Construction of a high-density genetic map for sesame based on large scale marker development by specific length amplified fragment (SLAF) sequencing

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Abstract

Background

The genetics and molecular biology of sesame has only recently begun to be studied even though sesame is an important oil seed crop. A high-density genetic map for sesame has not been published yet due to a lack of sufficient molecular markers. Specific length amplified fragment sequencing (SLAF-seq) is a recently developed high-resolution strategy for large-scale de novo SNP discovery and genotyping. SLAF-seq was employed in this study to obtain sufficient markers to construct a high-density genetic map for sesame.

Results

In total, 28.21 Gb of data containing 201,488,285 pair-end reads was obtained after sequencing. The average coverage for each SLAF marker was 23.48-fold in the male parent, 23.38-fold in the female parent, and 14.46-fold average in each F ₂ individual. In total, 71,793 high-quality SLAFs were detected of which 3,673 SLAFs were polymorphic and 1,272 of the polymorphic markers met the requirements for use in the construction of a genetic map. The final map included 1,233 markers on the 15 linkage groups (LGs) and was 1,474.87 cM in length with an average distance of 1.20 cM between adjacent markers. To our knowledge, this map is the densest genetic linkage map to date for sesame. 'SNP_only’ markers accounted for 87.51% of the markers on the map. A total of 205 markers on the map showed significant ( P < 0.05) segregation distortion.

Conclusions

We report here the first high-density genetic map for sesame. The map was constructed using an F ₂ population and the SLAF-seq approach, which allowed the efficient development of a large number of polymorphic markers in a short time. Results of this study will not only provide a platform for gene/QTL fine mapping, map-based gene isolation, and molecular breeding for sesame, but will also serve as a reference for positioning sequence scaffolds on a physical map, to assist in the process of assembling the sesame genome sequence.

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Most cited references 37

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SLAF-seq: An Efficient Method of Large-Scale De Novo SNP Discovery and Genotyping Using High-Throughput Sequencing

Xiaowen Sun, Dongyuan Liu, Xiaofeng Zhang … (2013)

Large-scale genotyping plays an important role in genetic association studies. It has provided new opportunities for gene discovery, especially when combined with high-throughput sequencing technologies. Here, we report an efficient solution for large-scale genotyping. We call it specific-locus amplified fragment sequencing (SLAF-seq). SLAF-seq technology has several distinguishing characteristics: i) deep sequencing to ensure genotyping accuracy; ii) reduced representation strategy to reduce sequencing costs; iii) pre-designed reduced representation scheme to optimize marker efficiency; and iv) double barcode system for large populations. In this study, we tested the efficiency of SLAF-seq on rice and soybean data. Both sets of results showed strong consistency between predicted and practical SLAFs and considerable genotyping accuracy. We also report the highest density genetic map yet created for any organism without a reference genome sequence, common carp in this case, using SLAF-seq data. We detected 50,530 high-quality SLAFs with 13,291 SNPs genotyped in 211 individual carp. The genetic map contained 5,885 markers with 0.68 cM intervals on average. A comparative genomics study between common carp genetic map and zebrafish genome sequence map showed high-quality SLAF-seq genotyping results. SLAF-seq provides a high-resolution strategy for large-scale genotyping and can be generally applicable to various species and populations.

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Characterization of the sesame (Sesamum indicum L.) global transcriptome using Illumina paired-end sequencing and development of EST-SSR markers

Wenliang Wei, Xiaoqiong Qi, Linhai Wang … (2011)

Background Sesame is an important oil crop, but limited transcriptomic and genomic data are currently available. This information is essential to clarify the fatty acid and lignan biosynthesis molecular mechanism. In addition, a shortage of sesame molecular markers limits the efficiency and accuracy of genetic breeding. High-throughput transcriptomic sequencing is essential to generate a large transcriptome sequence dataset for gene discovery and molecular marker development. Results Sesame transcriptomes from five tissues were sequenced using Illumina paired-end sequencing technology. The cleaned raw reads were assembled into a total of 86,222 unigenes with an average length of 629 bp. Of the unigenes, 46,584 (54.03%) had significant similarity with proteins in the NCBI nonredundant protein database and Swiss-Prot database (E-value < 10-5). Of these annotated unigenes, 10,805 and 27,588 unigenes were assigned to gene ontology categories and clusters of orthologous groups, respectively. In total, 22,003 (25.52%) unigenes were mapped onto 119 pathways using the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG). Furthermore, 44,750 unigenes showed homology to 15,460 Arabidopsis genes based on BLASTx analysis against The Arabidopsis Information Resource (TAIR, Version 10) and revealed relatively high gene coverage. In total, 7,702 unigenes were converted into SSR markers (EST-SSR). Dinucleotide SSRs were the dominant repeat motif (67.07%, 5,166), followed by trinucleotide (24.89%, 1,917), tetranucleotide (4.31%, 332), hexanucleotide (2.62%, 202), and pentanucleotide (1.10%, 85) SSRs. AG/CT (46.29%) was the dominant repeat motif, followed by AC/GT (16.07%), AT/AT (10.53%), AAG/CTT (6.23%), and AGG/CCT (3.39%). Fifty EST-SSRs were randomly selected to validate amplification and to determine the degree of polymorphism in the genomic DNA pools. Forty primer pairs successfully amplified DNA fragments and detected significant amounts of polymorphism among 24 sesame accessions. Conclusions This study demonstrates that Illumina paired-end sequencing is a fast and cost-effective approach to gene discovery and molecular marker development in non-model organisms. Our results provide a comprehensive sequence resource for sesame research.

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Construction and application for QTL analysis of a Restriction Site Associated DNA (RAD) linkage map in barley

Yada Chutimanitsakun, Rick Nipper, Alfonso Cuesta-Marcos … (2011)

Background Linkage maps are an integral resource for dissection of complex genetic traits in plant and animal species. Canonical map construction follows a well-established workflow: an initial discovery phase where genetic markers are mined from a small pool of individuals, followed by genotyping of selected mapping populations using sets of marker panels. A newly developed sequence-based marker technology, Restriction site Associated DNA (RAD), enables synchronous single nucleotide polymorphism (SNP) marker discovery and genotyping using massively parallel sequencing. The objective of this research was to assess the utility of RAD markers for linkage map construction, employing barley as a model system. Using the published high density EST-based SNP map in the Oregon Wolfe Barley (OWB) mapping population as a reference, we created a RAD map using a limited set of prior markers to establish linakge group identity, integrated the RAD and prior data, and used both maps for detection of quantitative trait loci (QTL). Results Using the RAD protocol in tandem with the Illumina sequence by synthesis platform, a total of 530 SNP markers were identified from initial scans of the OWB parental inbred lines - the "dominant" and "recessive" marker stocks - and scored in a 93 member doubled haploid (DH) mapping population. RAD sequence data from the structured population was converted into allele genotypes from which a genetic map was constructed. The assembled RAD-only map consists of 445 markers with an average interval length of 5 cM, while an integrated map includes 463 RAD loci and 2383 prior markers. Sequenced RAD markers are distributed across all seven chromosomes, with polymorphic loci emanating from both coding and noncoding regions in the Hordeum genome. Total map lengths are comparable and the order of common markers is identical in both maps. The same large-effect QTL for reproductive fitness traits were detected with both maps and the majority of these QTL were coincident with a dwarfing gene (ZEO) and the VRS1 gene, which determines the two-row and six-row germplasm groups of barley. Conclusions We demonstrate how sequenced RAD markers can be leveraged to produce high quality linkage maps for detection of single gene loci and QTLs. By combining SNP discovery and genotyping into parallel sequencing events, RAD markers should be a useful molecular breeding tool for a range of crop species. Expected improvements in cost and throughput of second and third-generation sequencing technologies will enable more powerful applications of the sequenced RAD marker system, including improvements in de novo genome assembly, development of ultra-high density genetic maps and association mapping.

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Author and article information

Contributors

Yanxin Zhang

Linhai Wang

Huaigen Xin

Donghua Li

Chouxian Ma

Xia Ding

Weiguo Hong

Xiurong Zhang

Journal

Journal ID (nlm-ta): BMC Plant Biol

Journal ID (iso-abbrev): BMC Plant Biol

Title: BMC Plant Biology

Publisher: BioMed Central

ISSN (Electronic): 1471-2229

Publication date Collection: 2013

Publication date (Electronic): 24 September 2013

Volume: 13

Page: 141

Affiliations

[1 ]Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, No.2 Xudong 2nd Rd, 430062 Wuhan, China

[2 ]Biomarker Technologies Corporation, 101300 Beijing, China

Article

Publisher ID: 1471-2229-13-141

DOI: 10.1186/1471-2229-13-141

PMC ID: 3852768

PubMed ID: 24060091

SO-VID: dd189562-d49a-4495-b652-4c5de959ca65

License:

This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Construction of a high-density genetic map for sesame based on large scale marker development by specific length amplified fragment (SLAF) sequencing

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Arabidopsis genomics

Most cited references 37

SLAF-seq: An Efficient Method of Large-Scale De Novo SNP Discovery and Genotyping Using High-Throughput Sequencing

Characterization of the sesame (Sesamum indicum L.) global transcriptome using Illumina paired-end sequencing and development of EST-SSR markers

Construction and application for QTL analysis of a Restriction Site Associated DNA (RAD) linkage map in barley

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