Ultra-high Density Linkage Map Research Articles

A genome-wide linkage map and QTL mapping for growth traits of Asian rice-field eel (Monopterus albus)

The rice-field eel (Monopterus albus) is showing increasingly important economic value as a freshwater aquaculture fish in Asia. It is necessary to strengthen the genetic breeding research to ensure the quantity and quality of fry supply for the expanding production of rice-field eel. However, the genetic breeding research of this species is still in its infancy. In this study, a genome-wide linkage analysis was carried out to locate its growth-related genes, which may provide guidance for high-yield breeding. In addition, the linkage map was used to mount the supercontigs of the draft genome of rice-field eel. A couple of adults and their 199 six-month-old offspring were used to construct the first ultra-high density linkage map by genome-wide resequencing. The consensus map with a total length of 5161 cM includes 1,603,560 SNPs distributed in 11,522 bins, covering almost all recombination events in the population. Using this map, up to 97.91% of the supercontigs were mounted to chromosomes, which show a good collinearity with the draft genome of rice-field eel. Based on the growth-related traits of the offspring, 37 quantitative trait loci (QTLs) were identified in 18 regions of 4 linkage groups. These QTLs contained SNPs belonging to 58 candidate growth-related genes, among which anks1a, brpf3, plch2, creld1, fry, slc9a1, rragc, mst1r, pfkfb2 and dgkb are worthy of attentions.

An ultra-high density SNP-based linkage map for enhancing the pikeperch (Sander lucioperca) genome assembly to chromosome-scale

Pikeperch (Sander lucioperca) is a fish species with growing economic significance in the aquaculture industry. However, successful positioning of pikeperch in large-scale aquaculture requires advances in our understanding of its genome organization. In this study, an ultra-high density linkage map for pikeperch comprising 24 linkage groups and 1,023,625 single nucleotide polymorphisms markers was constructed after genotyping whole-genome sequencing data from 11 broodstock and 363 progeny, belonging to 6 full-sib families. The sex-specific linkage maps spanned a total of 2985.16 cM in females and 2540.47 cM in males with an average inter-marker distance of 0.0030 and 0.0026 cM, respectively. The sex-averaged map spanned a total of 2725.53 cM with an average inter-marker distance of 0.0028 cM. Furthermore, the sex-averaged map was used for improving the contiguity and accuracy of the current pikeperch genome assembly. Based on 723,360 markers, 706 contigs were anchored and oriented into 24 pseudomolecules, covering a total of 896.48 Mb and accounting for 99.47% of the assembled genome size. The overall contiguity of the assembly improved with a scaffold N50 length of 41.06 Mb. Finally, an updated annotation of protein-coding genes and repetitive elements of the enhanced genome assembly is provided at NCBI.

Mapping quantitative trait loci for yield-related traits and predicting candidate genes for grain weight in maize

Quantitative trait loci (QTLs) mapped in different genetic populations are of great significance for marker-assisted breeding. In this study, an F2:3 population were developed from the crossing of two maize inbred lines SG-5 and SG-7 and applied to QTL mapping for seven yield-related traits. The seven traits included 100-kernel weight, ear length, ear diameter, cob diameter, kernel row number, ear weight, and grain weight per plant. Based on an ultra-high density linkage map, a total of thirty-three QTLs were detected for the seven studied traits with composite interval mapping (CIM) method, and fifty-four QTLs were indentified with genome-wide composite interval mapping (GCIM) methods. For these QTLs, Fourteen were both detected by CIM and GCIM methods. Besides, eight of the thirty QTLs detected by CIM were identical to those previously mapped using a F2 population (generating from the same cross as the mapping population in this study), and fifteen were identical to the reported QTLs in other recent studies. For the fifty-four QTLs detected by GCIM, five of them were consistent with the QTLs mapped in the F2 population of SG-5 × SG-7, and twenty one had been reported in other recent studies. The stable QTLs associated with grain weight were located on maize chromosomes 2, 5, 7, and 9. In addition, differentially expressed genes (DEGs) between SG-5 and SG-7 were obtained from the transcriptomic profiling of grain at different developmental stages and overlaid onto the stable QTLs intervals to predict candidate genes for grain weight in maize. In the physical intervals of confirmed QTLs qKW-7, qEW-9, qEW-10, qGWP-6, qGWP-8, qGWP-10, qGWP-11 and qGWP-12, there were 213 DEGs in total. Finally, eight genes were predicted as candidate genes for grain size/weight. In summary, the stable QTLs would be reliable and the candidate genes predicted would be benefit for maker assisted breeding or cloning.

An ultra-high density map allowed for mapping QTL and candidate genes controlling dry latex yield in rubber tree

Rubber tree (Hevea brasiliensis) is the most important commercial producer of high-quality natural rubber in the world. Here, we identified 571,267 SNPs and 134,184 indels, and constructed the first ultra-high density genetic linkage map in rubber tree population. This map consisted of 6940 markers, with average marker density and recombination rate 0.30 cM and 0.97 cM/Mb, respectively. Then the whole genome QTL scanning for dry latex yield (DLY) trait were performed and seventeen repQTLs were obtained, among which qDFY-10 and qDFY-18-4 could explain up to 38.3% and 33.3% phenotypic variability, respectively. Numerous highly-promising QTL candidate genes were identified and then verified significant associated with DLY trait, including some first found in rubber tree, such as thioredoxin h, plastin-like protein, calmodulin binding protein, cytochrome-c oxidase and methylglutaconyl-CoA hydratase. This ultra-high density linkage map supplies a base for important traits QTL mapping and will be useful in the improvement of the assembly of genome sequences in rubber tree. The verification of candidate QTL genes showed great importance for understanding the possible function of mapped QTLs and the comprehensive relations between phenotypes and genotypes. Accurate QTL mapping will be enhance genetic studies that can be applied in rubber tree breeding programs.

Ultrahigh-Density Linkage Map Construction Using Low-Coverage Whole-Genome Sequencing of a Doubled Haploid Population: Case Study of Torafugu (Takifugu rubripes).

Next-generation sequencing enables genome-wide genotyping of a large population and further facilitates the construction of a genetic linkage map. Low-coverage whole-genome sequencing has been employed for genetic linkage map construction in several species. However, this strategy generally requires available high-quality reference genomes and/or designed inbred pedigree lines, which restrict the scope of application for non-model and unsequenced species. Here, using torafugu (Takifugu rubripes) as a test model, we propose a new strategy for ultrahigh-density genetic linkage map construction using low-coverage whole-genome sequencing of a haploid/doubled haploid (H/DH) population without above requirements. Low-coverage (≈1×) whole-genome sequencing data of 165 DH individuals were used for de novo assembly and further performed single nucleotide polymorphisms (SNPs) calling, resulting in the identification of 1,070,601 SNPs. Based on SNP genotypes and de novo assembly, genotypes were associated with short DNA segments and an ultrahigh-density linkage map was constructed containing information of 802,277 SNPs in 3090 unique positions. Comparative analyses showed near-perfect concordance between the present linkage map and the latest published torafugu genome (FUGU5). This strategy would facilitate ultrahigh-density linkage map construction in various sexually reproducing organisms for which H/DH populations can be generated.

Developing genome-wide SNPs and constructing an ultrahigh-density linkage map in oil palm

Oil palm (Elaeis guineensis Jacq.) is the leading oil-producing crops and the most important edible oil resource worldwide. DNA markers and genetic linkage maps are essential resources for marker-assisted selection to accelerate genetic improvement. We conducted RAD-seq on an Illumina NextSeq500 to discover genome-wide SNPs, and used the SNPs to construct a linkage map for an oil palm (Tenera) population derived from a cross between a Deli Dura and an AVROS Pisifera. The RAD-seq produced 1,076 million single-end reads across the breeding population containing 155 trees. Mining this dataset detected 510,251 loci. After filtering out loci with low accuracy and more than 20% missing data, 11,394 SNPs were retained. Using these SNPs, in combination with 188 anchor SNPs and 123 microsatellites, we constructed a linkage map containing 10,023 markers covering 16 chromosomes. The map length is 2,938.2 cM with an average marker space of 0.29 cM. The large number of SNPs will supply ample choices of DNA markers in analysing the genetic diversity, population structure and evolution of oil palm. This high-density linkage map will contribute to mapping quantitative trait loci (QTL) for important traits, thus accelerating oil palm genetic improvement.

Construction of an ultrahigh-density genetic linkage map for Jatropha curcas L. and identification of QTL for fruit yield

BackgroundAs an important biofuel plant, the demand for higher yield Jatropha curcas L. is rapidly increasing. However, genetic analysis of Jatropha and molecular breeding for higher yield have been hampered by the limited number of molecular markers available.ResultsAn ultrahigh-density linkage map for a Jatropha mapping population of 153 individuals was constructed and covered 1380.58 cM of the Jatropha genome, with average marker density of 0.403 cM. The genetic linkage map consisted of 3422 SNP and indel markers, which clustered into 11 linkage groups. With this map, 13 repeatable QTLs (reQTLs) for fruit yield traits were identified. Ten reQTLs, qNF-1, qNF-2a, qNF-2b, qNF-2c, qNF-3, qNF-4, qNF-6, qNF-7a, qNF-7b and qNF-8, that control the number of fruits (NF) mapped to LGs 1, 2, 3, 4, 6, 7 and 8, whereas three reQTLs, qTWF-1, qTWF-2 and qTWF-3, that control the total weight of fruits (TWF) mapped to LGs 1, 2 and 3, respectively. It is interesting that there are two candidate critical genes, which may regulate Jatropha fruit yield. We also identified three pleiotropic reQTL pairs associated with both the NF and TWF traits.ConclusionThis study is the first to report an ultrahigh-density Jatropha genetic linkage map construction, and the markers used in this study showed great potential for QTL mapping. Thirteen fruit-yield reQTLs and two important candidate genes were identified based on this linkage map. This genetic linkage map will be a useful tool for the localization of other economically important QTLs and candidate genes for Jatropha.

Erratum: An ultra-high density linkage map and QTL mapping for sex and growth-related traits of common carp (Cyprinus carpio).

Scientific Reports 6: Article number: 26693; published online: 26 May 2016; updated: 22 July 2016 In the original version of this Article, Affiliation 3 was omitted for Wenzhu Peng. The correct affiliations are listed below: College of Life Sciences, Shanghai Ocean University, Shanghai, 201306, China.

An ultra-high density linkage map and QTL mapping for sex and growth-related traits of common carp (Cyprinus carpio).

High density genetic linkage maps are essential for QTL fine mapping, comparative genomics and high quality genome sequence assembly. In this study, we constructed a high-density and high-resolution genetic linkage map with 28,194 SNP markers on 14,146 distinct loci for common carp based on high-throughput genotyping with the carp 250 K single nucleotide polymorphism (SNP) array in a mapping family. The genetic length of the consensus map was 10,595.94 cM with an average locus interval of 0.75 cM and an average marker interval of 0.38 cM. Comparative genomic analysis revealed high level of conserved syntenies between common carp and the closely related model species zebrafish and medaka. The genome scaffolds were anchored to the high-density linkage map, spanning 1,357 Mb of common carp reference genome. QTL mapping and association analysis identified 22 QTLs for growth-related traits and 7 QTLs for sex dimorphism. Candidate genes underlying growth-related traits were identified, including important regulators such as KISS2, IGF1, SMTLB, NPFFR1 and CPE. Candidate genes associated with sex dimorphism were also identified including 3KSR and DMRT2b. The high-density and high-resolution genetic linkage map provides an important tool for QTL fine mapping and positional cloning of economically important traits, and improving common carp genome assembly.

Ultrahigh-density linkage map for cultivated cucumber (Cucumis sativus L.) using a single-nucleotide polymorphism genotyping array.

Genotyping arrays are tools for high-throughput genotyping, which is beneficial in constructing saturated genetic maps and therefore high-resolution mapping of complex traits. Since the report of the first cucumber genome draft, genetic maps have been constructed mainly based on simple-sequence repeats (SSRs) or on combinations of SSRs and sequence-related amplified polymorphism (SRAP). In this study, we developed the first cucumber genotyping array consisting of 32,864 single-nucleotide polymorphisms (SNPs). These markers cover the cucumber genome with a median interval of ~2 Kb and have expected genotype calls in parents/F1 hybridizations as a training set. The training set was validated with Fluidigm technology and showed 96% concordance with the genotype calls in the parents/F1 hybridizations. Application of the genotyping array was illustrated by constructing a 598.7 cM genetic map based on a ‘9930’ × ‘Gy14’ recombinant inbred line (RIL) population comprised of 11,156 SNPs. Marker collinearity between the genetic map and reference genomes of the two parents was estimated at R2 = 0.97. We also used the array-derived genetic map to investigate chromosomal rearrangements, regional recombination rate, and specific regions with segregation distortions. Finally, 82% of the linkage-map bins were polymorphic in other cucumber variants, suggesting that the array can be applied for genotyping in other lines. The genotyping array presented here, together with the genotype calls of the parents/F1 hybridizations as a training set, should be a powerful tool in future studies with high-throughput cucumber genotyping. An ultrahigh-density linkage map constructed by this genotyping array on RIL population may be invaluable for assembly improvement, and for mapping important cucumber QTLs.

Parent-independent genotyping for constructing an ultrahigh-density linkage map based on population sequencing

Bar-coded multiplexed sequencing approaches based on new-generation sequencing technologies provide capacity to sequence a mapping population in a single sequencing run. However, such approaches usually generate low-coverage and error-prone sequences for each line in a population. Thus, it is a significant challenge to genotype individual lines in a population for linkage map construction based on low-coverage sequences without the availability of high-quality genotype data of the parental lines. In this paper, we report a method for constructing ultrahigh-density linkage maps composed of high-quality single-nucleotide polymorphisms (SNPs) based on low-coverage sequences of recombinant inbred lines. First, all potential SNPs were identified to obtain drafts of parental genotypes using a maximum parsimonious inference of recombination, making maximum use of SNP information found in the entire population. Second, high-quality SNPs were identified by filtering out low-quality ones by permutations involving resampling of windows of SNPs followed by Bayesian inference. Third, lines in the mapping population were genotyped using the high-quality SNPs assisted by a hidden Markov model. With 0.05x genome sequence per line, an ultrahigh-density linkage map composed of bins of high-quality SNPs using 238 recombinant inbred lines derived from a cross between two rice varieties was constructed. Using this map, a quantitative trait locus for grain width (GW5) was localized to its presumed genomic region in a bin of 200 kb, confirming the accuracy and quality of the map. This method is generally applicable in genetic map construction with low-coverage sequence data.