Genetic analysis and phenotypic associations for drought tolerance in Hordeum spontaneum introgression lines using SSR and SNP markers

Associations between markers and drought related traits were investigated on a set of 57 advanced barley breeding lines, carrying various levels of introgression from Hordeum spontaneum lines 41-1 and 41-5, the best sources of drought tolerance in the ICARDA barley breeding program, using 74 simple sequences repeats (SSR) and 20 single nucleotide polymorphism markers. The 57 lines were evaluated for grain yield and drought related traits for three years (2003/04, 2004/05, 2005/06) in nine Mediterranean low rainfall environments. A high level of polymorphism was found with SSR markers, and the mean polymorphism information content and gene diversity values were 0.67 and 0.71, respectively. The number of alleles per locus varied from 2 to 11, with an average of 5.8 alleles per marker. Considering all the 57 lines, the linkage disequilibrium (LD) analysis was significant at a comparison-wise P?<?0.01 level in nearly 9 % of the SSR marker pairs used and a decay of LD was observed to a value of r ²?<?0.2 at a genetic distance of 40?cM. The association analysis revealed a total of 147 significant marker?Ctrait associations for grain yield and drought related traits. A total of 72 (49 %) marker?Ctrait associations showed favorable effects of the exotic germplasm where the H. spontaneum lines contributed to an improvement of the trait under drought stress conditions. The number of significant marker?Ctrait associations per trait were: 12 for growth habit; 2 for growth vigor; 11 for peduncle extrusion; 5 for number of grains per spike; 20 for peduncle length; 16 for days to heading; 20 for plant height; 8 for spike length; 17 for thousand kernel weight; 30 for grain yield; 4 for harvest index and 2 for biological yield. The phenotypic variation explained by individual marker?Ctrait associations ranged from 7.6 % to 36.2 %. The identification of genomic regions associated with grain yield and drought related traits is useful for the genetic improvement of cultivars better adapted to drought stress environments. Thus, the present study is encouraging in identifying significant marker?Ctrait associations through LD based association mapping analysis, which could complement and augment previous quantitative trait loci information for the potential use of Marker Assisted Selection for drought.     

Genome-wide InDel marker system for application in rice breeding and mapping studies

In order to widen the genetic basis for local rice breeding programs, a set of 506 PCR-based novel insertion/deletion (InDel) markers were designed. The markers have size differences larger than 30 base-pairs and were generated by published genome sequences of rice (Oryza sativa L.) indica and japonica subspecies. Among the 506 InDel markers, 133 markers are polymorphic between the selected parents Taiken 2 and Taichung Sen 10, which are elite Taiwanese japonica and indica varieties, respectively. The InDel to amplicon size ratios, based on the japonica template, were in the range of 8.8–45.6 %, with an average of 21.6 %. As a result, the PCR product can be efficiently separated and easily scored by running 30 min of non-submerging electrophoresis on 2–3 % agarose gels. Commonly observed non-parental bands originated from heteroduplex are considered beneficial in distinguishing homo- and hetero-zygotes. A linkage map constructed solely on these markers arranged in 12 linkage groups and extended over 1,413.9 cM with an average distance of 9.8 cM between markers, based on a segregating population of 286 F₂ individuals derived from a cross between Taiken 2 and Taichung Sen 10. The result indicated that InDel markers with moderate size differences were abundant and uniformly distributed across the whole genome of rice. Due to their simplicity in design and robustness in genotyping, these InDel markers have been routinely used in our quantitative trait loci mapping studies and marker-assisted selection programs for rice. We suggest that InDel markers with moderate size differences could be a valuable alternative in addition to the widely used SSR and SNP markers for rice breeding, and particularly suitable for field laboratories at breeding stations and/or laboratories with limited resources.     

Association analysis of sugar yield-related traits in sorghum [Sorghum bicolor (L.)]

Association mapping is widely used for detecting QTLs in higher plants. In the present study a synthetic sorghum population containing 119 representative samples, including 43 sweet and 76 grain sorghum accessions originating mainly from China, USA and India, were genotyped using 51 simple-sequence repeat (SSR) markers. Linkage disequilibrium (LD) of pair-wise loci and population structure were analyzed, followed by association analysis of SSR loci and 3 sugar yield related traits using the TASSEL general linear model program. Results showed that: (i) different degrees of LD occurred among syntenic markers and also among nonsyntenic markers, indicating historical recombination among sorghum linkage groups; (ii) significant LD extended up to 7.31 cM; (iii) the collection of accessions was composed of three subgroups; (iv) four marker loci were associated with stalk sugar concentration, fresh stalk weight and stalk juice weight measured in different growing environments and could be used, therefore, in future marker assisted breeding programs. Several loci were also associated with two or more traits simultaneously, which might be due to tight linkage between different genes affecting these traits and/or pleiotropy. In addition, some associated markers were located close to QTLs previously mapped in family-based linkage mapping analyses.     

Fine mapping of the uniform immature fruit color gene u in cucumber (Cucumis sativus L.)

Mottled/uniform color at the flower end of immature fruit is a highly important external quality trait that affects the market value of cucumber. Genetic analysis of different F₂ and backcross populations revealed that one single recessive gene, u (uniform immature fruit color), determines the uniform immature fruit color trait in cucumber. Based on earlier studies, the u locus is located on chromosome 5 (Chr. 5). By combining bulked segregant analysis using 60 published molecular markers on Chr. 5, we found that eight markers are polymorphic and are linked to the u locus. In addition, we developed five new relevant polymorphic simple sequence repeat (SSR) markers between markers SSR16203 and SSR15818. Subsequently, the F₂ population (477 individuals) from the cross of S06 (uniform fruit color line) × S94 (mottled fruit color line) was used for fine mapping of the u gene. The u gene was mapped to a 313.2-kb region between markers SSR10 and SSR27, at a genetic distance of 0.8 and 0.5 cM, respectively. Moreover, validity analysis of the codominant markers SSR10 and SSR27 was performed using 50 lines with mottled/uniform fruit color, demonstrating that these two SSR markers can be used for marker-assisted selection of the mottled/uniform fruit color trait in cucumber breeding. The results of this study will facilitate the cloning of the u gene.     

Construction of an integrated genetic map based on maternal and paternal lineages of tea (Camellia sinensis)

Genetic study on important traits of tea is difficult because of its self-incompatibility in nature. Moreover, development of a new variety usually needs more than 20 years, since it takes many years from seedling to matured plants for trait investigation. Genetic map is an essential tool for genetic study and breeding. In this study, we have developed an integrated genetic map of tea (Camellia sinensis) using a segregating F1 population derived from a cross between two commercial cultivars (‘TTES 19’ and ‘TTES 8’). A total of 574 polymorphic markers (including SSR, CAPS, STS, AFLP, ISSR and RAPD), 69 markers with highly significant levels of segregation distortion (P < 0.001) (12.0 %) were excluded from further analyses. Of the 505 mapped markers, there were 265 paternal markers (52.5 %), 163 maternal markers (32.3 %), 65 doubly heterozygous dominant markers (12.9 %), and 12 co-dominant markers (2.4 %). The co-dominant markers and doubly heterozygous dominant markers were used as bridge loci for the integration of the paternal and maternal maps. The integrated map comprised 367 linked markers, including 36 SSR, 3 CAPS, 1 STS, 250 AFLP, 13 ISSR and 64 RAPD that were assigned to 18 linkage groups. The linkage groups represented a total map length of 4482.9 cM with a map density of 12.2 cM. This genetic map has the highest genetic coverage so far, which could be applied to comparative mapping, QTL mapping and marker assisted selection in the future.     

Bulked segregant analysis identifies molecular markers associated with early bulking in cassava (Manihot esculenta Crantz)

Late root bulking is a major factor leading to rejection and abandoning of improved cassava genotypes in sub-Saharan Africa. Early bulking (EB) varieties shorten the growth period from planting to harvesting, better fit into environments with short rainy season, and reduce exposure to biotic and abiotic stresses thereby increasing productivity. This study was carried out to identify molecular markers linked to EB in cassava. Nine cassava hybrid populations (COB-1–COB-9) were developed using six elite varieties (TMS 30572, TMS 97/2205, TMS 98/0505, TMS 30555, NR 8212 and NR 8083) from the African cassava germplasm as parents. The progeny in each of the nine populations (101–272 genotypes per population) were evaluated for EB at 7 months after planting at seedling, clonal, and preliminary stages of breeding evaluation at Umudike. The parameters measured are fresh root yield, harvest index, fresh shoot weight and number of storage roots per plant. The progeny in each of the nine populations were genotyped at 542 simple sequence repeat (SSR) marker loci. Bulked segregant analysis was used to identify the SSR markers associated with EB in the populations. Nine SSR markers (SSRY 106, (ESTs)SSRY 292, SSRY 239, (ESTs)SSRY 7, NS 194, (ESTs)SSRY 47, SSRY 63, SSRY 250, and NS 323) were found to be closely linked (r = 0.3–0.5; p < 0.05) to EB in six of the nine hybrid populations. Seven of the markers with 10 % or more coefficient of determination (R²) were linked to major quantitative trait loci associated with EB in cassava. The molecular markers identified in this study provide useful materials to select for EB in cassava and for further target-traits-improvement by pyramiding.     

Association analysis of fibre traits in Gossypium arboreum accessions

Advances in the use of diploid Asiatic species in cotton breeding require an understanding of the relatedness and ancestry of diploid cotton accessions, and identification of simple sequence repeat (SSR) markers associated with agronomically important phenotypic traits, for example, fibre quality. Fifty‐six Gossypium arboreum germplasm accessions introduced from nine regions of Africa, Asia and Europe were evaluated for eight fibre characters (lint percentage, lint colour, elongation, micronaire, strength, 50% span length, 2.5% span length and maturity%) and genotyped with 98 SSR markers. When viewed across all accessions most of the SSR markers were polymorphic. Population structure analysis identified six main clusters for the accessions which corresponded to different geographic regions, indicating agreement between genetic and predefined populations. The general linear model method was used to disclose marker–trait associations. Marker–trait associations were investigated by fitting single marker regression models for phenotypic traits on marker band intensities with correction for population structure. This paper illustrates the potential of association mapping in diploid cotton, because existing phenotypic data, a modest number of SSR markers, and a pioneering statistical analysis, identified interesting associations.     

QTLs conferring FOV 7 resistance detected by linkage and association mapping in Upland cotton

Fusarium wilt is a worldwide disease that affects cotton production. Molecular markers tightly linked to resistance genes can be used for marker-assisted and/or genomic selection. We performed both family-based linkage mapping and population-based association mapping (AM) to detect quantitative trait loci (QTLs) conferring resistance against Fusarium oxysporum f. sp. vasinfectum race 7 (FOV 7) in Upland cotton. To identify QTLs underlying FOV 7 resistance by linkage mapping, three Upland cotton cultivars/lines, Xuzhou 142, Yumian 21 and Shang 9901, were used to obtain the composite cross population, designated as Xuzhou 142/Yumian 21//Xuzhou 142/Shang 9901. A linkage map containing 185 simple sequence repeat loci and 40 linkage groups was constructed with an average distance of 7.5 cM between adjacent markers. Seven QTLs were detected by linkage mapping, explaining 2.9–6.6 % of the total phenotypic variance. We also performed marker–trait AM with the MLM model (Q + K) in a panel composed of 356 Upland cotton cultivars. In total, 27 loci were significantly associated with FOV 7 resistance at the α = 0.01 level (?log₁₀ P ≥ 2), which were distributed on 16 chromosomes and explained 1.48–12.99 % of phenotypic variation. Three of the 7 QTLs identified by linkage mapping could be detected in AM. We identified the favorable allele for each of the 27 associated loci and investigated the number of favorable alleles in each accession. The results should increase our understanding of the genetic basis of FOV resistance and facilitate future resistance breeding in Upland cotton.     

Genetic diversity and association mapping of agronomic yield traits in eighty six synthetic hexaploid wheat

Association mapping is a method to identify associations between target traits and genetic markers based on linkage disequilibrium (LD) of a quantitative trait locus. Synthetic hexaploid wheat (SHW) is derived from a cross between Triticum durum Desf. and Aegilops tauschii Coss. that enhances genetic diversity and broadens breeding resources. In this study, phenotypic diversity in 110 wheat accessions (86 SHW germplasm specimens and 24 conventional wheat varieties) was evaluated quantitatively for yield characteristics of grains per spike, thousand kernel weight, and spike length. Phenotypic data were collected over two years at two locations, and 1785 alleles were detected (mean 6.59), ranging from 3 to 11 alleles per locus. The average genetic diversity index was 0.749, with a range from 0.239 to 0.923. The polymorphic information content (PIC) ranged from 0.145 to 0.968, with a mean value of 0.695. The genetic diversity index and PIC indicated that genome B > D > A. Accessions were grouped into three subgroups based on STRUCTURE and unweighted pair-group with arithmetic mean clustering. The mean LD decay across the genome was 11.78 cM. Association mapping between traits and simple sequence repeat markers was performed using the generalized linear model approach. Forty-six SSR loci were significantly associated with the measured agronomic traits in two geographic locations. Together, these results broaden our knowledge of how to harness elite genes and genetic diversity in SHW in genomic and marker-assisted selection.     

Tagging quantitative trait loci for heading date and plant height in important breeding parents of rice (Oryza sativa)

Heading date and plant height are important determinants for plant growth rate. In this study, simple sequence repeat markers were used to tag quantitative trait loci (QTL) using a recombinant inbred line mapping population derived from two important breeding parents, genetic stock Kaybonnetlpa1-1 and indica cultivar Zhe733, using data collected under field and greenhouse conditions. Interval mapping, composite interval mapping, and multiple interval mapping were performed to map QTL for heading date and plant height, and to identify epistatic interactions between the QTL. qHD3.1 on chromosome 3 from KBNTlpa1-1 had the largest effect on heading date contributing an average of 28.4 % of the total phenotypic variation. qHD7.1, 7.2, and 8.1 also had a significant contribution to heading date from Zhe733 averaging 8.1, 12.8, and 12.8 % of the phenotypic variance, respectively, and there was a positive additive-by-additive epistatic interaction between qHD7.1 and qHD8.1. QTL, qPHT1.1 and qPHT3.1, for plant height were detected on chromosomes 1 and 3, respectively. qPHT1.1 contributed the largest effect representing 38.2 % of the total phenotypic variation. Comparison of the QTL identified in our study with previous results revealed that the chromosomal locations for QTL coincided closely with positions reported previously in other rice populations worldwide, suggesting that these QTL have coevolved and become domesticated. The tightly linked SSR markers that flank these QTL should be desirable for tagging heading date and plant height genes and facilitating their incorporation into advanced breeding lines using marker assisted selection.     

Association mapping for seed cotton yield,yield components and fibre quality traits in upland cotton (Gossypium hirsutum L.) genotypes

Association mapping has revolutionized human genetics and is increasingly used in plant genetics. It is an efficient way of determining the genetic basis of complex traits. In cotton so far numerous association mapping studies are available as compared to many other important crops. In our study, mapping was performed using cotton 63K infinium beadchip with 201 germplasm lines. Through fast STRUCTURE analysis, lines were grouped into 12 subgroups and revealed genome sharing among the groups. The critical value (R²) was set to 0.243 as threshold to claim LD between two loci. About 3.13% marker pairs recorded significantly high LD (R² = 1), and 8.19% of marker pairs were in the range of 0.3 to 0.99 R². In MLM, 349 significant marker–trait associations were detected as against 642 in GLM because of effectiveness in eliminating false associations in MLM. A total of 68 markers explained >10% phenotypic variation for yield and fibre quality traits. Phenotypic variation explained by markers was smaller, suggesting that they might be associated with minor QTLs.     

Genetic mapping of the <Emphasis Type="Italic">qGCR6</Emphasis> locus affecting glossiness of cooked rice

A japonica variety, Koshihikari, is known to have favorable eating quality. Two rice backcross inbred lines (BILs) developed from Koshihikari exhibited significantly different glossiness of cooked rice (GCR), an eating quality trait measured using the Toyo-taste meter. Genetic analysis indicated that the genetic composition of these two BILs differed only on the short arm of chromosome 6, which led to the identification of the qGCR6 locus. Through high-resolution genetic mapping, the qGCR6 locus was further delimited to a 43.9 kb chromosomal region containing ten putative genes. The DNA marker SNP2175, which tightly links to qGCR6, was developed and can be used in marker-assisted breeding programs.     

A microsatellite-based genome-wide analysis of genetic diversity and linkage disequilibrium in Upland cotton (Gossypium hirsutum L.) cultivars from major cotton-growing countries

To better understand the genetic diversity of the cultivated Upland cotton (Gossypium hirsutum L.) and its structure at the molecular level, 193 Upland cotton cultivars collected from 26 countries were genotyped using 448 microsatellite markers. These markers were selected based on their mapping positions in the high density G. hirsutum TM-1 × G. barbadense 3-79 map, and they covered the whole genome. In addition, the physical locations of these markers were also partially identified based on the reference sequence of the diploid G. raimondii (D₅) genome. The marker orders in the genetic map were largely in agreement with their orders in the physical map. These markers revealed 1,590 alleles belonging to 732 loci. Analysis of unique marker allele numbers indicated that the modern US Upland cotton had been losing its genetic diversity during the past century. Linkage disequilibrium (LD) between marker pairs was clearly un-even among chromosomes, and among regions within a chromosome. The average size of a LD block was 6.75 cM at r ² = 0.10. A neighbor-joining phylogenic tree of these cultivars was generated using marker allele frequencies based on Nei’s genetic distance. The cultivars were grouped into 15 groups according to the phylogenic tree. Grouping results were largely congruent with the breeding history and pedigrees of the cultivars with a few exceptions.     

Genome-wide association study of yield components and fibre quality traits in a cotton germplasm diversity panel

A genome-wide association study (GWAS) was conducted on a diversity panel of 103 cotton accessions over three seasons to determine genetic contributions to a range of cotton yield components including fibre quality, plant architecture and stomatal conductance traits. The accessions covered breeding lines, released cultivars and some obsolete cultivars that contributed significantly to modern breeding pools. They were genotyped with Illumina’s CottonSNP63 K single nucleotide polymorphism (SNP) assay. Broad-sense heritability was low for yield component traits (\(h_{B}^{2}\) = 0.14–0.43), except for gin turnout and boll weight (\(h_{B}^{2}\)) = 0.74 and 0.59, respectively), and low to high for fibre quality traits (\(h_{B}^{2}\) = 0.26–0.89). Population structure analysis revealed extensive admixture and cryptic relatedness amongst the accessions. Genome-wide linkage disequilibrium (LD) analyses showed LD decayed, on average, within a physical distance of 5 Mbp and reduced to 2 Mbp at r ² ≥ 0.2, suggesting that few markers are necessary for association mapping in cotton. A mixed linear model accounting for population structure and cryptic relatedness identified 17 and 50 significant SNP associations for fibre length and micronaire, respectively. GWAS failed to detect significant associations in other traits, with the contribution of any single SNP to the phenotypic falling below 5%. This may be due to the low level of DNA polymorphism in cotton and/or insufficient resolution provided by the cotton SNP chip. Whole genome sequencing combined with whole genomic selection approaches that do not require prior knowledge about the effect or function of individual SNPs may be better suited than GWAS for trait dissection and prediction in cotton breeding.     

棉花品种资源群体结构与连锁不平衡分析

用基因组扫描的方法,利用棉花9个连锁群上的79个微卫星标记(Simple sequence repeat,SSR),对收集的204份陆地棉品种(系)组成的品种资源群体进行群体结构和连锁不平衡(Linkage disequilibrium,LD)分析.结果表明:本研究群体可划分为3个群体,其中两个群体分别由3个亚群体组成...     

Mining of favorable marker alleles for flag leaf inclination in some rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.) accessions by association mapping

Flag leaf angle (FLA) in rice (Oryza sativa L.) is one of the important traits affecting F₁ seed production by mechanization. To elucidate the genetic mechanism of FLA and mine favorable marker alleles for F₁ seed production in rice, we performed a genome-wide association study using phenotypic data over 2 years and genotypic data of 262 pairs of simple sequence repeat (SSR) markers collected from 441 rice accessions. We detected seven SSR marker loci associated with FLA and four loci were novel. The four newly found loci were RM6266 on chromosome 3, RM348 on chromosome 4, RM258 on chromosome 10 and RM7303 on chromosome 11. We found a total of 27 favorable alleles, of which four, i.e., RM348-130 bp, RM7303-90 bp, RM258-180 bp, and RM4835-230 bp, had phenotypic effects larger than 10°. Nine combinations, which increased FLA by 45.7°–94.7° through pyramiding the favorable alleles contained in seven typical accessions, were predicted.     

QTL analysis for eating quality-related traits in an F2:3 population derived from waxy corn × sweet corn cross

In order to identify quantitative trait loci (QTL) for the eating quality of waxy corn and sweet corn (Zea mays L.), QTL analysis was conducted on an F₂ population derived from a cross between a waxy corn inbred line and a sweet corn inbred line. Ten QTLs for pericarp thickness (PER), amylose content (AMY), dextrose content (DEX) and sucrose content (SUC) were found in the 158 F₂ families. Among them, four QTLs, qAMY4 (10.43%), qAMY9 (19.33%), qDEX4 (21.31%) and qSUC4 (30.71%), may be considered as major QTLs. Three of these, qAMY4, qDEX4 and qSUC4, were found to be located within a region flanked by two adjacent SSR markers on chromosome 4 (umc1088 and bnlg1265), making this SSR marker pair a useful selection tool for screening the eating quality traits of AMY, DEX and SUC. The QTL for amylose content was found to be located between markers phi027 and umc1634, raising the possibility of its identity being the Wx1 gene, which encodes a granule-bound amylose synthase. The new QTLs identified by the present study could serve as useful molecular markers for selecting important eating quality traits in subsequent waxy corn breeding studies.     

小麦南大2419及其衍生品种(系)主要农艺性状的关联分析

筛选与小麦重要农艺性状相关联的SSR标记,对小麦分子标记辅助育种有重要的实践意义.本研究利用多态性较高的80个SSR标记,对南大2419及其71份衍生后代品种(系)进行基因型分析,采用TASSEL软件的MLM (Mixed linear model)方法对籽粒产量、千粒重、有效穗、穗粒数等8个主要农艺性状进行SSR标记...     

Genetic linkage map of Chinese native variety faba bean (Vicia faba L.) based on simple sequence repeat markers

Simple sequence repeat (SSR) marker is a powerful tool for construction of genetic linkage map which can be applied for quantitative trait loci (QTL) and marker‐assisted selection (MAS). In this study, a genetic map of faba bean was constructed with SSR markers using a 129 F₂ individuals population derived from the cross of Chinese native variety 91825 (large seed) and K1563 (small seed). By screening 11 551 SSR primers between two parents, 149 primer pairs were detected polymorphic and used for F₂ population analysis. This SSR‐based genetic linkage map consisted of 15 linkage groups with 128 SSR. The map encompassed 1587 cM with an average genetic distance of 12.4 cM. The genetic map generated in this study will be beneficial for genetic studies of faba bean for identification of marker‐locus‐trait associations as well as comparative mapping among faba bean, pea and grasspea.     

Identification and fine mapping of molecular markers closely linked to fruit spines size <Emphasis Type="Italic">ss</Emphasis> gene in cucumber (<Emphasis Type="Italic">Cucumis sativus</Emphasis> L.)

Fruit spine size is one of the importantly external quality traits effected the economic value of cucumber fruit. Morphological–cytological observation of the fruit spine size phenotype indicated that large spine formation arises from an increasing of spiny pedestal cell number caused by cell division, and best periods to accurately score fruit spine size trait was 4th day before flowering to 7th day after flowering according the continuous observation. Genetic analysis showed that a single dominant gene determined the fruit spine size trait in cucumber. BC₁ population (189 individuals) of two inbred lines (large spine PI197088 and small spine SA0422) was used for primary mapping of the SS/ss locus with 7 markers covering an interval of 37.1 cM. An F₂ segregating population of 1032 individuals constructed from the same two parents (PI197088 and SA0422) was used to fine mapping of the SS/ss locus. Six new markers linked to the gene were successfully screened for construction of a fine linkage map, in which the SS/ss locus was located in the region flanked by marker SE1 (3 recombinants) and SSR43 (2 recombinants) with a 189 kb physical distance. Markers from this study will be valuable for candidate gene cloning and marker-assisted selection for cucumber breeding.