QTL mapping of general combining abilities of four traits in maize using a high-density genetic map

General combining abilities （GCAs） are very important in utilization of heterosis in maize breeding. However, its genetic basis is unclear. In the present study, a set of 118 doubled haploid （DH） lines were induced from F1 generations produced from the cross between the inbred line Zheng 58 and the inbred line W499 belonging to the Reid subgroup. Using the MaizeSNP50 BeadChip, a high-density genetic map was constructed based on the DH population which included 1 147 bin markers with an average interval length of 2.00 cM. Meanwhile, the DH population was crossed with three testers including W16-5, HD568, and W556, which belong to the Sipingtou subgroup. The GCAs of the ear height （EH）, the kernel moisture content （KMC）, the kernel ratio （KR）, and the yield per plant （YPP） were estimated using these hybrids in three environments. Combining the high-density genetic map and the GCAs, a total of 14 QTLs were detected for the GCAs of the four traits. Especially, one pleiotropic QTL was identified on chromosome 1 between the SNP SYN16067 and the SNP PZE-101169244 which was simultaneously associated with the GCAs of the EH, the KR, and the YPP. These QTLs pave the way for further dissecting the genetic architecture underlying GCAs of the traits, and they may be used to enhance GCAs of inbred lines under the fixed heterotic pattern ReidxSipingtou in China through a marker-assisted selection approach.

QTL mapping for berry shape based on a high-density genetic map constructed by whole-genome resequencing in grape

Grape berry shape is an important agricultural trait.Clarifying its genetic basis is significant for cultivating grape varieties that meet market demands.However,the current study by forward genetics has not achieved in-depth results.Here,a high-density map was constructed to identify quantitative trait loci(QTLs)for berry shape.A total of 358709 polymorphic SNPs were obtained using whole-genome resequencing(WGS)based on 208 F2 individuals derived from round grape‘E42-6’and oblong grape‘Rizamat’.The 1635.65 cM high-density map was divided into 19 linkage groups with an average distance of 0.37 cM.Using this map,three significant QTLs for fruit shape index(ShI:ratio of berry length to berry width)identified over three years were mapped onto LG4 and LG5,including one stable QTL on Chr5 with the genomic region of 0.47–1.94 Mb.Combining with gene annotation and expression patterns based on RNA-seq data from two contrasting F2 individuals with round and oblong berry(their average ShI was 1.89 and 1.10,respectively)at four developmental stages,four candidate genes were selected from the above QTLs.They were mainly involved in DNA replication,cell wall modification,and phytohormone biosynthesis.Further analysis of RNA-seq data revealed that several important phytohormone synthesis and metabolic pathways were enriched based on differentially expressed genes(DEGs),which was consistent with the results of QTL mapping for genes related to plant hormone biosynthesis in the F2 population.Furthermore,a comparison of plant hormone content showed that there were significant differences in IAA and tZ content between the two contrasting F2 individuals at different developmental stages.Our findings provide molecular insights into the genetic variation in grape berry shape.Stable QTLs and their tightly linked markers offer the possibility of marker-assisted selection to accelerate berry shape breeding.

High-density genetic mapping identified a major locus for environmental sex expression in pumpkin(Cucurbita moschata Duch.)

Long-day length and high temperature inhibit sex expression in pumpkin(Cucurbita moschata Duch.),and therefore directly impact the production potential.In this study,female flowering patterns in photoperiod-insensitive(PPIS)and photoperiod-sensitive(PPS)germplasms differed significantly in a moderately long day and high temperature environment.However,both germplasms exhibited a similar response in short day with either low temperature or high temperature environment.Photoperiod sensitivity led to this difference in sex expression between the germplasms.For the traits of 1st female flowering node(FFFN)and number of female flowers(NFF),high-density linkage map construction and quantitative trait locus(QTL)mapping were performed using SLAF-seq technology and 162 F_(2) individuals generated from PPIS and PPS.In total,4655 SLAFs were selected and mapped on 20 linkage groups(LGs).The total map length was 2502.01 cM with an average interval distance of 0.75 cM.Major QTLs for both FFFN and NFF were detected on LG6 with intervals of 7.89 and 17.67 cM and PVE values of 30.5%and 22.9%,respectively.Further analyses of the major locus for FFFN revealed 73 protein-coding genes.Among them,4 were related to sex expression,photoperiod flowering,and hormone response.An InDel(insertion and deletion)marker partially correlated with FFFN of the F_(2) population was also developed.Our study identified the QTL for the sex expression response to environmental factors using the high-density linkage map.The identified candidate genes and markers will provide useful information about the molecular interaction between the environment and sex expression and for marker-assisted selection of pumpkin environment-insensitive resources.

Construction of the first high-density genetic map for growth related QTL analysis in Ancherythroculter nigrocauda

Ancherythroculter nigrocauda is a fish endemic to the upper areas of the Changjiang(Yangtze)River in China.Quantitative trait locus(QTL)mapping is a powerful tool to identify potential genes affecting traits of economic importance in domestic animals.In this study,a high-density genetic map was constructed with 5901 single nucleotide polymorphism(SNP)makers by sequencing 92 individual fish from a F1 family using the specific-locus amplified fragment sequencing approach.Initially,48 QTLs for total length,body length,body height,and body weight were identified according to the high density of the genetic map with 24 LGs,a total length of 3839.4 cM,and marker spacing of about 0.82 cM.These QTLs explained 27.1%-49.9%of phenotypic variance.The results of this study suggest that major QTLs are responsible for the growth of A.nigrocauda,and these are potentially useful in comparative genomics research,genome assembly,and marker-assisted breeding programs for this species.

QTL mapping for grain number per spikelet in wheat using a high-density genetic map

Grain number per spikelet (GNS) is a key determinant of grain yield in wheat.A recombinant inbred line population comprising 300 lines was developed from a cross between a high GNS variety H461 and Chinese Spring from which the reference genome assembly of bread wheat was obtained.Both parents and the recombinant inbred lines were genotyped using the wheat 55K single nucleotide polymorphism(SNP) array.A high-density genetic map containing 21,197 SNPs was obtained.These markers covered each of the 21 chromosomes with a total linkage distance of 3792.71 c M.Locations of these markers in this linkage map were highly consistent with their physical locations in the genome assembly of Chinese Spring.The two parents and the whole RIL population were assessed for GNS in two consecutive years at two different locations.Based on multi-environment phenotype data and best liner unbiased prediction values,three quantitative trait loci (QTL) for GNS were identified.One of them located on chromosomes 2B and the other two on 2D.Phenotypic variation explained by these loci varied from 3.07%to26.57%.One of these QTL,QGns.sicau-2D-2,was identified in each of all trials conducted.Based on the best linear unbiased prediction values,this locus explained 19.59%–26.57%of phenotypic variation.A KASP(Kompetitive Allele-Specific PCR) marker closely linked with this locus was generated and used to validate the effects of this locus in three different genetic backgrounds.The identified QTL and the KASP marker developed for it will be highly valuable in fine-mapping the locus and in exploiting it for markerassisted selection in wheat breeding programs.

High-density SNP genetic linkage map construction and quantitative trait locus mapping for resistance to cucumber mosaic virus in tobacco(Nicotiana tabacum L.)

Genetic linkage maps are essential for studies of genetics, genomic structure, and genomic evolution, and for mapping quantitative trait loci (QTL). Identification of molecular markers and construction of genetic linkage maps in tobacco (Nicotiana tabacum L.), a classical model plant and important economic crop, have remained limited. In the present study we identified a large number of single nucleotide polymorphism (SNP) markers and constructed a high-density SNP genetic map for tobacco using restriction site-associated DNA sequencing. In 1216.30 Gb of clean sequence obtained using the Illumina HiSeq 2000 sequencing platform, 99,647,735 SNPs were identified that differed between 203 sequenced plant genomes and the tobacco reference genome. Finally, 13,273 SNP markers were mapped on 24 high-density tobacco genetic linkage groups. The entire linkage map spanned 3421.80 cM, with a mean distance of 0.26 cM between adjacent markers. Compared with genetic linkage maps published previously, this version represents a considerable improvement in the number and density of markers. Seven QTL for resistance to cucumber mosaic virus (CMV) in tobacco were mapped to groups 5 and 8. This high-density genetic map is a promising tool for elucidation of the genetic bases of QTL and for molecular breeding in tobacco.

Construction of high-density SNP genetic maps and QTL mapping for dwarf-related traits in Litchi chinensis Sonn

Litchi chinensis Sonn is widely cultivated in subtropical regions and has an important economic value.A high-density genetic map is a valuable tool for mapping quantitative trait loci(QTL)and marker-assisted breeding programs.In this study,a single nucleotide polymorphism(SNP)-based high-density linkage map was constructed by a genotyping-by-sequencing(GBS)protocol using an F1 population of 178 progenies between two commercial litchi cultivars,‘Ziniangxi’(dwarf)and‘Feizixiao’(vigorous).The genetic map consisted of 3027 SNP markers with a total length of 1711.97 cM in 15 linkage groups(LGs)and an average marker distance of 0.57 cM.Based on this high-density linkage map and three years of phenotyping,a total of 37 QTLs were detected for eight dwarf-related traits,including length of new branch(LNB),diameter of new branch(DNB),length of common petiole(LCP),diameter of common petiole(DCP),length of internode(LI),length of single leaf(LSL),width of single leaf(WSL),and plant height(PH).These QTLs could explain 8.0 to 14.7%(mean=9.7%)of the phenotypic variation.Among them,several QTL clusters were observed,particularly on LG04 and LG11,which might show enrichment for genes regulating the dwarf-related traits in litchi.There were 126 candidate genes identified within the QTL regions,55 of which are differentially expressed genes by RNA-seq analysis between‘Ziniangxi’and‘Feizixiao’.These DEGs were found to participate in the regulation of cell development,material transportation,signal transduction,and plant morphogenesis,so they might play important roles in regulating plant dwarf-related traits.The high-density genetic map and QTLs identification related to dwarf traits can provide a valuable genetic resource and a basis for marker-assisted selection and genomic studies of litchi.

Construction of a high-density adzuki bean genetic map and evaluation of its utility based on a QTL analysis of seed size

Adzuki bean(Vigna angularis(Willd.)Ohwi&Ohashi)is an annual cultivated leguminous crop commonly grown in Asia and consumed worldwide.However,there has been limited research regarding adzuki bean genetics,which has prevented the efficient application of genes during breeding.In the present study,we constructed a high-density genetic map based on whole genome re-sequencing technology and validated its utility by mining QTLs related to seed size.Moreover,we analyzed the sequences flanking insertions/deletions(In Dels)to develop a set of PCR-based markers useful for characterizing adzuki bean genetics.A total of 2904 markers were mapped to 11 linkage groups(LGs).The total length of the map was 1365.0 cM,with an average distance between markers of 0.47 cM.Among the LGs,the number of markers ranged from 208(LG7)to 397(LG1)and the total distance ranged from 97.4 cM(LG9)to 155.6 cM(LG1).Twelve QTLs related to seed size were identified using the constructed map.The two major QTLs in LG2 and LG9 explained 22.1 and 18.8%of the total phenotypic variation,respectively.Ten minor QTLs in LG4,LG5 and LG6 explained 3.0–10.4%of the total phenotypic variation.A total of 9718 primer pairs were designed based on the sequences flanking In Dels.Among the 200 selected primer pairs,75 revealed polymorphisms in 24 adzuki bean germplasms.The genetic map constructed in this study will be useful for screening genes related to other traits.Furthermore,the QTL analysis of seed size and the novel markers described herein may be relevant for future molecular investigations of adzuki bean and will be useful for exploiting the mechanisms underlying legume seed development.

Construction of high-density genetic linkage map of Pyropia yezoensis(Bangiales,Rhodophyta)and identifi cation of red color trait QTLs in the thalli

Pyropia yezoensis is an important macroalga because of its extensive global distribution and economic importance.Color is an important trait in the thalli of P.yezoensis,it is also an effective marker to identify the hybridization in genetic breeding.In this study,a high-density genetic linkage map was constructed based on high-throughput single nucleotide polymorphism(SNP)markers,and used for analyzing the quantitative trait loci(QTLs)of red color trait in the thalli of P.yezoensis.The conchospore undergoes meiosis to develop into an ordered tetrad,and each cell has a haploid phenotype and can grow into a single individual.Based on this theory,F1 haploid population was used as the mapping population.The map included 531 SNP markers,394.57 cM long on average distance of 0.74 cM.Collinear analysis of the genetic linkage map and the physical map indicated that the coverage between the two maps was 79.42%.Furthermore,QTL mapping identified six QTLs for the chromosomal regions associated with the red color trait of the thalli.The value of phenotypic variance explained(PVE)by an individual QTL ranged from 4.71%-63.11%.And QTL qRed-1-1,with a PVE of 63.11%,was considered the major QTL.Thus,these data may provide a platform for gene and QTL fine mapping,and marker-assisted breeding in P.yezoensis in the future.

Construction of a high-density genetic linkage map and identification of flowering-related QTL in erect milkvetch(Astragalus adsurgens)

Erect milkvetch(Astragalus adsurgens)is a perennial legume forage crop with economic and ecological value in livestock grazing and soil-erosion control in arid and semiarid areas worldwide.Genomic information and molecular tools to support breeding and research in the species are limited.The objectives of this investigation were to map its genome using DNA markers and to identify quantitative trait loci(QTL)in the species.An F1 mapping population of 250 plants was developed from a cross between two parents with differing flowering-related traits.A high-density genetic linkage map containing 4821 markers on eight linkage groups(LGs)with a total genetic length of 1395 cM and a mean interval of 0.29 cM between adjacent markers was constructed with SLAF-seq technology.Comparative genomic analyses revealed the highest genome sequence similarity(8.71%)between erect milkvetch and Medicago truncatula,followed by Glycine max(7.65%),Cicer arietinum(7.53%),and Lupinus angustifolius(5.21%).A total of 64 significant QTL for flowering-related traits on six LGs were detected,accounting for 9.38 to 19.1%of the associated phenotype variation.Five and 48 key candidate genes for floret number and inflorescence length were identified based on the Glycyrrhiza uralensis genome.These candidate genes were involved in ubiquitination/degradation,pollen development,cell division,cytokinin biosynthetic process,and plant flowering.These findings shed light on the regulation of flowering traits in erect milkvetch and provide genomic resources for future molecular breeding of the crop.

Rapid Constructing a Genetic Linkage Map by AFLP Technique and Mapping a New Gene tms5

In this study, we reported the repaid construction of a molecular marker linkage map of rice (Oryza sativa L.). An F-2 population from the cross between Annong S-1 and Nanjing 11 was used to construct a genetic linkage map of rice. Total of 142 newly screened AFLP markers and 30 anchor markers (25 SSR markers and 5 RFLP markers) were mapped on the 12 chromosomes covering 1537.4 cM of rice genome. The average interval between these markers was 9.0 cM. The total work which usually was finished in more than one year was finished within only 3 months by one person. This is the first plant AFLP map developed in China. A new thermosensitive genic male sterile gene in rice, tms5, was Egged and mapped onto chromosome 2 during the development of the linkage map.

Construction of Molecular Genetic Linkage Map Based on an RIL Population of Rice and Detection of QTLs for Tiller Angle

In this study, an RIL （recombinant inbred line） population containing 240 lines was developed by single seed descent method （SSD） based on a parent com- bination of small-grain indica cultivar Kasalath and large-grain japanica cultivar TD70 with significant differences in plant type traits, to construct the molecular genetic linkage map. Totally 838 SSR （Simple Sequence Repeat） markers were used for polymorphism screening between parents, 302 SSR markers with polymorphism were detected, with a frequency of 36.04%; 141 SSR markers with clear amplified bands and uniform distribution in the genome were finally used for genotype analysis of the RIL population. According to the experimental results, the frequency of male and female genotype in this RIL population was respectively 53% and 47%, suggesting good balance in population structure. A molecular genetic linkage map of rice was constructed by 141 markers based on a RIL population of 240 lines, with a total genetic distance of about 1 832.47 cM covering all 12 chromosomes, an average genetic distance between markers of 12.70 cM and a range of genetic distance be- tween markers of 0.43-36.11 cM, which is consistent with basic requirements of quantitative trait locus （QTL） mapping. Except for few markers on chromosomes 1 and 8, the order and location of markers is similar to the published sequences of Nipponbare. QTL analysis for the tiller angle was conducted with this RIL population of 240 lines, and results showed that three QTLs controlling tiller angle were detected on chromosome 8, 9 and 11, which were named qTA8, qTA9 and qTA11, with a contribution rate of 4.10%, 26.08% and 4.35%, respectively. To be specific, qTA9 contained Tiller Angle Controlling （TAC1） gene. The construction of this molecular genetic linkage map laid the foundation for genetic analysis and QTL mapping of various traits in the progeny of indica and japonica.

DENSE DISPARITY MAP ESTIMATION USING GENETIC ALGORITHMS

An approach to addressing the stereo correspondence problem is presented using genetic algorithms (GAs) to obtain a dense disparity map. Different from previous methods, this approach casts the stereo matching as a multi-extrema optimization problem such that finding the fittest solution from a set of potential disparity maps. Among a wide variety of optimization techniques, GAs are proven to be potentially effective methods for the global optimization problems with large search space. With this idea, each disparity map is viewed as an individual and the disparity values are encoded as chromosomes, so each individual has lots of chromosomes in the approach. Then, several matching constraints are formulated into an objective function, and GAs are used to search the global optimal solution for the problem. Furthermore, the coarse-to-fine strategy has been embedded in the approach so as to reduce the matching ambiguity and the time consumption. Finally, experimental results on synthetic and real images show the performance of the work.

A genetic linkage map of hexaploid naked oat constructed with SSR markers

Naked oat is a unique health food crop in China. Using 202 F2 individuals derived from a hybrid between the variety 578 and the landrace Sanfensan, we constructed a genetic linkage map consisting of 22 linkage groups covering 2070.50 c M and including 208 simple sequence repeat(SSR) markers. The minimum distance between adjacent markers was0.01 c M and the average was 9.95 c M. Each linkage group contained 2–22 markers. The largest linkage group covered 174.40 c M and the shortest one covered 36.80 c M, with an average of 94.11 c M. Thirty-six markers(17.3%) showing distorted segregation were distributed across linkage groups LG5 to LG22. This map complements published oat genetic maps and is applicable for quantitative trait locus analysis, gene cloning and molecular marker-assisted selection.

Genetic Analysis and Molecular Mapping of a Novel Chlorophyll-Deficit Mutant Gene in Rice

A rice etiolation mutant 824ys featured with chlorophyll deficiency was identified from a normal green rice variety 824B. It showed whole green-yellow plant from the seedling stage, reduced number of tillers and longer growth duration. The contents of chlorophyll, chlorophyll a, chlorophyll b and net photosynthetic rate in leaves of the mutant obviously decreased, as well as the number of spikelets per panicle, seed setting rate and 1000-grain weight compared with its wild-type parent. Genetic analyses on F1 and F2 generations of 824ys crossed with three normal green varieties showed that the chlorophyll-deficit mutant character was controlled by a pair of recessive nuclear gene. Genetic mapping of the mutant gene was conducted by using microsatellite markers and F2 mapping population of 495R/824ys, and the mutant gene of 824ys was mapped on the short arm of rice chromosome 3. The genetic distances from the target gene to the markers RM218, RM282 and RM6959 were 25.6 cM, 5.2 cM and 21.8 cM, respectively. It was considered to be a new chlorophyll-deficit mutant gene and tentatively named as chill（t）.

Construction of Genetic Linkage Map Based on SSR Markers in Peanut(Arachis hypogaea L.)

Molecular genetic maps of crop species can be used in a variety of ways in breeding and genomic research such as identification and mapping of genes and quantitative trait loci （QTLs） for morphological, physiological and economic traits of crop species. However, a comprehensive genetic linkage map for cultivated peanut has not yet been developed due to the extremely low frequency of DNA polymorphism in cultivated peanut. In this study, 142 recombinant inbred lines （RILs） derived from a cross between Yueyou 13 and Zhenzhuhei were used as mapping population in peanut （Arachis hypogaea L.）. A total 652 pairs of genomic-SSR primer and 392 pairs of EST-SSR primer were used to detect the polymorphisms between the two parents. 141 SSR primer pairs, 127 genomic-SSR and 14 EST-SSR ones, which can be used to detect polymorphisms between the two parents, were selected to analyze the RILs population. Thus, a linkage genetic map which consists of 131 SSR loci in 20 linkage groups, with a coverage of 679 cM and an average of 6.12 cM of inter-maker distance was constructed. The putative functions of 12 EST-SSR markers located on the map were analyzed. Eleven showed homology to gene sequences deposited in GenBank. This is the first report of construction of a comprehensive genetic map with SSR markers in peanut （Arachis hypogaea L.）. The map presented here will provide a genetic framework for mapping the qualitative and quantitative trait in peanut.

Genetic Analysis and Primary Mapping of pms4, a Photoperiod-Sensitive Genic Male Sterility Gene in Rice (Oryza sativa)

To understand the genetic characteristics of a new photoperiod-sensitive genic male sterile line Mian 9S, some reciprocal crosses were made between Mian 9S and six indica rice materials, Yangdao 6, Luhui 602, Shuihui 527, Mianhui 725, Fuhui 838 and Yixiang 1B. Genetic analysis results suggested that the photoperiod-sensitive genic male sterility （PGMS） of Mian 9S was controlled by a single recessive nuclear gene. Thus, the F2 population derived from the cross of Yangdao 6/Mian 9S was used to map the PGMS gene in Mian 9S. By using SSR markers, the PGMS gene of Mian 9S was mapped on one side of the markers, RM6659 and RM1305, on rice chromosome 4, with the genetic distances of 3.0 cM and 3.5 cM, respectively. The gene was a novel PGMS gene and designated tentatively as pms4. In addition, the application of the pms4 gene was discussed.

Morphological Structure and Genetic Mapping of New Leaf-Color Mutant Gene in Rice (Oryza sativa)

Leaf-color mutations are a widely-observed class of mutations, playing an important role in the study of chlorophyll biosynthesis and plant chloroplast structure, function, genetics and development. A naturally-occurring leaf-color rice mutant, Baihuaidao 7, was analyzed. Mutant plants typically exhibited a green-white-green leaf-color progression, but this phenotype was only expressed in the presence of a stress signal induced by mechanical scarification such as transplantation. Prior to the appearance of white ~eaves, mutant plant growth, leaf color, chlorophyll content, and chloroplast ultrastructure appeared to be identical to those of the wild type. After the changeover to white leaf color, an examination of the mutated leaves revealed a decrease in total chlorophyll, chlorophyll a, chlorophyll b, and carotenoid content, a reduction in the number of chloroplast grana lamella and grana, and a gradual degradation of the thylakoid lamellas. At maturity, the mutant plant was etiolated and dwarfed compared with wild-type plants. Genetic analysis indicated that the leaf mutant character is controlled by a recessive nuclear gene. Genetic mapping of the mutant gene was performed using an F2 population derived from a Baihuaidao 7 ~ Jiangxi 1587 cross. The mutant gene was mapped to rice chromosome 11, positioned between InDel markers L59.2-7 and L64.8-11, which are separated by approximately 740.5 kb. The mutant gene is believed to be a new leaf-color mutant gene in rice, and is tentatively designated as gwgl.

Construction of a Genetic Linkage Map of Mango Based on SRAP, AFLP and ISSR Markers

A hybrid Ft population was obtained by crossing between mango cultivars ＇ Jinhuang＇ and ＇ Guifei＇. Among the hybrid population, 98 F1 plants were selected as the mapping population. The molecular genetic map of mango was constructed by linkage analysis by SRAP, AFLP and ISSR markers using Joinmap4. 0 software. The genetic linkage map consisted of 33 linkage groups, with a total genetic distance of 1 561.1 cM. Moreover, the genetic linkage map involved 245 polymorphic markers, including 149 SRAP markers, 90 AFLP markers and 6 ISSR markers, with an average genetic distance of 6.37 cM. This study laid a founda- tion for further investigation of the important agronomic traits of mango.

Genetic Analysis and Mapping of TWH Gene in Rice Twisted Hull Mutant

A mutant with twisted hulls was found in a breeding population of rice （Oryza sativa L.）. The mutant shows less grain weight and inferior grain quality in addition to twisted hulls. Genetic analysis indicated that the phenotype of mutant was controlled by a single recessive gene （temporarily designated as TW（H）. To map the TWH gene, an F2 population was generated by crossing the twh mutant to R725, an indica rice variety with normal hulls. For bulked segregant analysis, the bulk of mutant plants was prepared by mixing equal amount of plant tissue from 10 twisted-hull plants and the bulk of normal plants was obtained by pooling equal amount tissue of 10 normal-hull plants. Two hundred and seven pairs of simple sequence repeat （SSR） primers, which are distributed on 12 rice chromosomes, were used for polymorphism analysis of the parents and the two bulks. The TWH locus was initially mapped close to the SSR marker RM526 on chromosome 2. Therefore, further mapping was performed using 50 pairs of SSR primers around the marker RM526. The TWH was delimited between the SSR markers RM14128 and RM208 on the long arm of chromosome 2 at the genetic distances of 1.4 cM and 2.7 cM, respectively. These results provide the foundation for further fine mapping, cloning and functional analysis of the TWH gene.