Chromosome Segment Substitution Line Population Research Articles

Unravelling the genetic basis and regulation networks related to fibre quality improvement using chromosome segment substitution lines in cotton.

The elucidation of genetic architecture and molecular regulatory networks underlying complex traits remains a significant challenge in life science, largely due to the substantial background effects that arise from epistasis and gene-environment interactions. The chromosome segment substitution line (CSSL) is an ideal material for genetic and molecular dissection of complex traits due to its near-isogenic properties; yet a comprehensive analysis, from the basic identification of substitution segments to advanced regulatory network, is still insufficient. Here, we developed two cotton CSSL populations on the Gossypium hirsutum background, representing wide adaptation and high lint yield, with introgression from G. barbadense, representing superior fibre quality. We sequenced 99 CSSLs that demonstrated significant differences from G. hirsutum in fibre, and characterized 836 dynamic fibre transcriptomes in three crucial developmental stages. We developed a workflow for precise resolution of chromosomal substitution segments; the genome sequencing revealed substitutions collectively representing 87.25% of the G.barbadense genome. Together, the genomic and transcriptomic survey identified 18 novel fibre-quality-related quantitative trait loci with high genetic contributions and the comprehensive landscape of fibre development regulation. Furthermore, analysis determined unique cis-expression patterns in CSSLs to be the driving force for fibre quality alteration; building upon this, the co-expression regulatory network revealed biological relationships among the noted pathways and accurately described the molecular interactions of GhHOX3, GhRDL1 and GhEXPA1 during fibre elongation, along with reliable predictions for their interactions with GhTBA8A5. Our study will enhance more strategic employment of CSSL in crop molecular biology and breeding programmes.

Haplotype-resolved gapless genome and chromosome segment substitution lines facilitate gene identification in wild rice

The abundant genetic variation harbored by wild rice (Oryza rufipogon) has provided a reservoir of useful genes for rice breeding. However, the genome of wild rice has not yet been comprehensively assessed. Here, we report the haplotype-resolved gapless genome assembly and annotation of wild rice Y476. In addition, we develop two sets of chromosome segment substitution lines (CSSLs) using Y476 as the donor parent and cultivated rice as the recurrent parents. By analyzing the gapless reference genome and CSSL population, we identify 254 QTLs associated with agronomic traits, biotic and abiotic stresses. We clone a receptor-like kinase gene associated with rice blast resistance and confirm its wild rice allele improves rice blast resistance. Collectively, our study provides a haplotype-resolved gapless reference genome and demonstrates a highly efficient platform for gene identification from wild rice.

Identification of Candidate Genes for Salt Tolerance at Seedling Stage in Rice Using QTL-Seq and Chromosome Segment Substitution Line-Derived Population

Rice is a staple food for more than half of the world’s population. However, the pervasive problem of salinity is severely undermining rice production, especially in coastal and low-lying areas where soil salinization is widespread. This stress, exacerbated by climate change, necessitates the development of salt-tolerant rice varieties to ensure food security. In this study, an F2:3 population (n = 454) from a cross of KDML105 and its chromosome segment substitution line (CSSL) was used to identify genomic regions associated with salt tolerance at the seedling stage. Using the QTL-seq approach, a QTL significantly associated with salt tolerance was identified on chromosome 1. Annotation of candidate genes in this region revealed the potential regulators of salt tolerance, including MIKC-type MADS domain proteins, calmodulin-binding transcription factors, and NB-ARC domain-containing proteins. These and other identified genes provide insights into the genetic basis of salt tolerance. This study underscores the importance of using advanced genomics tools and CSSL populations in the study of complex traits such as salt tolerance in rice. Several candidate genes identified in this study could be used in further studies on molecular or physiological mechanisms related to the salt response and tolerance mechanism in rice. Additionally, these genes could also be utilized in plant breeding programs for salt tolerance.

A wild rice CSSL population facilitated identification of salt tolerance genes and rice germplasm innovation.

Salt stress is one of the major factors that limits rice production. Therefore, identification of salt-tolerant alleles from wild rice is important for rice breeding. In this study, we constructed a set of chromosome segment substitution lines (CSSLs) using wild rice as the donor parent and cultivated rice Nipponbare (Nip) as the recurrent parent. Salt tolerance germinability (STG) was evaluated, and its association with genotypes was determined using this CSSL population. We identified 17 QTLs related to STG. By integrating the transcriptome and genome data, four candidate genes were identified, including the previously reported AGO2 and WRKY53. Compared with Nip, wild rice AGO2 has a structure variation in its promoter region and the expression levels were upregulated under salt treatments; wild rice WRKY53 also has natural variation in its promoter region, and the expression levels were downregulated under salt treatments. Wild rice AGO2 and WRKY53 alleles have combined effects for improving salt tolerance at the germination stage. One CSSL line, CSSL118 that harbors these two alleles was selected. Compared with the background parent Nip, CSSL118 showed comprehensive salt tolerance and higher yield, with improved transcript levels of reactive oxygen species scavenging genes. Our results provided promising genes and germplasm resources for future rice salt tolerance breeding.

Evaluation of Grain-Filling-Related Traits Using Taichung 65 x DV85 Chromosome Segment Substitution Lines (TD-CSSLs) of Rice.

Grain yield of rice consists of sink capacity and grain filling. There are some genes known to contribute to sink capacity, but few genes associated with grain filling are known. We conducted a genetic analysis on yield-related traits by using a chromosome segment substitution line population that have introgression from DV85, an aus variety of rice, in the background of T65, a japonica variety. Refined whole-genome genotypes of the 43 TD-CSSLs were obtained by genotyping-by-sequencing. The effects of previously detected quantitative trait loci (QTLs), qNSC1 and qNSC2, were confirmed by the amount of non-structural carbohydrate (NSC) at 5 days after heading (DAH). The CSSL for qSWTR11, the QTL for decrease in shoot weight during the maturity stage, showed the highest NSC at 5 DAH and lowest at 35 DAH. The brown rice yield of these lines were not stably significant. Most of the sink-related traits correlated between the 2 tested years, but most of the grain-filling traits did not show correlation between the 2 years. Correlation analysis revealed that the sink capacity is stable and primarily determines the yield, and grain filling is more affected by the environment. In addition, biomass production before heading and during the maturity stage contributes to higher yield in TD-CSSLs, and the amount of translocation of stem reserve does not affect much to the yield. We conclude that higher NSC at the heading stage and rapid decrease in shoot biomass during the maturity stage did not directly contribute to the yield formation in the japonica genetic background.

A QTL allele from wild soybean enhances protein content without reducing the oil content

Abstract Soybean is one of the chief crops producing protein and oil for human consumption. Wild soybean, the ancestor of cultivated soybean, possesses high seed protein content; therefore, it is a valuable genetic resource that could enhance protein content in the cultivated varieties. To identify the genes responsible for increasing protein content in wild soybean, a population comprising 113 BC4F6 chromosome segment substitution lines (CSSL) was developed from a cross between soybean cultivar ‘Jackson’ and wild soybean accession JWS156-1. The CSSL population was cultivated in the field conditions for 3 years (2018, 2019 and 2020), and the seeds harvested from each line were analysed for protein and oil contents by InfraTec Nova instrument. Quantitative trait locus (QTL) analysis with 243 SSR markers identified 12 QTLs associated with seed protein, oil and protein + oil contents. Among these QTLs, qPro8 and qPro19, two major and stable QTLs for protein content, were detected on chromosomes 8 and 19, respectively. No QTL for oil content was detected in the vicinity of qPro19, indicating that qPro19 did not influence the seed oil content. The effect of qPro19 was validated using near-isogenic lines (NILs) of qPro19. By introducing the qPro19 allele from wild soybean into another soybean variety, ‘Tachiyutaka’, a BC4 line, T-678, that showed enhanced seed protein content, without reducing the seed oil content. This study implied that the qPro19 allele from wild soybean could be a potential genetic resource for breeding programmes aimed to improve soybean seed quality.

Fine mapping and candidate gene analysis of proportion of four-seed pods by soybean CSSLs.

Soybean yield, as one of the most important and consistent breeding goals, can be greatly affected by the proportion of four-seed pods (PoFSP). In this study, QTL mapping was performed by PoFSP data and BLUE (Best Linear Unbiased Estimator) value of the chromosome segment substitution line population (CSSLs) constructed previously by the laboratory from 2016 to 2018, and phenotype-based bulked segregant analysis (BSA) was performed using the plant lines with PoFSP extreme phenotype. Totally, 5 ICIM QTLs were repeatedly detected, and 6 BSA QTLs were identified in CSSLs. For QTL (qPoFSP13-1) repeated in ICIM and BSA results, the secondary segregation populations were constructed for fine mapping and the interval was reduced to 100Kb. The mapping results showed that the QTL had an additive effect of gain from wild parents. A total of 14 genes were annotated in the delimited interval by fine mapping. Sequence analysis showed that all 14 genes had genetic variation in promoter region or CDS region. The qRT-PCR results showed that a total of 5 candidate genes were differentially expressed between the plant lines having antagonistic extreme phenotype (High PoFSP > 35.92%, low PoFSP< 17.56%). The results of haplotype analysis showed that all five genes had two or more major haplotypes in the resource population. Significant analysis of phenotypic differences between major haplotypes showed all five candidate genes had haplotype differences. And the genotypes of the major haplotypes with relatively high PoFSP of each gene were similar to those of wild soybean. The results of this study were of great significance to the study of candidate genes affecting soybean PoFSP, and provided a basis for the study of molecular marker-assisted selection (MAS) breeding and four-seed pods domestication.

Characterization of chromosome segment substitution lines reveals candidate genes associated with the nodule number in soybean

Soybean is one of the most important food crops worldwide. Like other legumes, soybean can form symbiotic relationships with Rhizobium species. Nitrogen fixation of soybean via its symbiosis with Rhizobium is pivotal for sustainable agriculture. Type III effectors (T3Es) are essential regulators of the establishment of the symbiosis, and nodule number is a feature of nitrogen-affected nodulation. However, genes encoding T3Es at quantitative trait loci (QTLs) related to nodulation have rarely been identified. Chromosome segment substitution lines (CSSLs) have a common genetic background but only a few loci with heterogeneous genetic information; thus, they are suitable materials for identifying candidate genes at a target locus. In this study, a CSSL population was used to identify the QTLs related to nodule number in soybean. Single nucleotide polymorphism (SNP) markers and candidate genes within the QTLs interval were detected, and it was determined which genes showed differential expression between isolines. Four candidate genes (GmCDPK28, GmNAC1, GmbHLH, and GmERF5) linked to the SNPs were identified as being related to nodule traits and pivotal processes and pathways involved in symbiosis establishment. A candidate gene (GmERF5) encoding a transcription factor that may interact directly with the T3E NopAA was identified. The confirmed CSSLs with important segments and candidate genes identified in this study are valuable resources for further studies on the genetic network and T3Es involved in the signaling pathway that is essential for symbiosis establishment.

Insights into the Regulation of Rice Seed Storability by Seed Tissue-Specific Transcriptomic and Metabolic Profiling.

Non-dormant seeds are continuously aging and deteriorating during storage, leading to declining seed vigor, which is a challenge for the rice seed industry. Improving the storability of seeds is of great significance to ensure the quality of rice and national food security. Through a set of chromosome segment substitution lines population constructed using japonica rice NIP as donor parent and indica rice ZS97 as recurrent parent, we performed seed storability QTL analysis and selected four non-storable NILs to further investigate the storability regulatory mechanisms underlying it. The seeds were divided into four tissues, which were the embryo, endosperm, aleurone layer, and hull, and tissue-specific transcriptome and metabolome analyses were performed on them. By exploring the common differentially expressed genes and differentially accumulated metabolites, as well as the KEGG pathway of the four non-storable NILs, we revealed that the phenylpropanoid biosynthesis pathway and diterpenoid biosynthesis pathway played a central role in regulating seed storability. Integrated analysis pinpointed 12 candidate genes that may take part in seed storability. The comprehensive analysis disclosed the divergent and synergistic effect of different seed tissues in the regulation of rice storability.

Precise genetic mapping of Rf18(t), a new fertility restorer gene from 'Nipponbare' for wild abortive cytoplasmic male sterility in rice (Oryza sativa L.).

We mapped Rf18(t), a Restorer-of-fertility gene for wild abortive cytoplasmic male sterility from the japonica maintainer 'Nipponbare', to chromosome 1. The best candidate gene, LOC_Os01g71320, is predicted to encode hexokinase. Three-line hybrid rice obtained through cytoplasmic male sterility (CMS) has helped increase the yield of rice globally, and the wild abortive (WA)-type cytoplasm from wild rice (Oryza rufipogon Griff.) is used widely in three-line indica hybrids. The identification and mapping of the Restorer-of-fertility (Rf) genes in maintainer lines aided in uncovering the genetic basis of fertility restoration of WA-type CMS and the development of WA-type hybrids. In this study, we identified a new Rf gene, Rf18(t), for WA-type CMS from the japonica maintainer line 'Nipponbare' using a chromosome segment substitution line population derived from a cross between the indica line 9311 and 'Nipponbare.' Using a substitution mapping strategy, Rf18(t) was delimited to a 48-kb chromosomal region flanked by molecular marker loci ID01M28791 and ID01M28845 on chromosome 1. By comparative sequence analyses, we propose that LOC_Os01g71320 is the most likely candidate gene for Rf18(t), and it is predicted to encode hexokinase. Furthermore, Rf18(t) was found to function in fertility restoration probably by a posttranscriptional mechanism and its function is dependent on the genetic background of 9311. These results broaden our knowledge on the mechanism of fertility restoration of WA-type CMS lines and will facilitate the development of WA-type rice hybrids.

Detection of QTLs for Plant Height Architecture Traits in Rice (Oryza sativa L.) by Association Mapping and the RSTEP-LRT Method.

Plant height (PH) and its component traits are critical determinants of lodging resistance and strongly influence yield in rice. The genetic architecture of PH and its component traits were mined in two mapping populations. In the natural population composed of 504 accessions, a total of forty simple sequence repeat (SSR) markers associated with PH and its component traits were detected across two environments via association mapping. Allele RM305-210 bp on chromosome 5 for PH had the largest phenotypic effect value (PEV) (−51.42 cm) with a reducing effect. Allele RM3533-220 bp on chromosome 9 for panicle length and allele RM264-120 bp on chromosome 8 for the length of upper first elongated internode (1IN) showed the highest positive PEV. Among the elongated internodes with negative effects being desirable, the allele RM348-130 bp showed the largest PEV (−7.48 cm) for the length of upper second elongated internode. In the chromosome segment substitution line population consisting of 53 lines, a total of nine QTLs were detected across two environments, with the phenotypic variance explained (PVE) ranging 10.07–28.42%. Among the detected QTLs, q1IN-7 explained the largest PVE (28.42%) for the 1IN, with an additive of 5.31 cm. The favorable allele RM257-125 bp on chromosome 9 for the 1IN increasing was detected in both populations. The favorable alleles provided here could be used to shape PH architecture against lodging.

Whole-Genome Sequencing of 117 Chromosome Segment Substitution Lines for Genetic Analyses of Complex Traits in Rice

Rice is one of the most important food crops in Asia. Genetic analyses of complex traits and molecular breeding studies in rice greatly rely on the construction of various genetic populations. Chromosome segment substitution lines (CSSLs) serve as a powerful genetic population for quantitative trait locus (QTL) mapping in rice. Moreover, CSSLs containing target genomic regions can be used as improved varieties in rice breeding. In this study, we developed a set of CSSLs consisting of 117 lines derived from the recipient ‘Huanghuazhan’ (HHZ) and the donor ‘Basmati Surkb 89–15’ (BAS). The 117 lines were extensively genotyped by whole-genome resequencing, and a high-density genotype map was constructed for the CSSL population. The 117 CSSLs covered 99.78% of the BAS genome. Each line contained a single segment, and the average segment length was 6.02 Mb. Using the CSSL population, we investigated three agronomic traits in Shanghai and Hangzhou, China, and a total of 25 QTLs were detected in both environments. Among those QTLs, we found that RFT1 was the causal gene for heading date variance between HHZ and BAS. RFT1 from BAS was found to contain a loss-of-function allele based on yeast two-hybrid assay, and its causal variation was a P to S change in the 94th amino acid of the RFT1 protein. The combination of high-throughput genotyping and marker-assisted selection (MAS) is a highly efficient way to construct CSSLs in rice, and extensively genotyped CSSLs will be a powerful tool for the genetic mapping of agronomic traits and molecular breeding for target QTLs/genes.

Identification and Fine-Mapping of a Novel QTL, qMrdd2, That Confers Resistance to Maize Rough Dwarf Disease.

Maize rough dwarf disease (MRDD) is caused by a virus and seriously affects maize quality and yield worldwide. MRDD can be most effectively controlled with disease-resistant hybrids of corn. Here, MRDD-resistant (Qi319) and -susceptible (Ye478) parental inbred maize lines and their 314 recombinant inbred lines (RILs) that were derived from a cross between them were evaluated across three environments. A stable resistance QTL, qMrdd2, was identified and mapped using best linear unbiased prediction (BLUP) values to a 0.55-Mb region between the markers MK807 and MK811 on chromosome 2 (B73 RefGen_v3) and was found to explain 8.6 to 11.0% of the total phenotypic variance in MRDD resistance. We validated the effect of qMrdd2 using a chromosome segment substitution line (CSSL) that was derived from a cross between maize inbred Qi319 as the MRDD resistance donor and Ye478 as the recipient. Disease severity index of the CSSL haplotype II harboring qMrdd2 was significantly lower than that of the susceptible parent Ye478. Subsequently, we fine-mapped qMrdd2 to a 315-kb region flanked by the markers RD81 and RD87, thus testing recombinant-derived progeny using selfed backcrossed families. In this study, we identified a novel QTL for MRDD resistance by combining the RIL and CSSL populations, thus providing important genetic information that can be used for breeding MRDD-resistant varieties of maize.

Identifying Wild Versus Cultivated Gene-Alleles Conferring Seed Coat Color and Days to Flowering in Soybean.

Annual wild soybean (G. soja) is the ancestor of the cultivated soybean (G. max). To reveal the genetic changes from soja to max, an improved wild soybean chromosome segment substitution line (CSSL) population, SojaCSSLP5, composed of 177 CSSLs with 182 SSR markers (SSR-map), was developed based on SojaCSSLP1 generated from NN1138-2(max)×N24852(soja). The SojaCSSLP5 was genotyped further through whole-genome resequencing, resulting in a physical map with 1366 SNPLDBs (SNP linkage-disequilibrium blocks), which are composed of more markers/segments, shorter marker length and more recombination breakpoints than the SSR-map and caused 721 new wild substituted segments. Using the SNPLDB-map, two loci co-segregating with seed-coat color (SCC) and six loci for days to flowering (DTF) with 88.02% phenotypic contribution were identified. Integrated with parental RNA-seq and DNA-resequencing, two SCC and six DTF candidate genes, including three previously cloned (G, E2 and GmPRR3B) and five newly detected ones, were predicted and verified at nucleotide mutant level, and then demonstrated with the consistency between gene-alleles and their phenotypes in SojaCSSLP5. In total, six of the eight genes were identified with the parental allele-pairs coincided to those in 303 germplasm accessions, then were further demonstrated by the consistency between gene-alleles and germplasm phenotypes. Accordingly, the CSSL population integrated with parental DNA and RNA sequencing data was demonstrated to be an efficient platform in identifying candidate wild vs. cultivated gene-alleles.

Genetic analysis of rice seed recovery under low-temperature conditions using a new CSSL population with a high-density genetic map in rice

Developing new population types based on interspecific introgressions has been suggested to facilitate the discovery of novel allelic sources for traits of agronomic importance. In this study, a chromosome segment substitution line (CSSL) rice population carrying contiguous chromosomal segments of japonica Nipponbare in the genetic background of indica 9311 was developed. Meanwhile, a high-density genetic map including 655 bins covering a distance of 1480.2 Mb, with an average of 0.60 Mb between markers, was generated through a whole-genome resequencing approach. Compared with previous CSSL populations constructed from 9311 or Nipponbare, this population has a higher marker density and can effectively meet the requirements of high QTL mapping resolution. On the basis of the newly constructed high-density genetic map, a total of three major QTLs (qGP4, qGP5, and qBL4) conferring seed growth capacity after low-temperature (5 °C) treatment were detected. Among these, two QTLs shared by Nipponbare, qGP4 and qGP5, could increase germination percentage, while one QTL shared by Nipponbare, qBL4, could increase bud length after cold stress; qGP4 and qBL4 were first reported in our study. These QTLs could be highly valuable genetic factors for cold tolerance improvement in rice lines. The CSSLs carrying these QTLs not only provide an opportunity for map-based cloning of important cold tolerance QTLs but also supply useful inbred lines to improve cold tolerance in 9311. These CSSLs represent good sources of japonica tolerance QTLs (genes) for use in improving cold tolerance in indica rice directly, avoiding the genetic drag arising from incompatible epistasis effects.

Fine-Mapping of a Wild Genomic Region Involved in Pod and Seed Size Reduction on Chromosome A07 in Peanut (Arachis hypogaea L.).

Fruit and seed size are important yield component traits that have been selected during crop domestication. In previous studies, Advanced Backcross Quantitative Trait Loci (AB-QTL) and Chromosome Segment Substitution Line (CSSL) populations were developed in peanut by crossing the cultivated variety Fleur11 and a synthetic wild allotetraploid (Arachis ipaensis × Arachis duranensis)4x. In the AB-QTL population, a major QTL for pod and seed size was detected in a ~5 Mb interval in the proximal region of chromosome A07. In the CSSL population, the line 12CS_091, which carries the QTL region and that produces smaller pods and seeds than Fleur11, was identified. In this study, we used a two-step strategy to fine-map the seed size QTL region on chromosome A07. We developed new SSR and SNP markers, as well as near-isogenic lines (NILs) in the target QTL region. We first located the QTL in ~1 Mb region between two SSR markers, thanks to the genotyping of a large F2 population of 2172 individuals and a single marker analysis approach. We then used nine new SNP markers evenly distributed in the refined QTL region to genotype 490 F3 plants derived from 88 F2, and we selected 10 NILs. The phenotyping of the NILs and marker/trait association allowed us to narrowing down the QTL region to a 168.37 kb chromosome segment, between the SNPs Aradu_A07_1148327 and Aradu_A07_1316694. This region contains 22 predicted genes. Among these genes, Aradu.DN3DB and Aradu.RLZ61, which encode a transcriptional regulator STERILE APETALA-like (SAP) and an F-box SNEEZY (SNE), respectively, were of particular interest. The function of these genes in regulating the variation of fruit and seed size is discussed. This study will contribute to a better knowledge of genes that have been targeted during peanut domestication.

Detection of QTLs for Outcrossing-Related Traits in CSSL Population Derived from Primitive Japonica Accession Ludao in the Genetic Background of O. sativa spp. Japonica Restorer C-bao Using RSTEP-LRT Method

The outcrossing traits in rice (Oryza sativa L.) affect the yield of hybrid seed production. Using a cytoplasmic male sterile (CMS) line with good outcrossing traits, such as short flag leaf length (FLL), narrow flag leaf width (FLW), wide flag leaf angle (FLA), and elongated panicle neck length (PNL), for hybrid rice seed production, it is possible to avoid the procedure of cutting flag leaves and make the supplementary pollination feasible by machine. In this study, a japonica restorer C-bao as the receptor parent and a primitive japonica accession Ludao as the donor parent were used to construct a chromosome segment substitution line (CSSL) population. The CSSL population was used to detect quantitative trait loci (QTLs) for the four outcrossing traits using a likelihood ratio test based on the stepwise regression (RSTEP-LRT) method. The CSSL population constructed consisted of 163 lines covering 90.7% of the donor genome. Among the seven QTLs detected in the CSSL population, four QTLs were detected in both years. qFLL-4 explained 6.70% of the two-year-averaged phenotypic variance, and the alleles from Ludao decreased FLL 5.1 cm. qFLA-1.1 and qFLA-1.2 explained 7.85% and 21.29% of the 2-year-averaged phenotypic variance respectively, and the alleles from Ludao increased FLA 17.38° and 31.50°. qPNL-8 explained 8.87% of the 2-year-averaged phenotypic variance, and the alleles from Ludao increased PNL 4.44 cm. These favorable alleles identified could be used to improve the outcrossing traits of parents for hybrid rice seed production in rice.

Detection of QTLs for outcrossing-related traits in rice (Oryza sativa L.) by association mapping and the RSTEP-LRT method

The outcrossing traits in rice affect the yield of hybrid rice seed production. In this study, 30 quantitative trait loci (QTLs) were detected in a natural population composed of 522 accessions by using a mixed linear model for the four outcrossing-related traits in 2017 and 2018. We detected 3, 4, 9 and 14 QTLs for flag leaf length (FLL), flag leaf width (FLW), flag leaf angle (FLA) and panicle neck length (PNL), respectively, among which 26 QTLs were novel. The favorable alleles and their carrier varieties were noticed with the largest phenotypic effect value. For FLL, marker allele RM6997-105 bp on chromosome 4 carried by Yue96 cultivar showed the highest phenotypic effect value (PEV), allele RM562-280 bp on chromosome 1 carried by Wanzhongqiu genotype for FLW, allele RM152-375 bp on chromosome 8 in typical carrier variety Zhongzuo93 for FLA and allele RM134-150 bp on chromosome 7 carried by Yue34 cultivar for PNL showed the highest PEVs. Additionally, nine QTLs were detected for the 4 traits with percentage of phenotypic variance explained (PVE) ranging from 16.24% (RM21) to 28.75% (RM5479) using a chromosome segment substitution line (CSSL) population. Among these detected QTLs, qFLL-12 and qFLW-12 showed 19.85% and 19.57% PVE, respectively. Four QTLs (qFLA-10, qFLA-11, qFLA-12.1, qFLA-12.2) detected for FLA showed PVEs ranging from 16.24% to 28.75%. For PNL, qPNL-7, qPNL-12.1, qPNL-12.2 were observed with PVEs of 20.14%, 17.39% and 28.36%, respectively. The favorable allele RM125-145 bp on chromosome 7 for PNL was detected in both the natural population and CSSL population and was carried by Yue38 accession and parent Xiushui79. The detected favorable alleles could be used to improve target traits.

Using Maize Chromosome Segment Substitution Line Populations for the Identification of Loci Associated with Multiple Disease Resistance.

Southern Leaf Blight (SLB), Northern Leaf Blight (NLB), and Gray Leaf Spot (GLS) caused by Cochliobolus heterostrophus, Setosphaeria turcica, and Cercospora zeae-maydis respectively, are among the most important diseases of corn worldwide. Previously, moderately high and significantly positive genetic correlations between resistance levels to each of these diseases were identified in a panel of 253 diverse maize inbred lines. The goal of this study was to identify loci underlying disease resistance in some of the most multiple disease resistant (MDR) lines by the creation of chromosome segment substitution line (CSSL) populations in multiple disease susceptible (MDS) backgrounds. Four MDR lines (NC304, NC344, Ki3, NC262) were used as donor parents and two MDS lines (Oh7B, H100) were used as recurrent parents to produce eight BC3F4:5 CSSL populations comprising 1,611 lines in total. Each population was genotyped and assessed for each disease in replicated trials in two environments. Moderate to high heritabilities on an entry mean basis were observed (0.32 to 0.83). Several lines in each population were significantly more resistant than the MDS parental lines for each disease. Multiple quantitative trait loci (QTL) for disease resistance were detected for each disease in most of the populations. Seventeen QTL were associated with variation in resistance to more than one disease (SLB/NLB: 2; SLB/GLS: 7; NLB/GLS: 2 and 6 to all three diseases). For most populations and most disease combinations, significant correlations were observed between disease scores and also between marker effects for each disease. The number of lines that were resistant to more than one disease was significantly higher than would be expected by chance. Using the results from individual QTL analyses, a composite statistic based on Mahalanobis distance (Md) was used to identify joint marker associations with multiple diseases. Across all populations and diseases, 246 markers had significant Md values. However further analysis revealed that most of these associations were due to strong QTL effects on a single disease. Together, these findings reinforce our previous conclusions that loci associated with resistance to different diseases are clustered in the genome more often than would be expected by chance. Nevertheless true MDR loci which have significant effects on more than one disease are still much rarer than loci with single disease effects.

QSE7 is a major quantitative trait locus (QTL) influencing stigma exsertion rate in rice (Oryza sativa L.)

Stigma exsertion is a key determinant to increase the efficiency of commercial hybrid rice seed production. The major quantitative trait locus (QTL) qSE7 for stigma exsertion rate was previously detected on the chromosome 7 using 75 Chromosome Segment Substitution Lines (CSSLs) derived from a cross between the high stigma exsertion indica maintainer XieqingzaoB (XQZB) and low stigma exsertion indica restorer Zhonghui9308 (ZH9308). The C51 line, a CSSL population with an introgression from XQZB, was backcrossed with ZH9308 to produce the secondary F2 (BC5F2) and F2:3 (BC5F2:3) populations. As a result, the Near Isogenic Line (NIL qSE7XB) was developed. Analysis indicated qSE7 acted as a single Mendelian factor and decreased the stigma exsertion. We hypothesized qSE7 regulates single, dual, and total stigma exsertion rate, provided experimental support. qSE7 was mapped and localized between RM5436 and RM5499 markers, within a physical distance of 1000-kb. With use of new insertion-deletion (InDel) markers and analysis of the heterozygous and phenotypic data, it was ultimately dissected to a 322.9-kb region between InDel SER4-1 and RM5436. The results are useful for additional identification and isolation of this candidate gene controlling stigma exsertion rate, and provide a basis for further fine mapping, gene cloning, and Marker Assisted Selection (MAS) breeding later.