Recent advances in soybean transformation and their application to molecular breeding and genomic analysis

Herbicide-resistant transgenic soybean plants hold a leading market share in the USA and other countries, but soybean has been regarded as recalcitrant to transformation for many years. The cumulative and, at times, exponential advances in genetic manipulation have made possible further choices for soybean transformation. The most widely and routinely used transformation systems are cotyledonary node–Agrobacterium-mediated transformation and somatic embryo–particle-bombardment-mediated transformation. These ready systems enable us to improve seed qualities and agronomic characteristics by transgenic approaches. In addition, with the accumulation of soybean genomic resources, convenient or promising approaches will be requisite for the determination and use of gene function in soybean. In this article, we describe recent advances in and problems of soybean transformation, and survey the current transgenic approaches for applied and basic research in Japan.     

QTL mapping and associated marker selection for the efficacy of green plant regeneration in anther culture of rice

Anther culturability of rice is a quantitative trait controlled by nuclear‐encoded genes. The identification of quantitative trait loci (QTL) and associated marker selection for anther culturability is important for increasing the efficiency of green plant regeneration from microspores. QTL associated with the capacity for green plant regeneration in anther culture of rice were mapped on chromosomes 3 and 10 using 164 recombinant inbred (RI) lines from a cross between ‘Milyang 23’ and ‘Gihobyeo’. The quantitative trait locus located on chromosome 10 was detected repeatedly when three anther culture methods were applied and was tightly linked to the markers, RG323, RG241 and RZ400. Associations between these markers and the efficacy of green plant regeneration in 43 rice cultivars and two F₂ populations, ‘MG RI036’/‘Milyang 23’, and ‘MG RI036’;/‘IR 36’ were analysed. One of these markers, RZ400, was able to identify effectively genotypes with good (> 10.0%) and poor (< 3.0%) regenerability, based on the marker genotypes in the cultivars and two F₂ populations. This marker enables the screening of rice germplasm for anther culturability and introgression into elite lines in breeding programmes.     

QTL mapping and marker-assisted selection for Fusarium head blight resistance in wheat: a review

During the past decade, numerous studies have been published on molecular mapping of Fusarium head blight (FHB) resistance in wheat. We summarize the relevant findings from 52 quantitative trait loci (QTL) mapping studies, nine research articles on marker-assisted selection and seven on marker-assisted germplasm evaluation. QTL for FHB resistance were found on all wheat chromosomes except chromosome 7D. Some QTL were found in several independent mapping studies indicating that such QTL are stable and therefore useful in breeding programmes. We summarize and update current knowledge on the genetics of FHB resistance in wheat resulting from QTL mapping investigations and review and suggest FHB breeding strategies based on the available information and DNA markers.     

外来入侵植物与地下生态系统相互影响的研究进展

摘要：生物入侵已成为世界性的生态、经济问题,是人类当前面临的巨大挑战。作为入侵生物中一个重要组分的外来入侵植物,其入侵不仅改变了入侵地地上植物群落的多样性,而且对入侵地的地下生态系统也产生了深刻影响。国内外的生态学家对于外来植物的入侵虽已提出多个机制假说,但真正机理还未明确。近年来兴起的外来入侵植物与入侵地土壤生态过程相互影响的研究为外来植物入侵机理的揭示提供了新思路。本文从两个方面综述了近年来对外来入侵植物与入侵地地下生态系统相互作用的研究结果：（1）入侵植物与入侵地土壤微生物的相互影响：外来入侵植物可通过破坏土著植物与土壤微生物间的共生关系、分泌化感物质影响入侵地微生物群落结构和功能、逃避原产地土传天敌、改变入侵地的微生物群落结构进而间接改变土壤养分循环等途径实现入侵;（2）入侵植物与入侵地土壤养分的相互影响：主要是入侵植物与入侵地土壤氮、磷、钾等几种大量元素及其他元素之间的相互作用。在综述国内外研究的基础上,探讨了外来入侵植物入侵机理研究中存在的问题及未来的研究方向,以期为外来植物入侵的预防、控制与生境恢复提供依据。     

Cold stress tolerance of soybeans during flowering: QTL mapping and efficient selection strategies under controlled conditions

Breeding soybeans for higher latitudes requires cultivars with an increased chilling stress tolerance, especially when flowering occurs. Phenotyping in climate chambers to select for this trait is labour‐intensive and requires an optimal allocation of resources due to limited space. We screened a diversity panel of 35 early maturity cultivars and a biparental population of 103 RILs for their cold stress tolerance at flowering stage. Pod number under control and stress conditions is highly heritable and showed only a weak correlation between the two treatments. Based on different testing scenarios, we could show that testing more genotypes with less replicates yields much higher responses to selection and hence should be pursued in such climate‐controlled experiments. We identified quantitative trait loci (QTL) for pod number under both conditions (chromosomes 7 and 13) and a cold tolerance‐specific QTL (chromosome 11). Furthermore, we performed genomic predictions using different test set scenarios and prediction models, showing that genomic prediction is a promising tool to select for cold stress tolerance, particularly if known QTL can be used as fixed effects in the model.     

An introduction to markers, quantitative trait loci (QTL) mapping and marker-assisted selection for crop improvement: The basic concepts

Recognizing the enormous potential of DNA markers in plant breeding, many agricultural research centers and plant breeding institutes have adopted the capacity for marker development and marker-assisted selection (MAS). However, due to rapid developments in marker technology, statistical methodology for identifying quantitative trait loci (QTLs) and the jargon used by molecular biologists, the utility of DNA markers in plant breeding may not be clearly understood by non-molecular biologists. This review provides an introduction to DNA markers and the concept of polymorphism, linkage analysis and map construction, the principles of QTL analysis and how markers may be applied in breeding programs using MAS. This review has been specifically written for readers who have only a basic knowledge of molecular biology and/or plant genetics. Its format is therefore ideal for conventional plant breeders, physiologists, pathologists, other plant scientists and students.     

Genetic mapping,marker assisted selection and allelic relationships for the Pu6 gene conferring rust resistance in sunflower

Rust resistance in the sunflower line P386 is controlled by Pu₆, a gene which was reported to segregate independently from other rust resistant genes, such as R₄. The objectives of this work were to map Pu₆, to provide and validate molecular tools for its identification, and to determine the linkage relationship of Pu₆ and R₄. Genetic mapping of Pu₆ with six markers covered 24.8 cM of genetic distance on the lower end of linkage Group 13 of the sunflower consensus map. The marker most closely linked to Pu₆ was ORS316 at 2.5 cM in the distal position. ORS316 presented five alleles when was assayed with a representative set of resistant and susceptible lines. Allelism test between Pu₆ and R₄ indicated that both genes are linked at a genetic distance of 6.25 cM. This is the first confirmation based on an allelism test that at least two members of the R_adv/R₄/R₁₁/ R_13a/R_13b/Pu₆ cluster of genes are at different loci. A fine elucidation of the architecture of this complex locus will allow designing and constructing completely new genomic regions combining genes from different resistant sources and the elimination of the linkage drag around each resistant gene.     

The role of molecular markers and marker assisted selection in breeding for organic agriculture

Plant geneticists consider molecular marker assisted selection a useful additional tool in plant breeding programs to make selection more efficient. Standards for organic agriculture do not exclude the use of molecular markers as such, however for the organic sector the appropriateness of molecular markers is not self-evident and is often debated. Organic and low-input farming conditions require breeding for robust and flexible varieties, which may be hampered by too much focus on the molecular level. Pros and contras for application of molecular markers in breeding for organic agriculture was the topic of a recent European plant breeding workshop. The participants evaluated strengths, weaknesses, opportunities, and threats of the use of molecular markers and we formalized their inputs into breeder’s perspectives and perspectives seen from the organic sector’s standpoint. Clear strengths were identified, e.g. better knowledge about gene pool of breeding material, more efficient introgression of new resistance genes from wild relatives and testing pyramided genes. There were also common concerns among breeders aiming at breeding for organic and/or conventional agriculture, such as the increasing competition and cost investments to get access to marker technology, and the need for bridging the gap between phenotyping and genotyping especially with complex and quantitative inherited traits such as nutrient-efficiency. A major conclusion of the authors is that more interaction and mutual understanding between organic and molecular oriented breeders is necessary and can benefit both research communities.     

Kazusa Marker DataBase: a database for genomics,genetics, and molecular breeding in plants

In order to provide useful genomic information for agronomical plants, we have established a database, the Kazusa Marker DataBase (http://marker.kazusa.or.jp). This database includes information on DNA markers, e.g., SSR and SNP markers, genetic linkage maps, and physical maps, that were developed at the Kazusa DNA Research Institute. Keyword searches for the markers, sequence data used for marker development, and experimental conditions are also available through this database. Currently, 10 plant species have been targeted: tomato (Solanum lycopersicum), pepper (Capsicum annuum), strawberry (Fragaria × ananassa), radish (Raphanus sativus), Lotus japonicus, soybean (Glycine max), peanut (Arachis hypogaea), red clover (Trifolium pratense), white clover (Trifolium repens), and eucalyptus (Eucalyptus camaldulensis). In addition, the number of plant species registered in this database will be increased as our research progresses. The Kazusa Marker DataBase will be a useful tool for both basic and applied sciences, such as genomics, genetics, and molecular breeding in crops.     

Molecular mapping of QTL for southwestern corn borer resistance, plant height and flowering in tropical maize

Development of multiple insect resistance in tropical maize represents a major effort of the maize breeding programme at CIMMYT. Resistance to the southwestern corn borer (SWCB) is polygenically controlled with primarily additive gene action. Our main objective was to identify quantitative trait loci (QTL) involved in resistance to SWCB. Other objectives were to detect QTL in the same population for plant height, female flowering, and the anthesis-silking interval (ASI). A population of 472 F₂ individuals derived from a cross between the susceptible line Ki3 and the resistant inbred CML139, was restriction fragment length polymorphism (RFLP) genotyped using 110 maize probes. F₃ families were rated for leaf-feeding damage after artificial infestation at one location in three consecutive years. Height and flowering were measured in protected trials in two locations. QTL analyses were conducted using joint composite interval mapping. Seven QTL on chromosomes 3, 5, 6, 8, and 9 explained 30% of the phenotypic variance (σ²_p) for SWCB damage. Most QTL alleles conferring resistance were contributed from CML139. QTL showed dominance, partial dominance and additive gene action. Eleven QTL dispersed across the genome were determined to affect plant height and explained 43% of σ²_p. Four of these were in close proximity to loci with qualitative effects on plant height. Thirteen QTL (50% of sigma;²_p) were identified for days to female flowering and nine (30% of σ²_p) for ASI. Our results, along with those from other mapping studies at CIMMYT, are allowing us to formulate marker-assisted selection schemes to complement the breeding efforts for such complex traits as borer resistance.     

QTL mapping for chromium-induced growth and zinc, and chromium distribution in seedlings of a rice DH population

Chromium contamination in soil has become a severe threat to crop production and food safety. The experiment was conducted using a rice DH population to detect the QTLs associated with Cr tolerance. Seventeen putative QTLs associated with growth traits included three additive loci and fourteen epistatic loci. These loci were distributed on 11 rice chromosomes, and their contribution to the phenotypic variation ranged from 2.44 to 10.08%. Two QTLs located at the similar genetic region on chromosome ten were associated with shoot Cr concentration and translocation from roots to shoots, respectively; and they accounted for 11.65 and 11.22% of the phenotypic variation. In addition, six QTLs related to Zn concentration and translocation was found on chromosomes 1, 2, 4, 5, 7 and 12. Meanwhile epistatic effect existed in the two additive QTLs of qRZC1 and qRZC7. Most of QTLs controlling Zn concentration had small genotypic variance and qSRZ4 related to Zn translocation showed growth condition-dependent expression.     

Molecular mapping and gene action of Scm1 and Scm2, two major QTL contributing to SCMV resistance in maize

Sugarcane mosaic virus (SCMV) is one of the most important virus diseases of maize in Europe. In this study, the gene action at two major quantitative trait loci (QTL) affecting resistance to SCMV in maize was mapped and characterized. A total of 121 F₃ lines from cross F7 (susceptible) × FAP1360A (resistant) were evaluated for SCMV resistance in replicated field trials across two environments under artificial inoculation at seven scoring dates. The genotypic variance was always highly significant and heritability increased up to 0.92 for later scoring dates. The method of composite interval mapping was employed for QTL mapping using four simple sequence repeat (SSR) markers flanking two regions identified in a previous study with cross D145 × D32. The presence of two QTL for SCMV resistance, one on chromosome 6 (Scml region) and one on chromosome 2 (Scm2 region), was confirmed. These two QTL together explained between 15% (first score) and 62% (final score) of the phenotypic variance at various stages of plant development. Gene action was additive for the Scm1 region but completely dominant for the Scm2 region. Comparison of results of this study with those obtained for cross D145 × D32 suggested that the resistance alleles in the two populations are identical for the Scm1 region but different for the Scm2 region.     

Genetics, fertility behaviour and molecular marker analysis of a new TGMS line, TS6, in rice

The thermosensitive genic male sterility (TGMS) system has great potential for revolutionizing hybrid rice production through simple, less expensive and more efficient seed production technology. For the successful utilization of this novel male sterility system, knowledge of the breeding and fertility behaviour of a TGMS line is essential. In this study, the fertility transformation behaviour, the critical fertility and sterility temperatures and the mode of inheritance of male sterility were studied for a new TGMS line, TS6, identified at Tamil Nadu Agricultural University, Coimbatore, India. The pollen and spikelet fertilities recorded on plants raised at fortnightly intervals revealed that this line was completely sterile for 78 consecutive days (35/22 to 32/23°C, maximum/minimum temperatures) and reverted to fertile when the temperature was 30/18°C. It remained fertile continuously for 69 days and the maximum pollen and spikelet fertilities recorded were 75 and 70%, respectively. The fertility was highly influenced by daily maximum temperature followed by average and minimum temperatures. It was not influenced by relative humidity, sunshine hours or photoperiod. The critical temperature inducing sterility and fertility was 26.7 and 25.5°C, respectively. The male sterility in TS6 was inherited as a monogenic recessive in the F₂ and BC₁ populations of TS6 × MRST9 as well as TS6 × IR68281B. Using bulked segregant analysis on an F₂ population of TS6 × MRST9, an RAPD marker, OPC05₂₉₆₂, was identified to be associated with TGMS in TS6.     

利用DH和IF_2群体检测油菜籽粒油酸、亚油酸、亚麻酸含量QTL

以德国冬性甘蓝型油菜Express和中国半冬性甘蓝型油菜SWU07为亲本构建包含261个株系的DH群体和包含234个株系的IF2群体,检测不同年份条件下油菜籽粒油酸、亚油酸、亚麻酸含量相关的QTL。在DH群体4年环境下共检测出71个QTL,在IF2群体2年环境下共检测出4个QTL。去掉在不同年份和群体中置信区间相互重叠的QTL之后,共得到3个品质性状的51个QTL,其中有15个在2年以上环境中被检测到。这些QTL分别分布在13个连锁群上,其中与油酸含量相关的18个,分布于A01、A02、A04、A05、A07、A09、C01连锁群,揭示3.44%～13.97%的表型变异;与亚油酸相关的12个,分布于A02、A06、A09、C01、C02连锁群,揭示3.84%～19.51%的表型变异;与亚麻酸相关的21个,分布于A01、A02、A03、A04、A05、A08、A09、C01、C02、C03、C06连锁群,揭示2.86%～11.91%的表型变异。这些结果将为油菜脂肪酸品质改良提供更多遗传信息。     

Comparative molecular marker-based genetic mapping of flavanone 3-hydroxylase genes in wheat,rye and barley

The F3 h gene encoding flavanone 3-hydroxylase, one of the key enzymes of the flavonoid biosynthesis pathway, is involved in plant defense response, however, it has not yet been genetically mapped in such important crop species as wheat, barley and rye. In the current study, the F3 h genes were for the first time genetically mapped in these species, using microsatellite and RFLP markers. The three wheat F3 h homoeologous copies F3 h-A1, F3 h-B1 and F3 h-D1, and rye F3 h-R1 were mapped close to the microsatellite loci Xgwm0877 and Xgwm1067 on chromosomes 2AL, 2BL, 2DL, and 2RL, respectively. Wheat F3 h-G1 and barley F3 h-H1 were also mapped at the homoeologous F3 h-1 position on chromosomes 2GL and 2HL, respectively. The non-homoeologous F3 h gene (F3 h-B2) was mapped on wheat chromosome 2BL about 40 cM distal to the F3 h-1 map position. The results obtained in the current study are important for further studies aimed on manipulation with F3 h expression (and, hence, plant defense) in wheat, barley and rye.     

Identification for H. villosa chromatin in wheat lines using genomic in situ hybridization, C-banding and gliadin electrophoresis techniques

Detection of H. villosa chromosomes in telosomic addition and translocation lines of common wheat was undertaken using genomic in situ hybridization (GISH), C-banding techniques and polyacrylamide gels electrophoresis. The result of GISH on mitotic metaphase cells of the addition line `95039' indicated that the added telochromosomes originated from H. villosa, and it was probably 6VS or 7Vs of H. villosa according to the C-banding pattern. Furthermore, the analysis of gliadin profiles demonstrated that the telochromosome was 6VS. A pair of 1RS/1BL translocation chromosome was also found in `95039'. In addition, mitotic GISH analysis showed that the 6VS/6AL translocation chromosome remained unchanged after being transferred into new wheat background. This revised version was published online in August 2006 with corrections to the Cover Date.     

QTL mapping and marker analysis of main stem height and the first lateral branch length in peanut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis> L.)

Main stem height (MSH) and the first lateral branch length (LBL) of peanut (Arachis hypogaea L.) are important traits affecting plant shape and yield. Identification of quantitative trait loci (QTLs) related to these two traits, the prediction and cloning of candidate genes, and identification of plant height-related molecular markers are the basis for analysis of the molecular genetic mechanism of plant shape in peanut. In this study, a population of 151 recombinant inbred lines from a single seed, derived from a cross between variety 79266 (P1) and its variant progeny D893 (P2), was used to construct a peanut genetic map. The map consisted of 231 simple sequence repeat markers in 23 linkage groups, had a total length of 905.18 cM with average and minimum marker intervals of 3.92 and 0.1 cM, respectively. There were 11 and 16 QTLs detected in six environments for MSH and LBH with 6.26–22.53 and 5.89–21.63% phenotypic variation explained (PVE), respectively. Seven QTLs were detected in two or more environments: 3 QTLs for MSH (including Qmsh-14-3) with 7.66–22.53% PVE, and 4 QTLs for LBL (including Qllb-11-1) with 6.12–21.63% PVE. Qmsh-14-3 was steadily detected in five environments, localized between two markers, ARS376 and SEQ4G02, exhibited a genetic distance of 0.59–2.59 and 4.11–6.11 cM from the two markers, respectively. Qllb-11-1 was steadily detected in five environments, localized between two markers, ARS203 and AHS1413, exhibited a genetic distance of 1.06–3.06 and 0.23–2.23 cM from the two markers, respectively. There were 220 germplasm accessions used to detect the relationship between genotype and phenotypic values of traits at the marker loci ARS376 and SEQ4G02, average values of MSH and LBL were significantly higher for germplasm with the P1 compared to the P2 genotype. Determination of the marker loci ARS203 and AHS1413 indicated that average values of LBL were greater in germplasm with the P1 than the P2 genotype. The results provide references for fine mapping of QTLs for MSH and LBL, as well as breeding optimum plant-types in peanut.     

Inheritance of resistance to angular leaf spot in common bean and validation of the utility of resistance linked markers for marker assisted selection out side the mapping population

Inheritance of resistance to angular leaf spot (ALS) disease caused by Phaeoisariopsis griseola (Sacc.) Ferr was investigated in two common bean cultivars, Mexico 54 and BAT 332. Both Andean and Mesoamerican backgrounds were used to determine the stability of the resistance gene in each of the two cultivars. Resistance to P. griseola was phenotypically evaluated by artificial inoculation with one of the most widely distributed pathotypes, 63–39. Evaluation of the parental genotypes, F₁, F₂ and backcross populations revealed that the resistance to angular leaf spot in the cultivars Mexico 54 and BAT 332 to pathotype 63–39 is controlled by a single dominant gene, when both the Andean and Mesoamerican backgrounds were used. Allelism test showed that ALS resistance in Mexico 54 and BAT 332 to pathotype 63–39 was conditioned by the same resistance locus. Resistant and susceptible segregating populations generated using Mexico 54 resistant parent were selected for DNA extraction and amplification to check for the presence /absence of the SCAR OPN02 and RAPD OPE04 markers linked to the Phg-2 resistance gene. The results indicated that the SCAR OPN02 was not polymorphic in the study populations and therefore of limited application in selecting resistant genotypes in such populations. On the other hand, the RAPD OPE04 marker was observed in all resistant individuals and was absent in those scored susceptible based on virulence data. Use of the RAPD OPE04 marker in marker-assisted selection is underway.