Genome-wide association mapping and genomic prediction of stalk rot in two mid-altitude tropical maize populations

Maize stalk rot reduces grain yield and quality.Information about the genetics of resistance to maize stalk rot could help breeders design effective breeding strategies for the trait.Genomic prediction may be a more effective breeding strategy for stalk-rot resistance than marker-assisted selection.We performed a genome-wide association study(GWAS)and genomic prediction of resistance in testcross hybrids of 677 inbred lines from the Tuxpe?o and non-Tuxpe?o heterotic pools grown in three environments and genotyped with 200,681 single-nucleotide polymorphisms(SNPs).Eighteen SNPs associated with stalk rot shared genomic regions with gene families previously associated with plant biotic and abiotic responses.More favorable SNP haplotypes traced to tropical than to temperate progenitors of the inbred lines.Incorporating genotype-by-environment(G×E)interaction increased genomic prediction accuracy.

QTL Identification of the Insensitive Response to Photoperiod and Temperature in Soybean by Association Mapping

The insensitive response to photoperiod and temperature is an important quantitative trait for soybean in wide adaptation breeding. The natural variation in response to photoperiod and temperature was detected using 275 accessions of soybean [Glycine max （L.） Merrill] from China. Genome-wide association mapping, based on population structure analysis, was carried out using 118 SSR markers by the TASSEL GLM （general linear model） program. Nine SSR markers （P〈0.01） were associated with the value of the response to photoperiod and temperature （VRPT） caused by days to flowering （DF）, among which, Satt308 （LG M）, Sattl50 （LG M） and Satt440 （LG l）, were identified in both 2006 and 2007. Twelve SSR markers （P〈0.01） were associated with VRPT caused by days to maturity （DM）, among which three markers, Satt387 （LG N）, Satt307 （LG C2） and AW310961 （LG J）, were detected in both 2006 and 2007. In addition, a total of 20 elite alleles were screened out over 2006 and 2007 for being associated with an insensitive response to photoperiod and temperature （IRPT） caused by DF and a total of seven different elite alleles were screened out for being associated with IRPT caused by DM. Among these elite alleles, five alleles, Satt150-244, Satt308-164, Satt308-206, Satt440-176, and Satt440-206, were associated with IRPT caused by DF and were identified in both years, but only one allele, Satt307-170, was identified as being associated with an IRPT caused by DM. Based on these elite alleles, a set of typical accessions were screened out. The result about the genetic basis of IRPT is meaningful for soybean wide adaption breeding.

Genotype × Environment Interactions for Agronomic Traits of Rice Revealed by Association Mapping

Agronomic traits are important determinants to rice yield, which are controlled by complex genetic factors as well as genotype by environment （G × E） interaction effects. The G × E effects for agronomic traits of rice have been dissected with various approaches, but not with the current available approach, the association studies. In this study, a total of 32 655 single nucleotide polymorphisms were used to carry out associations with 14 agronomic traits among 20 rice accessions in two environments. The G × E interaction effects for all the agronomic traits were at highly significant levels （P〈0.01）, accounting for 3.4%-22.3% of the total sum of squares except for the length of brown rice. Twenty three putative quantitative trait loci （QTLs）, including five previously known and several new promising associations, were identified for 10 of 14 traits. Analysis of the relationships between the traits for which QTLs and the genotype effects could be identified suggested that the higher the genotypic effect, the higher the possibility to identify QTLs for the given trait. The new QTLs detected in this study will facilitate dissection of the complex agronomic traits and may give insight into the G × E effects with association mapping.

Association Mapping and Marker Development of Genes forStarch Lysophospholipid Synthesis in Rice

Phospholipids are a major kind of lipids in rice grains and have fundamental nutritional andfunctional benefits to the plant. Their lyso forms （lysophospholipids, LPLs） often form inclusion complexeswith amylose or independently influence the physicochemical and functional properties of rice starch.However, the genetic basis for LPL synthesis in rice endosperm is largely unknown. Here, we performeda preliminary association test of 13 LPL compositions among 20 rice accessions, and identified 22putative main-effect quantitative trait loci responsible for all LPLs except for LPC14：0 and LPE14：0. Fivederived cleaved amplified polymorphic sequences and one insertion/deletion marker for threeLPL-synthesis-related candidate genes were developed. Association analysis revealed two markerssignificantly associated with starch LPL traits. These results provide an insight into the genetic basis ofphospholipid biosynthesis in rice and may contribute to the rice quality breeding programs usingfunctional markers.

Combined linkage and association mapping reveals two major QTL for stripe rust adult plant resistance in Shaanmai 155 and their haplotype variation in common wheat germplasm

The development and deployment of diverse resistance sources in new wheat cultivars underpin the durable control of stripe rust.In the present study,two loci for adult plant resistance(APR),QYr SM155.1 and QYr SM155.2,were identified in the Chinese wheat breeding line Shaanmai 155.QYr SM155.1 was mapped to a 3.0-c M interval between the single-nucleotide polymorphism(SNP)markers AX-109583610 and AX-110907562 on chromosome arm 2 BL.QYr SM155.2 was mapped to a 2.1-c M interval flanked by the SNP markers AX-110378556 and AX-86173526 on chromosome arm 7 AS.A genome-wide association study was used to identify markers associated with APR in a panel of 411 spring wheat lines.Thirteen and 11 SNPs were significantly associated with QYr SM155.1 and QYr SM155.2,respectively,corresponding to physical intervals of 653.75–655.52 Mb on 2 BL and 81.63–83.93 Mb on7 AS.To characterize the haplotype variation and the distribution of these QTL,haplotype analysis was performed based on these SNPs in an independent panel of 1101 worldwide wheat accessions.Three major haplotypes(2 B_h1,2 B_h2,and 2 B_h3)for QYr SM155.1 and four major haplotypes(7 A_h1,7 A_h2,7 A_h3,and 7 A_h4)for QYr SM155.2 were identified.Accessions individually harboring QYr SM155.1_h1 and QYr SM155.2_h1 haplotypes and their combination displayed resistance.Additional assays of 1306 current Chinese cultivars and breeding lines using markers flanking QYr SM155.1 and QYr SM155.2 indicated that the resistance haplotypes of the two QTL were present in respectively 1.45%and 14.16%of lines.Increasing resistance haplotype frequencies at these two loci using marker-assisted selection should benefit wheat production in China.

Population structure and association mapping to detect QTL controlling tomato spotted wilt virus resistance in cultivated peanuts

Tomato spotted wilt(TSW)is a serious virus disease of peanut in the United States.Breeding for TSWV resistance would be facilitated by the implementation of marker-assisted selection in breeding programs;however,genes associated with resistance have not been identified.Association mapping is a type of genetic mapping that can exploit relationships between markers and traits in many lineages.The objectives of this study were to examine genetic diversity and population structure in the U.S.peanut mini-core collection using simple sequence repeat(SSR)markers,and to conduct association mapping between SSR markers and TSWV resistance in cultivated peanuts.One hundred and thirty-three SSR markers were used for genotyping 104 accessions.Four subpopulations,generally corresponding to botanical varieties,were classified by population structure analysis.Association mapping analysis indicated that five markers:pP GPseq5D5,GM1135,GM1991,TC23C08,and TC24C06,were consistently associated with TSW resistance by the Q,PCA,Q+K,and PCA+K models.These markers together explained 36.4%of the phenotypic variance.Moreover,pP GPseq5D5 and GM1991 were associated with both visual symptoms of TSWV and ELISA values with a high R^2.The potential of these markers for use in a marker-assisted selection program to breed peanut for resistance to TSWV is discussed.

Mining Elite Alleles of Growth Duration and Productive Panicle Number per Plant by Association Mapping with Conditional Phenotypic Value in Japonica Rice

To provide genetic information and materials for breeding hybrid japonica rice with wide adaptability and strong competitive advantage of yield, elite alleles and their carrier varieties of growth duration （GD） and productive panicle number per plant （PN） were detected. A natural population composed of 94 japonica varieties was phenotyped for the GD, PN and plant height （PH） in two environments. The conditional phenotypic data were transferred by the linear model method in software QGAStation 1.0, and association mapping based on the unconditional and conditional phenotype values of GD and PN was analyzed by using general linear model in software TASSEL. A total of 34 simple sequence repeat （SSR） marker loci associated with GD and PN were detected in the two environments. Among them, 15 were associated with GD, and 19 were associated with PN. Four elite alleles of RM8095-120bp, RM7102-176bp, RM72-170bp and RM72-178bp were associated with GD, and their carrier varieties were Hongmangshajing, Nipponbare, Hongmangshajing and Nannongjing 62401, respectively. These elite alleles from the carrier varieties can shorten GD by 2.03-9.93 d when they were introduced into improved materials. RM72-182bp associated with PN was an elite allele, and its carrier variety was Xiaoqingzhong. It can increase PN by three when introduced into improved materials. Moreover, these elite alleles can be used to improve target traits without influencing another two traits.

Genetic and Association Mapping Study of English Grain Aphid Resistance and Tolerance in Bread Wheat Germplasm

English grain aphid （EGA） is a destructive insect pest of wheat. To identify the loci associated with EGA resistance and tolerance, 70 bread wheat accessions mainly from central Asia were evaluated for EGA resistance and tolerance traits at two locations, and genotyped with 51 SSR markers. Totally, three accessions showed high or moderate levels resistance and 17 genotypes displayed highly or moderately tolerate to EGA. Genetic diversity of these lines was investigated also. After 97 SSR loci which evenly covered all wheat chromosomes were scanned for association, four SSR loci were significantly associated with EGA resistance and four with EGA tolerance. After association analysis was conducted with dynamic aphid densities, we found four loci Xgwm192b, Xgwm391, Xbarc98, and Xgwm613b were detected continuously at different growing stages of wheat. In addition, the loci of EGA resistance/tolerance and Russian wheat aphid resistance were compared. The results generated in this study would be helpful for utilization of the EGA resistance/tolerance germplasm, and for development of mapping populations in EGA resistance breeding programs.

Dissecting the genetic basis of maize deep-sowing tolerance by combining association mapping and gene expression analysis

Deep-sowing is an important method for avoiding drought stress in crop species,including maize.Identifying candidate genes is the groundwork for investigating the molecular mechanism underlying maize deep-sowing tolerance.This study evaluated four traits(mesocotyl length at 10 and 20 cm planting depths and seedling emergence rate on days 6 and 12)related to deep-sowing tolerance using a large maize population containing 386 inbred lines genotyped with 0.5 million high-quality single nucleotide polymorphisms(SNPs).The genomewide association study detected that 273 SNPs were in linkage disequilibrium(LD)with the genetic basis of maize deep-sowing tolerance.The RNA-sequencing analysis identified 1944 and 2098 differentially expressed genes(DEGs)in two comparisons,which shared 281 DEGs.By comparing the genomic locations of the 273 SNPs with those of the 281 DEGs,we identified seven candidate genes,of which GRMZM2G119769 encoded a sucrose non-fermenting 1 kinase interactor-like protein.GRMZM2G119769 was selected as the candidate gene because its homologs in other plants were related to organ length,auxin,or light response.Candidate gene association mapping revealed that natural variations in GRMZM2G119769 were related to phenotypic variations in maize mesocotyl length.Gene expression of GRMZM2G119769 was higher in deep-sowing tolerant inbred lines.These results suggest that GRMZM2G119769 is the most likely candidate gene.This study provides information on the deep-sowing tolerance of maize germplasms and identifies candidate genes,which would be useful for further research on maize deep-sowing tolerance.

Association mapping of quantitative trait loci for yield-related agronomic traits in rice(Oryza sativa L.)

High yield in rice mainly depends on large grain weight, ideal plant architecture and proper flowering time adapting to various geographic regions. To help achieve higher yield, phenotype variations of heading date（HD）, plant architecture and grain shape in a panel of 416 rice accessions were investigated in this study. A total of 143 markers including 100 simple sequence repeat（SSR） markers and 43 gene-tagged markers were employed in association mapping to detect quantitative trait loci（QTL） responsible for these variations. Among the 7 subpopulations, POP5 in japonica group showed the largest values of HD and grain width（GW）, but the smallest values of grain length（GL） and grain length to width ratio（GLW）. Among the six indica groups, POP7 had the largest values of HD, GL, GLW, and 1 000-grain weight（TGW）. A total of 27 QTLs were detected underlying these phenotypic variations in single year, while 12 of them could be detected in 2006 and 2007. GS3 marker was closely associated with GL, GW and GLW, and widely distributed in different groups. The starch synthesis related gene markers, SSI, SSIIa, SBE1, AGPL4, and ISA1, were linked to plant height（PH）, panicle length（PL）, flag leaf length（FLL）, GW, and GLW. The SSR markers, RM267, RM340 and RM346, were linked to at least two traits. Therefore, these new markers will probably be used to improve rice grain yield or plant architecture when performing marker-assisted selection of proper alleles.

Deletion of a 1,049 bp sequence from the 5′UTR upstream of the SiHEC3 gene induces a seed non-shattering mutation in sesame

Sesame is a labor intensive crop with limited mechanized harvesting mainly due to the seed shattering(SS)trait.In this study,we performed a genetic analysis of the seed-shattering resistance(SR)trait with a SR sesame mutant 12M07.Unlike the SS type,the parenchyma cells in the abscission zone of the 12M07 mutant are arranged loosely but adhere to the seed coat.Inheritance analysis of six generations derived from 12M07(SR)×Xiangcheng Dazibai(SS)showed that the SR trait is recessive and controlled by a single gene pair.Association mapping of the F2population with 888,619 variants(single-nucleotide polymorphisms(SNPs)and insertion-deletion(InDels))and 31,884 structural variations(SVs)determined that only SV12002 in the 5′upstream region of gene Sindi0765000(named SiHEC3)in Chr.3 was significantly associated with the SR trait.SiHEC3 encodes the bHLH transcription factor.A 1,049 bp deletion occurred in the 5′UTR of Sihec3 in 12M07.SiHEC3 is mainly expressed in developing placental tissues,with the expression peaking in capsules at 45 days after pollination.A dual-luciferase reporter assay in tobacco confirmed that the promoter activity of Sihec3 was reduced because of the deletion of the 1,049 bp promoter sequence.Protein–protein interaction network analysis showed that HEC3 is co-expressed with nine key proteins,such as SHATTERPROOF1(SHP1)and SEEDSTICK(STK)which participate in the secondary wall biosynthesis of the abscission layer in plants.The findings of this study show the important function of Sihec3corresponding with the SR trait and supply the genetic information for breeding new varieties that are amenable to mechanized harvesting in sesame and other crops.

Diversity, Structure, and Marker-Trait Association Analysis of the Maize Recombinant Inbred Line Population

Association mapping has emerged as a new tool to elucidate complex quantitative trait loci in maize, but there are few reports about systematic association analysis for the specific SSR markers with agronomic traits of interest in China. We investigated the morphological and genetic diversity and population structure for 76 maize recombinant inbred lines, and then association analysis were further performed between 48 simple sequence repeat loci and 17 morphological traits, consisting of nine ear-related traits and eight other traits. The 48 SSR markers were screened out and further classified into two groups including a group of loci in regions harboring reported quantitative trait loci that affect ear shape and a group of markers distributing on the whole genome randomly. The result indicated that the population of recombinant inbred lines was structured, showing five subpopulations. Our association results revealed that there were 82, 59, and 40 significant associations detected by K-test, logistic regression, and both analysis, respectively. When the 17 traits were considered separately, the significant associations between Q-SSRs and E-traits were raised to 27.8%, whereas the other groups of combinations ranged between 2.3 and 6.3%. As the proportion of significant associations is higher among the Q-SSR subset of markers and the subset of traits related to ear shape than those for all of the other combinations, we conclude that this approach is valid for establishing true positive marker-trait relationships. Our results also demonstrated that association mapping could complement and enhance previous QTL information for marker-assisted selection.

An Adaptive Sequential Replacement Method for Variable Selection in Linear Regression Analysis

With the rapid development of DNA technologies, high throughput genomic data have become a powerful leverage to locate desirable genetic loci associated with traits of importance in various crop species. However, current genetic association mapping analyses are focused on identifying individual QTLs. This study aimed to identify a set of QTLs or genetic markers, which can capture genetic variability for marker-assisted selection. Selecting a set with k loci that can maximize genetic variation out of high throughput genomic data is a challenging issue. In this study, we proposed an adaptive sequential replacement (ASR) method, which is considered a variant of the sequential replacement (SR) method. Through Monte Carlo simulation and comparing with four other selection methods: exhaustive, SR method, forward, and backward methods we found that the ASR method sustains consistent and repeatable results comparable to the exhaustive method with much reduced computational intensity.

DISCRETE BIDIRECTIONAL ASSOCIATIVE MEMORY WITH LEARNING FUNCTION

In this paper we propose a new discrete bidirectional associative memory (DBAM) which is derived from our previous continuous linear bidirectional associative memory (LBAM). The DBAM performs bidirectionally the optimal associative mapping proposed by Kohonen. Like LBAM and NBAM proposed by one of the present authors,the present BAM ensures the guaranteed recall of all stored patterns,and possesses far higher capacity compared with other existing BAMs,and like NBAM, has the strong ability to suppress the noise occurring in the output patterns and therefore reduce largely the spurious patterns. The derivation of DBAM is given and the stability of DBAM is proved. We also derive a learning algorithm for DBAM,which has iterative form and make the network learn new patterns easily. Compared with NBAM the present BAM can be easily implemented by software.

Linkage Disequilibrium in Wild and Cultured Populations of Pacific Oyster(Crassostrea gigas)

Linkage disequilibrium(LD) can be applied for mapping the actual genes responsible for variation of economically important traits through association mapping.The feasibility and efficacy of association studies are strongly dependent on the extent of LD which determines the number and density of markers in the studied population,as well as the experimental design for an association analysis.In this study,we first characterized the extent of LD in a wild population and a cultured mass-selected line of Pacific oyster(Crassostrea gigas).A total of 88 wild and 96 cultured individuals were selected to assess the level of genome-wide LD with 53 microsatellites,respectively.For syntenic marker pairs,no significant association was observed in the wild population;however,three significant associations occurred in the cultured population,and the significant LD extended up to 12.7 c M,indicating that strong artificial selection is a key force for substantial increase of genome-wide LD in cultured population.The difference of LD between wild and cultured populations showed that association studies in Pacific oyster can be achieved with reasonable marker densities at a relatively low cost by choosing an association mapping population.Furthermore,the frequent occurrence of LD between non-syntenic loci and rare alleles encourages the joint application of linkage analysis and LD mapping when mapping genes in oyster.The information on the linkage disequilibrium in the cultured population is useful for future association mapping in oyster.

Molecular Tagging and Mapping of Quantitative Trait Loci for Lint Percentage and Morphological Marker Genes in Upland Cotton

Using 219 F2 Individuals developed by crossing the genetic standard line TM-1 and the multiple dominant marker line T586 In Gossyplum hirsutum L., a genetic linkage map with 19 linkage groups was constructed based on simple sequence repeat （SSR） markers. Compared with our tetraploid backboned molecular genetic map from a （TM-1xHal 7124）xTM-1 BC1 population, 17 of the 19 I｜nkage groups were combined and anchored to 12 chromosomes （sub-genomes）. Of these groups, four morphological marker genes In T586 had been mapped Into the molecular linkage map. Meanwhile, three quantitative trait loci for lint percentage were tagged and mapped separately on the A03 linkage group and chromosome 6.

Association analysis of fiber quality traits and exploration of elite alleles in Upland cotton cultivars/accessions(Gossypium hirsutum L.)

Exploring the elite al eles and germplasm acces-sions related to fiber quality traits wil accelerate the breeding of cotton for fiber quality improvement. In this study, 99 Gossypium hirsutum L. accessions with diverse origins were used to perform association analysis of fiber quality traits using 97 polymorphic microsatel ite marker primer pairs. A total of 107 significant marker-trait associations were detected for three fiber quality traits under three different environments, with 70 detected in two or three environments and 37 detected in only one environment. Among the 70 significant marker-trait associations, 52.86% were reported previously, implying that these are stable loci for target traits. Furthermore, we detected a large number of elite al eles associated simulta-neously with two or three traits. These elite al eles were mainly from accessions col ected in China, introduced to China from the United States, or rare al eles with a frequency of less than＆amp;nbsp;5%. No one cultivar contained more than half of the elite al eles, but 10 accessions were col ected from China and the two introduced from the United States did contain more than half of these al eles. Therefore, there is great potential for mining elite al eles from germplasm accessions for use in fiber quality improvement in modern cotton breeding.

Genome-wide Association Analysis of Ten Chilling Tolerance Indices at the Germination and Seedling Stages in Maize

Maize seedlings are very sensitive to chilling, especially during the transition phase from heterotrophic to autotrophic growth. Genetic dissection of the genetic basis of chilling tolerance would provide useful information for genetic improvement of maize inbreds. In this study, genome-wide association analysis was conducted to explore the genetic architecture of maize chilling tolerance at the seed germination and seedling stages with an association panel of 125 inbreds. Ten tolerance indices （ratios of the performance of 10 germination rates and seedling growth-related traits under chilling stress and control conditions） were investigated to assess the ability of chilling tolerance of the inbreds, and a total of 43 single nucleotide polymorphisms associated with chilling tolerance were detected, with none of them being related to chilling tolerance at both the germination and seedling stages simultaneously. Correlation analysis also revealed that the genetic basis of chilling tolerance at the seed germination stage is generally different from that at the seedling stage. In addition, a total of 40 candidate genes involving 31 of the 43 single nucleotide polymorphisms were predicted, and were grouped into five categories according to their functions. The possible roles of these candidate genes in chilling tolerance were also discussed.

Association Analysis for Quality Traits in a Diverse Panel of Chinese Sesame(Sesamum indicum L.)Germplasm

The main objective of this study was to evaluate the potential of a sesame （Sesamum indicum L.） panel for association analysis, and investigate the genetic basis of oil content （OC）, protein content, oleic acid concentration, and linoleic acid concentration using association mapping. A panel of 216 sesame accessions was phenotyped in a multi-environment trial and fingerprinted with 608 polymorphic loci produced by 79 primers, including simple sequence repeats （SSRs）, sequence-related amplified polymorphisms （SRAPs）, and amplified fragment length polymorphisms （AFLPs）. Population structure analysis revealed two subgroups in the population. The Q model performed better for its ability to re-identify associations for the four traits at highly significant P-values compared to the other three mixed models. And a total of 35 and 25 associations for the four traits in 2010 and 2011 were identified, respectively, with the Q model after Bonferroni correction. Among those associations, only one for OC was re-identified in two environments, and several markers associated simultaneously with multiple traits were discovered. These results suggest the power and stability of the Q model for association analysis of nutritional traits in this sesame panel for its slight population stratification and familial relationship, which could aid in dissecting complex traits, and could help to develop strategies for improving nutritional quality.

Association Analysis of the Amino Acid Contents in Rice

The main objective of the present study was to identify simple sequence repeat （SSR） markers associated with the amino acid content of rice （Oryza sativa L.）. SSR markers were selected by prescreening for the relationship to amino acid content. Eighty-four rice landrace accessions from Korea were evaluated for 16 kinds of amino acids in brown rice and ganotyped with 25 SSR markers. Analysis of population structure revealed four subgroups in the population. Linkage disequilibrium （LD） patterns and distributions are of fundamental importance for genome-wide mapping associations. The mean r^2 value for all intrachromosomal loci pairs was 0.033. LD between linked markers decreased with distance. Marker-trait associations were investigated using the unified mixed-model approach, considering both population structure （Q） and kinship （K）. A total of 42 marker-trait associations with amino acids （P 〈 0.05） were identified using 15 different SSR markers covering three chromosomes and explaining more than 40% of the total variation. These results suggest that association analysis in rice is a viable alternative to quantitative trait loci mapping and should help rice breeders develop strategies for improving rice quality.