Mapping and use of QTLs controlling pod dehiscence in soybean

While the cultivated soybean, Glycine max (L.) Merr., is more recalcitrant to pod dehiscence (shattering-resistant) than wild soybean, Glycine soja Sieb. & Zucc., there is also significant genetic variation in shattering resistance among cultivated soybean cultivars. To reveal the genetic basis and develop DNA markers for pod dehiscence, several research groups have conducted quantitative trait locus (QTL) analysis using segregated populations derived from crosses between G. max accessions or between a G. max and G. soja accession. In the populations of G. max, a major QTL was repeatedly identified near SSR marker Sat_366 on linkage group J (chromosome 16). Minor QTLs were also detected in several studies, although less commonality was found for the magnitudes of effect and location. In G. max × G. soja populations, only QTLs with a relatively small effect were detected. The major QTL found in G. max was further fine-mapped, leading to the development of specific markers for the shattering resistance allele at this locus. The markers were used in a breeding program, resulting in the production of near-isogenic lines with shattering resistance and genetic backgrounds of Japanese elite cultivars. The markers and lines developed will hopefully contribute to the rapid production of a variety of shattering-resistant soybean cultivars.     

QTL analysis of flooding tolerance in soybean at an early vegetative growth stage

Soybean cultivars are sensitive to flooding stress and their seed yields are substantially reduced in response to the stress. This study was conducted to investigate the genetic basis of flooding tolerance at an early vegetative growth stage. Sixty recombinant inbred lines derived from a cross between a relatively tolerant cv. ‘Misuzudaizu’ and a sensitive cv. ‘Moshidou Gong 503’ were grown in pots in a vinyl plastic greenhouse in 2002 and 2003. At the two‐leaf stage, half of the pots were waterlogged by water placed in plastic containers and adjusted to 5 cm above the soil surface. After 3 weeks of treatment, the pots were returned to the greenhouse and grown until maturity. Flooding tolerance was evaluated by dividing the seed weight of the treated plants by that of the control plants. Quantitative trait loci (QTL) analysis using 360 genetic markers revealed three QTLs for flooding tolerance, ft1 to ft3 in 2002. The ft1 (molecular linkage group C2) was reproducible and an additional four QTLs, ft4 to ft7, were found in 2003. The ft1 had a high LOD score in both years (15.41 and 7.57) and accounted for 49.2% and 30.5% of the total variance, respectively. A large QTL for days to flowering was consistently observed across treatments and years at a similar position to ft1. Comparing the relative location with markers, the maturity gene probably corresponds to E1. Late maturity may have conferred a longer growth period for recovery from flooding stress.     

Inheritance of the four-seeded-pod trait in a soybean mutant and marker-assisted selection for this trait

A high seeds‐per‐pod value is a crucial component of soybean seed yield, but reliable information is lacking on the inheritance and early selection for the four‐seeded pod (4SP) trait. The inheritance of 4SP was followed in crosses originating from an EMS‐derived mutant line (E182, 15% 4SP) with its parental cultivar, ‘Ludou No. 4’ (no 4SP), and the efficiencies of both a molecular and morphological marker selection were determined. The plants of two F₂ populations (with E182 as one of the parents) showed a segregation ratio of 3 : 1 (low to high SP values), indicating a single recessive locus; this mode of inheritance was confirmed by subsequent analysis of the F_{2 : 3} families. Among four microsatellite markers linked with the 4SP trait, Sat_107 was the closest to the 4SP locus (3.2 ± 1.11 cM). Both the molecular marker Sat_107 and the morphological marker, narrow leaflet (NL) associated with the 4SP locus were effective in selecting the 4SP trait, although with different efficiencies. The molecular marker was efficient in selecting soybean progenies from the cross in which one parent was the E182 mutant line because it was tightly linked to the mutated 4SP locus. In the other cross, with 4SP not derived from E182, the efficiency was reduced in comparison with the morphological marker.     

Identification of SSR markers linked to the Phytophthora resistance gene Rps1-d in soybean

To identify markers for the Phytophthora resistance gene, Rps1‐d, 123 F_{2 : 3} families were produced from a cross between Glycine max (L.) Merr. ‘Tanbakuro’ (a Japanese traditional black soybean) and PI103091 (Rps1‐d) as an experimental population. The results of virulence tests produced 33 homozygous resistant, 61 segregating and 29 homozygous susceptible F_{2 : 3} families. The chi‐squared test gave a goodness‐of‐fit for the expected ratio of 1 : 2 : 1 for resistant, segregating and susceptible traits, suggesting that the inheritance of Rps1‐d is controlled by a monogenic dominant gene. Simple sequence repeat (SSR) analyses of this trait were carried out using the cultivars ‘Tanbakuro’ and PI103091. Sixteen SSR primers, which produced 19 polymorphic fragments between the two parents, were identified from 41 SSR primers in MLG N. Eight SSR markers were related to Rps1‐d, based on 32 of the 123 F_{2 : 3} families, consisting of 16 homozygous resistant and 16 homozygous susceptible lines. The remaining 91 families were analysed for these eight markers, and a linkage map was constructed using all 123 F_{2 : 3} families. The length of this linkage group is 44.0 cM. The closest markers, Sat_186 and Satt152, are mapped at 5.7 cM and 11.5 cM, respectively, on either side of the Rps1‐d gene. Three‐way contingency table analysis indicates that dual‐marker‐assisted selection using these two flanking markers would be efficient.     

The presence or absence of the soybean Kunitz trypsin inhibitor as a quantitative trait locus for seed protein content

Numerous quantitative trait loci (QTL) for various characters have recently been reported in different crop plants. However, information is limited about the molecular mechanisms behind QTL, because most of them have only been detected at a statistical level. Therefore, progeny from a cross between two soybean genotypes segregating for the presence vs. absence of the Kunitz trypsin inhibitor, a 21.5 kDa protein, have been analysed for possible effects of that protein on agronomic and seed quality characters. Protein content was reduced by, on average, 4.5 g/kg in segregants lacking the Kunitz protein, whereas oil content and other characters remained unaffected. This finding can be interpreted as a ‘model QTL’ for variation in seed protein content, because the molecular and genetic backgrounds of the soybean Kunitz trypsin inhibitor are well understood.     

Identification of AFLP markers linked to the epistatic suppressor gene of a recessive genic male sterility in rapeseed and conversion to SCAR markers

A recessive epistatic genic male sterility (REGMS) two‐type line, 9012AB, has been used for rapeseed hybrid seed production in China. The male sterility of 9012AB is controlled by two recessive duplicate sterile genes (ms1 and ms2) interacting with one recessive epistatic suppressor gene (esp). Homozygosity at the esp locus (espesp) suppresses the expression of the recessive male sterility trait in homozygous ms1ms1ms2 ms2 plants. In this study, we used a combination of bulked segregant analyses and amplified fragment length polymorphism (AFLP) to identify markers linked to the suppressor gene in a BC₁ population. From the survey of 1024 AFLP primer combinations, eight markers tightly linked to the target gene were identified. The two closest markers flanking both sides of Esp, P9M5₃₇₀ and S16M14₇₈₀, had a genetic distance of 1.4 cM and 2.1 cM, respectively. The AFLP fragment from P4M8₁₉₀, which co‐segregated with the target gene was converted into a sequence characterized amplified region marker. The availability of linked molecular markers will facilitate the utilization of REGMS in hybrid breeding in Brassica napus.     

Development of STS markers and QTL validation for common bacterial blight resistance in common bean

Common bacterial blight (CBB) of common bean ( Phaseolus vulgaris L.), is one of the major diseases that decrease yield and quality. A major quantitative trait locus (QTL) for CBB resistance from line XAN 159 was transferred into two bean lines, HR45 and HR67. Previous studies identified that two markers are linked to this QTL but the chromosome location was not consistent. To identify more tightly linked markers and to verify the chromosome location, 65 additional markers were mapped using 81 recombinant inbred lines (RILs) derived from a cross HR67 × OAC95-4. The QTL was mapped to a 13 cM region on chromosome 1 and defined by eight molecular markers that explained 25–52% of the phenotypic variation. Six tightly linked amplified fragment length polymorphism markers (0.6–9.7 cM from the QTL peak) were converted into seven sequence tagged site markers, three of which were mapped to this QTL. Five tightly linked markers were used to screen 907 F₂ plants derived from a cross HR45 × 'OAC Rex' and four of them were linked to each other within 4.2 cM. These markers may be useful in marker-assisted selection and map-based cloning of this major QTL.     

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.     

Mapping and validation of PCR-based markers associated with a major QTL for seed dormancy in wheat

Seed dormancy is one of the important factors controlling pre-harvest sprouting (PHS) resistance in wheat. We identified a major quantitative trait locus (QTL) for seed dormancy on the long arm of wheat chromosome 4A (4AL) via simple sequence repeat (SSR)-based genetic mapping using doubled haploid lines from a cross between Japanese PHS resistant variety ‘Kitamoe’ and the Alpine non-resistant variety “Münstertaler” (K/M). The QTL explained 43.3% of total phenotypic variation for seed dormancy under greenhouse conditions. SSR markers flanking the QTL were assigned to the chromosome long arm fraction length 0.59–0.66 on the basis of chromosome deletion analysis, suggesting that the gene(s) controlling seed dormancy are probably located within this region. Under greenhouse conditions, the QTL explained 28.5 and 39.0% of total phenotypic variation for seed dormancy in Haruyutaka/Leader (HT/L) and OS21-5/Haruyokoi (O/HK) populations, respectively. However, in field conditions, the effect was relatively low or not significant in both the K/M and HT/L populations. These markers were considered to be widely useful in common with various genetic backgrounds for improvement of seed dormancy through the use of marker-assisted selection. Further detailed research using near isogenic lines will be needed to define how this major QTL interacts with environmental conditions in our area.     

利用染色体片段代换系定位棉花抗黄萎病QTL

通过陆地棉与海岛棉杂交并与陆地棉回交,获得1个染色体片段代换系苏VR043,抗江苏非落叶型黄萎病菌系BP2。分子检测表明,苏VR043兼有陆地棉苏棉8号的遗传背景和海7124的D4染色体片段。以苏VR043和苏棉8号为亲本构建包含176个单株的F2群体,通过标记分析和F2:3家系抗病鉴定,发现抗病性状与标记NAU3392连锁,p值为2.3×10-12。根据棉花D组染色体测序结果,参照置换区段的基因组序列,合成92对SSR引物;PCR扩增分析发现,有5对引物在苏棉8号和苏VR043之间表现多态性,并将抗病主效QTL定位在标记ZHX32和NAU3392之间,标记间遗传距离为3.2 c M,QTL解释表型变异64.8%。同时参照棉花D组测序结果,提取对应的置换区段序列,预测包含63个基因,基因聚类分析表明7个基因参与抗逆反应,其中1个基因与抗病相关。     

Analysis of quantitative trait loci associated with leaflet types in two recombinant inbred lines of soybean

Leaf area, length and width affect the photosynthetic capability of a plant and so increasing the photosynthetic rate per unit leaf area may improve seed yield in soybean. In this study, simple sequence repeat (SSR) markers were used to identify the genomic regions significantly associated with the quantitative trait locus (QTL) that controls length, width and the length/width ratio of the terminal and lateral leaflet in two segregating F_2:10 recombinant inbred line (RIL) populations, ‘Keounolkong’ × ‘Shinpaldalkong’ (K/S) and ‘Keounolkong’ × ‘Iksan10’ (K/I). In the K/S population, one QTL was identified for terminal leaflet length (TLL), two for lateral leaflet length (LLL), four for terminal leaflet width (TLW), four for lateral leaflet width (LLW), two for terminal leaflet length/width ratio (TLR) and four for lateral leaflet length/width ratio (LLR), with total phenotypic variations of 7.43, 10.9, 26.57, 23.46, 20.25 and 23.31%, respectively. In the K/I population, two QTLs were identified for TLL, two for LLL, three for TLW, and two for LLW, four for TLR and two for LLR with total phenotypic variations of 29.89, 22.77, 18.5, 12.15, 22.96 and 17.85%, respectively. Only a few QTLs coincided among the leaflet traits and no relationships were observed between the two populations. Many QTLs were associated with leaflet traits but each single QTL made only a minimal contribution. Thus, pyramiding the favourable alleles for leaflet traits in soybean breeding programmes may accelerate vegetative growth and perhaps lead to higher yields by maximizing total photosynthetic performance.     

Genetic diversity of resistance genes controlling fusarium head blight with simple sequence repeat markers in thirty-six wheat accessions from east asian origin

Summary Fusarium head blight (FHB) is a serious disease of wheat worldwide that may cause substantial yield and quality losses. Breeding for FHB-resistant cultivars is the most cost-effective approach to control FHB. The objective of the present study was to determine the relationship of resistance between new resistant sources and Sumai 3 using five simple sequence repeat (SSR) markers closely linked to the major QTL for FHB resistance on chromosome arms 3BS and 6BS. All five SSR markers were highly polymorphic between Sumai 3 (and its derivatives) and susceptible Canadian wheat lines. Most of the Sumai 3-derived Chinese wheat accessions and three Canadian FHB-resistant lines had all the Sumai 3 SSR marker alleles on chromosome arms 3BS and 6BS. The Chinese landrace Wangshuibai and two Japanese accessions Nobeokabozu and Nyu Bai had the same banding patterns as Sumai 3 for all five SSR marker alleles, and another Chinese landrace Fangshanmai had three of the five SSR markers in common with Sumai 3, and therefore most likely carries the same QTL as Sumai 3 on 3BS and 6BS. The Brazilian cultivar Frontana had no alleles in common with Sumai 3 on either QTL, and the Chinese landrace Hongheshang had only one of the five SSR markers in common with Sumai 3, therefore likely carrying resistance genes different from Sumai 3. The Italian cultivar Funo is not the donor of either the 3BS QTL or 6BS QTL. All five SSR seem to be effective candidates for marker-assisted selection to increase the level of resistance to FHB in wheat breeding programs.     

Two RAPD markers linked to a major fertility restorer gene in pepper

Both major and minor genes control the restoring of fertility in the cytoplasmic male-sterility system in pepper (Capsicum annuum L.). Bulked segregant analysis (BSA) was applied to identify molecular markers linked to a major restorer gene (Rf) using the F₂ population of NiujiaojiaoNo.21 (rfrf)/Xiangtanwan (RfRf). Two random amplified polymorphic DNA (RAPD) markers linked to this allele were detected with 520 decamer primers with arbitrary sequences. OP13₁₄₀₀ is a tightly linked marker with a genetic distance of0.37 cm. OW19₈₀₀ is on the opposite side with a distance of 8.12 cm. Both markers were repeatable and easy to score. A panel of genotypes, including 13elite inbred lines with different fertility restoring ability, were assayed for the presence ofOP13₁₄₀₀ and OW19₈₀₀. The markers are absent in all sweet pepper lines, indicating that they will be most helpful for transferring Rf into sweet pepper lines. With the aid of these markers, the size of the backcross population for testcrosses can be minimized. Furthermore, these markers will be useful in genetic analysis of the minor genes. This revised version was published online in July 2006 with corrections to the Cover Date.     

Confirmation of a major QTL on chromosome 10 for maize kernel row number in different environments

Maize kernel row number (KRN) is an important agronomic trait. In this study, 13 quantitative trait loci (QTL) for maize KRN were identified in different environments using F_2:3 and F_2:4 populations developed from two inbred lines. These QTL are distributed on chromosomes 2,3,5,8 and 10, and the genic effects are additive or partially dominant. Using the BC₃F_2:3 populations developed from the same parental lines, QTL of KRN located on chromosomes 5 and 10 were also identified in two environments. Three BC₅F_2:3 populations were used to confirm the major QTL for KRN between ssr1430 and umc1077 on chromosome 10(qKRN10). This result will facilitate the fine mapping and map‐based cloning of this major QTL in the future.     

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.     

Identification of AFLP and RAPD markers linked to anthracnose resistance in grapes and their conversion to SCAR markers

Resistance to anthracnose or black spot ( Elsinoe ampelina ), a serious fungal pathogen in viticulture and table grape production, was investigated on 25 grape cultivars. Bioassays performed with culture filtrates produced by the pathogen revealed 14 resistant genotypes. In most plants resistance originated from Vitis labrucsa but also genotypes with V. rupestris and V. riparia  ×  V. rupestris background showed resistance. Genetic analysis was conducted in F₁, S₁ and BC₁ plants developed from various cultivars. In total, 326 F₁ plants were evaluated, 172 genotypes proofed to be resistant, whereas 154 were susceptible to anthracnose. A Mendelian segregation ratio of 1 : 1 (χ² = 0.30–0.65) indicating that anthracnose resistance is controlled by a single dominant gene. To facilitate the use of marker-assisted selection in grape-breeding PCR-based markers were developed by random amplified polymorphic DNA and amplified fragment length polymorphism in bulk segregant analysis. Finally, OPB 15₁₂₄₇ was developed as a sequence characterized amplified region marker being diagnostic for the locus of resistance to anthracnose in all resistant genotypes tested. Within the 25 grape cultivars OPB 15₁₂₄₇ is diagnostic in the genetic background of both V. labrucsa and V. rupestris and V. riparia  ×  V. rupestris .     

AFLP-based STS markers closely linked to a fertility restoration locus (Rfm1) for cytoplasmic male sterility in barley

The Rfm1 gene restores the fertility of the msm1 and msm² male‐sterile cytoplasms in barley. Rfm1 is located on the short arm of chromosome 6H. To develop molecular markers tightly linked to Rfm1 for use in sophisticated marker‐assisted selection and map‐based cloning, an amplified fragment‐length polymorphism (AFLP) marker system with isogenic lines and a segregating BC₁F₁ population was used. Nine hundred primer combinations were screened and a linkage map was constructed around the Rfm1 locus by using 25 recombinant plants selected from 214 BC₁F₁ plants. Three AFLP markers were identified, e34m2, e46m19 and e48m17, linked to the locus. The most closely linked markers were e34m2, at 1.0 cM distally and e46m19, at 1.1 cM proximally. The two AFLP markers were converted to dominant STS markers. These markers should accelerate programmes for breeding restorer lines and will be useful for map‐based cloning.     

RAPD markers linked to resistance to downy mildew disease in soybean

To determine and utilize RAPD markers linked to resistance to downymildew incited by Peronospora manshurica in soybean, a resistantcultivar `AGS129' was crossed to a susceptible cultivar `Nakhon Sawan 1'(NS1). F₂ and BC₁ populations were advanced from the F₁ and evaluatedfor resistance to the disease. ²-test demonstrated that the resistancewas controlled by a single dominant gene (Rpmx). Near-isogenic lines(NILs) and bulked segregant analysis (BSA) were used to identify RAPDmarkers linked to the gene. Six DNA bulks namely F₅(R), F₅(S),BC₆F₃(R), BC₆F₃(S), F₂(R) and F₂(S) were set up by pooling equalamount of DNA from 8 randomly selected plants of each disease responsetype. A total of 180 random sequence decamer oligonucleotide primerswere used for RAPD analysis. Primer OPH-02 (5 TCGGACGTGA 3 andOPP-10 (5 TCCCGCCTAC 3) generated OPH-02₁₂₅₀ and OPP-10₈₃₁fragments in donor parent and resistant bulks, but not in the recurrentparent and susceptible ones. Co-segregation analysis using 102 segregatingF₂ progenies confirmed that both markers were linked to the Rpmxgene controlling downy mildew disease resistance with a genetic distance of4.9 cm and 23.1 cm, respectively. Marker OPH-02₁₂₅₀ was presentin 13 of 16 resistant soybean cultivars and absent in susceptible cultivars,thus confirming a potential for MAS outside the mapping population.