than one QTL associated with citrus Phytophthora gummosis resistance showed that inheritance of the resistance is quantitative.     

Identification of QTLs associated with male strobilus abundance in Cryptomeria japonica

It has become increasingly important to understand the factors controlling the abundance of male strobili in Cryptomeria japonica since allergic reactions to the species?? pollen have become prevalent in Japan. There is considerable inter-annual variation in the abundance of male strobili, so it is important to investigate and validate quantitative trait loci (QTL) for strobili abundance across different years. Data on third generation families derived from an individual that produces abundant male strobili were studied using Kruskal?CWallis tests and Bayesian regression analysis, resulting in the detection of a single major QTL that was observed over multiple years. This QTL explained between 10.4 and 26.5?% of the phenotypic variation in male strobilus abundance. The consistency and significance of this QTL??s effects suggest that it has a major role in male strobilus production in C. japonica, although we also detected other QTLs, indicating that strobilus production is subject to complex and multifactorial regulation. These QTL may also affect precocity because we were unable to separate their effects from the genetic variability in male strobilus production. The major QTL that we detected occurs in a region homologous to a QTL detected in a previous study examining trees with a different genetic heritage following artificial induction of male strobilus production. This result further supports the importance of the QTL detected in the present study. Identifying the important genes in this QTL will therefore significantly increase our understanding of the genetic mechanisms of pollen production in C. japonica.     

Identification of QTLs associated with physiological nitrogen use efficiency in rice

Demand for low-input sustainable crop cultivation is increasing to meet the need for environment-friendly agriculture. Consequently, developing genotypes with high nutrient use efficiency is one of the major objectives of crop breeding programs. This study was conducted to identify QTLs for traits associated with physiological nitrogen use efficiency (PNUE). A recombinant inbred population (DT-RILs) between Dasanbyeo (a tongil type rice, derived from an indica x japonica cross and similar to indica in its genetic make-up) and TR22183 (a Chinese japonica variety) consisting of 166 F8 lines was developed and used for mapping. A frame map of 1,409 cM containing 113 SSR and 103 STS markers with an average interval of 6.5 cM between adjacent marker loci was constructed using the DT-RILs. The RILs were cultivated in ordinary-N (N-P2O5-K2O = 100-80-80 kg/ha) and low-N (N-P2O5-K2O= 50-80-80 kg/ha) (100 kg/ha) conditions. PNUE was positively correlated with the harvest index and grain yield in both conditions. Twenty single QTLs (S-QTLs) and 58 pairs of epistatic loci (E-QTLs) were identified for the nitrogen concentration of grain, nitrogen concentration of straw, nitrogen content of shoot, harvest index, grain yield, straw yield and PNUE in both conditions. The phenotypic variance explained by these S-QTLs and E-QTLs ranged from 11.1 to 44.3% and from 16.0% to 63.6% , respectively. The total phenotypic variance explained by all the QTLs for each trait ranged from 35.8% to 71.3%, showing that the expression of PNUE and related characters depends significantly upon genetic factors. Both S-QTLs and E-QTLs may be useful for marker-assisted selection (MAS) to develop higher PNUE genotypes.     

Identification of QTLs conferring resistance to downy mildews of maize in Asia

Downy mildew is one of the most destructive diseases of maize in subtropical and tropical regions in Asia. As a prerequisite for improving downy mildew resistance in maize, we analyzed quantitative trait loci (QTLs) involved in resistance to the important downy mildew pathogens--Peronosclerospora sorghi (sorghum downy mildew) and P. heteropogoni (Rajasthan downy mildew) in India, P. maydis (Java downy mildew) in Indonesia, P. zeae in Thailand and P. philippinensis in the Philippines--using a recombinant inbred line population derived from a cross between Ki3 (downy mildew resistant) and CML139 (susceptible). Resistance was evaluated as percentage disease incidence in replicated field trials at five downy mildew 'hotspots' in the four countries. Heritability estimates of individual environments ranged from 0.58 to 0.75 with an across environment heritability of 0.50. Composite interval mapping was applied for QTL detection using a previously constructed restriction fragment length polymorphism linkage map. The investigation resulted in the identification of six genomic regions on chromosomes 1, 2, 6, 7 and 10 involved in the resistance to the downy mildews under study, explaining, in total, 26-57% of the phenotypic variance for disease response. Most QTL alleles conferring resistance to the downy mildews were from Ki3. All QTLs showed significant QTL x environment interactions, suggesting that the expression of the QTL may be environment-dependent. A strong QTL on chromosome 6 was stable across environments, significantly affecting disease resistance at the five locations in four Asian countries. Simple-sequence repeat markers tightly linked to this QTL were identified for potential use in marker-assisted selection.