Construction of a high-density genetic linkage map and QTL mapping of growth and cold tolerance traits in Takifugu fasciatus.

Takifugu fasciatus is an aquaculture species with high economic value. In recent years, problems such as environmental pollution and inbreeding have caused a serious decline in T. fasciatus germplasm resources. In this study, a high-density genetic linkage map was constructed by whole-genome resequencing. The map consists of 4891 bin markers distributed across 22 linkage groups (LGs), with a total genetic coverage of 2381.353 cM and a mean density of 0.535 cM. Quantitative trait locus (QTL) localization analysis showed that a total of 19 QTLs associated with growth traits of T. fasciatus in the genome-wide significance threshold range, distributed on 11 LGs. In addition, 11 QTLs associated with cold tolerance traits were identified, each scattered on a different LG. Furthermore, we used QTL localization analysis to screen out three candidate genes (IGF1, IGF2, ADGRB) related to growth in T. fasciatus. Meanwhile, we screened three candidate genes (HSP90, HSP70, and HMGB1) related to T. fasciatus cold tolerance. Our study can provide a theoretical basis for the selection and breeding of cold-tolerant or fast-growing T. fasciatus.

Construction of the first high-density genetic linkage map and QTL mapping of flavonoid and leaf-size related traits in Epimedium.

BACKGROUND: Leaves are the main medicinal organ in Epimedium herbs, and leaf flavonoid content is an important criterion of Epimedium herbs. However, the underlying genes that regulate leaf size and flavonoid content are unclear, which limits the use of breeding for Epimedium development. This study focuses on QTL mapping of flavonoid and leaf-size related traits in Epimedium. RESULTS: We constructed the first high-density genetic map (HDGM) using 109 F1 hybrids of Epimedium leptorrhizum and Epimedium sagittatum over three years (2019-2021). Using 5,271 single nucleotide polymorphism (SNP) markers, an HDGM with an overall distance of 2,366.07 cM and a mean gap of 0.612 cM was generated by utilizing genotyping by sequencing (GBS) technology. Every year for three years, 46 stable quantitative trait loci (QTLs) for leaf size and flavonoid contents were discovered, including 31 stable loci for Epimedin C (EC), one stable locus for total flavone content (TFC), 12 stable loci for leaf length (LL), and two stable loci for leaf area (LA). For flavonoid content and leaf size, the phenotypic variance explained for these loci varied between 4.00 and 16.80% and 14.95 and 17.34%, respectively. CONCLUSIONS: Forty-six stable QTLs for leaf size and flavonoid content traits were repeatedly detected over three years. The HDGM and stable QTLs are laying the basis for breeding and gene investigation in Epimedium and will contribute to accelerating the identification of desirable genotypes for Epimedium breeding.

Development of a 135K SNP genotyping array for Actinidia arguta and its applications for genetic mapping and QTL analysis in kiwifruit.

Kiwifruit (Actinidia spp) is a woody, perennial and deciduous vine. In this genus, there are multiple ploidy levels but the main cultivated cultivars are polyploid. Despite the availability of many genomic resources in kiwifruit, SNP genotyping is still a challenge given these different levels of polyploidy. Recent advances in SNP array technologies have offered a high-throughput genotyping platform for genome-wide DNA polymorphisms. In this study, we developed a high-density SNP genotyping array to facilitate genetic studies and breeding applications in kiwifruit. SNP discovery was performed by genome-wide DNA sequencing of 40 kiwifruit genotypes. The identified SNPs were stringently filtered for sequence quality, predicted conversion performance and distribution over the available Actinidia chinensis genome. A total of 134 729 unique SNPs were put on the array. The array was evaluated by genotyping 400 kiwifruit individuals. We performed a multidimensional scaling analysis to assess the diversity of kiwifruit germplasm, showing that the array was effective to distinguish kiwifruit accessions. Using a tetraploid F1 population, we constructed an integrated linkage map covering 3060.9 cM across 29 linkage groups and performed QTL analysis for the sex locus that has been identified on Linkage Group 3 (LG3) in Actinidia arguta. Finally, our dataset presented evidence of tetrasomic inheritance with partial preferential pairing in A. arguta. In conclusion, we developed and evaluated a 135K SNP genotyping array for kiwifruit. It has the advantage of a comprehensive design that can be an effective tool in genetic studies and breeding applications in this high-value crop.

The roles of recombination and selection in shaping genomic divergence in an incipient ecological species complex.

Speciation genomic studies have revealed that genomes of diverging lineages are shaped jointly by the actions of gene flow and selection. These evolutionary forces acting in concert with processes such as recombination and genome features such as gene density shape a mosaic landscape of divergence. We investigated the roles of recombination and gene density in shaping the patterns of differentiation and divergence between the cyclically parthenogenetic ecological sister-taxa, Daphnia pulicaria and Daphnia pulex. First, we assembled a phased chromosome-scale genome assembly using trio-binning for D. pulicaria and constructed a genetic map using an F2-intercross panel to understand sex-specific recombination rate heterogeneity. Finally, we used a ddRADseq data set with broad geographic sampling of D. pulicaria, D. pulex, and their hybrids to understand the patterns of genome-scale divergence and demographic parameters. Our study provides the first sex-specific estimates of recombination rates for a cyclical parthenogen, and unlike other eukaryotic species, we observed male-biased heterochiasmy in D. pulicaria, which may be related to this somewhat unique breeding mode. Additionally, regions of high gene density and recombination are generally more divergent than regions of suppressed recombination. Outlier analysis indicated that divergent genomic regions are probably driven by selection on D. pulicaria, the derived lineage colonizing a novel lake habitat. Together, our study supports a scenario of selection acting on genes related to local adaptation shaping genome-wide patterns of differentiation despite high local recombination rates in this species complex. Finally, we discuss the limitations of our data in light of demographic uncertainty.

Identification of a genomic region linked with sex determination of Undaria pinnatifida (Alariaceae) through genomic resequencing and genetic linkage analyses of a segregating gametophyte family.

Different from the traditional knowledge about kelp, three sexual phenotypes (female, male, and monoecious) exist in the haploid gametophytes of Undaria pinnatifida. However, the sex-determining mechanisms remain unknown. Genetic linkage mapping is an efficient tool to identify sex-linked regions. In the present study, we resequenced a segregating gametophyte family based on the male genome of U. pinnatifida. A high-density genetic linkage map was constructed using 9887 SNPs, with an average distance of 0.41 cM between adjacent SNPs. On the basis of this genetic map and using the composite interval mapping method, we identified 62 SNPs significantly linked with the sexual phenotype. They were located at a position of 67.67 cM on the linkage group 23, corresponding to a physical range of 14.67 Mbp on the HiC_Scaffold_23 of the genome. Reanalysis of the previous specific length amplified fragment sequencing data according to the reference genome led to the identification of a sex-linked genomic region that encompassed the above-mentioned 14.67 Mbp region. Hence, this overlapped genomic range was likely the sex-determining region. Within this region, 129 genes were retrieved and 39 of them were annotated with explicit function, including the potential male sex-determining gene-encoding high mobility group (HMG) domain protein. Relative expression analysis of the HMG gene showed that its expression was higher in male gametophytes during the vegetative phase and monoecious gametophytes during both the vegetative and gametogenesis phases, but significantly lower in male gametophytes during the gametogenesis phase. These results provide a foundation for deciphering the sex-determining mechanism of U. pinnatifida.

Genetic Mapping of a Novel Gene PmAege7M from Aegilops geniculata Conferring Resistance to Wheat Powdery Mildew.

Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is one of the most damaging foliage diseases of wheat across the world. Aegilops geniculata Roth is a valuable gene resource for enhancing wheat resistance to powdery mildew. This study identified Ae. geniculata accession PI 487224 as immune and PI 487228 as susceptible to powdery mildew. Genetic analysis of the F1, F2, and F2:3 progeny derived from PI 487224 × PI 487228 showed that powdery mildew resistance in PI 487224 was controlled by two independent dominant genes located on two different nonhomologous chromosomes. By combing bulked segregant RNA-Seq, genetic linkage analysis of a single resistance gene segregation population, and marker analysis of a set of 14 wheat-Ae. geniculata chromosome addition lines, one of the resistance genes, temperately designated PmAege7M, was mapped to a 4.9-cM interval flanked by markers STS7-55926 and SNP7-45792/STS7-65911 on the long arm of chromosome 7 Mg of PI 487224, spanning 604.73 to 622.82 Mb on the 7D long arm based on the Ae. tauschii reference genome (Aet_v4.0). The map and closely linked markers of PmAege7M from Ae. geniculata in this study will facilitate the transfer of PmAege7M into common wheat and fine mapping of the gene.

QTL mapping and identification of genes associated with the resistance to Acanthoscelides obtectus in cultivated common bean using a high-density genetic linkage map.

BACKGROUND: Common bean (Phaseolus vulgaris L.) is an important agricultural product with large nutritional value, and the insect pest Acanthoscelides obtectus (Say) seriously affects its product quality and commodity quality during storage. Few researches on genes of bruchid resistance have investigated in common bean cultivars. RESULTS: In this study, a bruchid-resistant cultivar black kidney bean and a highly susceptible accession Longyundou3 from different gene banks were crossed to construct a recombinant inbred line population. The genetic analysis indicated a quantitative inheritance of the bruchid resistance trait controlled by polygenes. A high-density genetic map of a total map distance of 1283.68 cM with an average interval of 0.61 cM between each marker was constructed using an F6 population of 157 recombinant inbred lines. The map has 3106 bin markers, containing 2,234,769 SNPs. Using the high-density genetic map, a new quantitative trait locus for the resistance to Acanthoscelides obtectus was identified on chromosome 6. New molecular markers based on the candidate region were developed, and this locus was further delimited to an interval of 122.3 kb between SSR markers I6-4 and I6-16 using an F2 population. This region comprised five genes. Phvul.006G003700, which encodes a bifunctional inhibitor, may be a potential candidate gene for bruchid resistance. Sequencing analysis of candidate gene identified a 5 bp insertion-deletion in promoter of gene Phvul.006G003700 between two parents. Expression analysis of candidate gene revealed that the expression level of Phvul.006G003700 in bruchid-resistant parent was markedly higher than that in bruchid-susceptible parent both in dry seeds and leaves. CONCLUSIONS: A high-density genetic linkage map was constructed utilizing whole-genome resequencing and one new QTL for bruchid resistance was identified on chromosome 6 in common bean cultivar. Phvul.006G003700 (encoding a bifunctional inhibitor) may be a potential candidate gene. These results may form the basis for further research to reveal the bruchid resistance molecular mechanism of common bean.

QTL mapping for growth-related traits by constructing the first genetic linkage map in Simao pine.

BACKGROUND: Simao pine is one of the primary economic tree species for resin and timber production in southwest China. The exploitation and utilization of Simao pine are constrained by the relatively lacking of genetic information. Construction a fine genetic linkage map and detecting quantitative trait locis (QTLs) for growth-related traits is a prerequisite section of Simao Pine's molecular breeding program. RESULTS: In our study, a high-resolution Simao pine genetic map employed specific locus amplified fragment sequencing (SLAF-seq) technology and based on an F1 pseudo-testcross population has been constructed. There were 11,544 SNPs assigned to 12 linkage groups (LGs), and the total length of the map was 2,062.85 cM with a mean distance of 0.37 cM between markers. According to the phenotypic variation analysis for three consecutive years, a total of seventeen QTLs for four traits were detected. Among 17 QTLs, there were six for plant height (Dh.16.1, Dh16.2, Dh17.1, Dh18.1-3), five for basal diameter (Dbd.17.1-5), four for needle length (Dnl17.1-3, Dnl18.1) and two for needle diameter (Dnd17.1 and Dnd18.1) respectively. These QTLs individually explained phenotypic variance from 11.0-16.3%, and the logarithm of odds (LOD) value ranged from 2.52 to 3.87. CONCLUSIONS: In our study, a fine genetic map of Simao pine applied the technology of SLAF-seq has been constructed for the first time. Based on the map, a total of 17 QTLs for four growth-related traits were identified. It provides helpful information for genomic studies and marker-assisted selection (MAS) in Simao pine.

Quantitative trait loci (QTL) analysis of leaf related traits in spinach (Spinacia oleracea L.).

BACKGROUND: Spinach (Spinacia oleracea L.) is an important leafy vegetable crop, and leaf-related traits including leaf length, leaf width, and petiole length, are important commercial traits. However, the underlying genes remain unclear. The objective of the study was to conduct QTL mapping of leaf-related traits in spinach. RESULTS: A BC1 population was used to construct the linkage map and for QTL mapping of leaf length, leaf width, petiole length, and the ratio of leaf length to width in 2015 and 2019. Two genetic linkage maps were constructed by specific locus amplified fragment sequencing (SLAF-seq), and kompetitive allele specific PCR (KASP) technology, respectively using BC1 population in 2015. Based on the results of 2015, the specific linkage groups (LG) detected QTLs were generated using BC1 population in 2019. A total of 13 QTLs were detected for leaf-related traits, only five QTLs being repeatedly detected in multiple years or linkage maps. Interestingly, the major QTLs of leaf length, petiole length, and the ratio of leaf length to width were highly associated with the same SNP markers (KM3102838, KM1360385 and KM2191098). A major QTL of leaf width was mapped on chromosome 1 from 41.470-42.045 Mb. And 44 genes were identified within the region. Based on the GO analysis, these genes were significantly enriched on ribonuclease, lyase activity, phosphodiester bond hydrolysis process, and cell wall component, thus it might change cell size to determine leaves shape. CONCLUSIONS: Five QTLs for leaf-related traits were repeatedly detected at least two years or linkage maps. The major QTLs of leaf length, petiole length, and the ratio of leaf length to width were mapped on the same loci. And three genes (Spo10792, Spo21018, and Spo21019) were identified as important candidate genes for leaf width.

A new genetic locus for self-compatibility in the outcrossing grass species perennial ryegrass (Lolium perenne).

BACKGROUND: Self-incompatibility (SI) is a physiological mechanism that many flowering plants employ to prevent self-fertilization and maintain heterozygosity. In the grass family this is known to be controlled by a two locus (S-Z) system; however, the SI system is intrinsically leaky. Modifier genes of both the S and Z loci and a further locus, T, are known to override SI leading to self-fertilization and self-seed production. This has implications for the ecological and evolutionary success as well as the commercial breeding of grasses. Here we report a study where the genetic control of self-compatibility (SC) was determined from the results of self-pollinating an F2 population of perennial ryegrass from two independently derived inbred lines produced by single-seed descent. METHODS: In vitro self-pollinations of 73 fertile plants were analysed. A genetic association analysis was made with a panel of 1863 single-nucleotide polymorphism (SNP) markers, generated through genotype-by-sequencing methodology. Markers were placed on a recombination map of seven linkage groups (LGs) created using Joinmap v.5. The seed set on self- and open-pollinated inflorescences was determined on 143 plants, including the 73 plants analysed for self-pollination response. KEY RESULTS: Self-pollinations revealed a bimodal distribution of percentage SC with peaks at 50 and 100 %. A single quantitative trait locus (QTL) was identified with peak association for marker 6S14665z17875_11873 that mapped to LG 6. Peak position was associated with maximum marker segregation distortion. The self-compatible plants were equally fecund after self- and open pollination. CONCLUSIONS: This is the first report in the Poaceae family of an SC locus located on LG 6. This new SC QTL discovery, as well as indicating the complex nature of the pollen-stigma recognition process and its evolutionary significance, provides an additional source of SC for breeding perennial ryegrass.

High-Density Genetic Linkage Map Construction and White Rot Resistance Quantitative Trait Loci Mapping for Genus Vitis Based on Restriction Site-Associated DNA Sequencing.

Grape white rot (Coniothyrium diplodiella) is a major fungal disease affecting grape yield and quality. Quantitative trait locus (QTL) analysis is an important method for studying important horticultural traits of grapevine. This study was conducted to construct a high-density map and conduct QTL mapping for grapevine white rot resistance. A mapping population with 177 genotypes was developed from interspecific hybridization of a white rot-resistant cultivar (Vitis vinifera × V. labrusca 'Zhuosexiang') and white rot-susceptible cultivar (V. vinifera 'Victoria'). Single-nucleotide polymorphism (SNP) markers were developed by restriction site-associated DNA sequencing. The female, male, and integrated maps contained 2,501, 4,110, and 6,249 SNP markers with average genetic distances of adjacent markers of 1.25, 0.77, and 0.50 cM, respectively. QTL mapping was conducted based on white rot resistance identification of 177 individuals in July and August of 2017 and 2018. Notably, one stable QTL related to white rot resistance was detected and located on linkage group LG14. The phenotypic variance ranged from 12.93 to 13.43%. An SNP marker (chr14_3929380), which cosegregated with white rot resistance, was discovered and shows potential for use in marker-assisted selection to generate new grapevine cultivars with resistance to white rot.

A high-density genetic linkage map for Chinese perch (Siniperca chuatsi) using 2.3K genotyping-by-sequencing SNPs.

Chinese perch, Siniperca chuatsi (Basilewsky), is one of the most commercially important cultured fishes in China. In the present study, a high-density genetic linkage map of Chinese perch was constructed by genotyping-by-sequencing technique with an F1 mapping panel containing 190 progenies. A total of 2328 SNPs were assigned to 24 linkage groups (LGs), agreeing with the chromosome haploid number in this species (n = 24). The sex-averaged map covered 97.9% of the Chinese perch genome, with the length of 1694.3 cM and a marker density of 0.7 cM/locus. The number of markers per LG ranged from 57 to 222, with a mean of 97. The length of LGs varied from 43.2 to 108.2 cM, with a mean size of 70.6 cM. The recombination rate of females was 1.5:1, which was higher than that of males. To better understand the distribution pattern of segregation distortion between the two sexes of Chinese perch, the skewed markers were retained and used to reconstruct the sex-specific maps. The 16 segregation distortion regions were identified on 10 LGs of the female map, while 12 segregation distortion regions on eight LGs of the male map. Among these LGs, six LGs matched between the sex-specific maps. This high-density linkage map could provide a solid basis for identifying QTL associated with economically important traits, and for implementing marker-assisted selection breeding of Chinese perch.

Development of an SSR marker-based genetic linkage map and identification of a QTL associated with flowering time in Eustoma.

Lisianthus (Eustoma grandiflorum) is an important floricultural crop cultivated worldwide. Despite its commercial importance, few DNA markers are available for molecular genetic research. In this study, we constructed a genetic linkage map and to detect quantitative trait loci (QTLs) for important agronomic traits of lisianthus. To develop simple sequence repeat (SSR) markers, we used 454-pyrosequencing technology to obtain genomic shotgun sequences and subsequently identified 8263 putative SSRs. A total of 3990 primer pairs were designed in silico and 1189 unique primer pairs were extracted through a BLAST search. Amplification was successful for more than 1000 primer pairs, and ultimately 278 SSR markers exhibited polymorphism between the two lisianthus accessions evaluated. Based on these markers, a genetic linkage map was constructed using a breeding population derived from crosses between the two accessions, for which flowering time differed (>140 days when grown under 20°C). We detected one QTL associated with flowering time (phenotypic variance, 27%; LOD value, 3.7). The SSR marker located at this QTL may account for variation in flowering time among accessions (i.e., three accessions whose nodes of the first flower were over 30 had late-flowering alleles of this QTL).

Construction of a high-density genetic map with whole genome sequencing in Nicotiana tabacum L.

Tobacco (Nicotiana tabacum L.) is an essential commercial crop and an ideal model plant for biological mechanism studies. As an allopolyploid species, tobacco harbors a massive and complex genome, which makes the application of molecular markers complicated and challenging. In our study, we performed whole-genome sequencing of an intraspecific recombinant inbred line (RIL) population, a F1 generation and their parents. With the Nicotiana tabacum (K326 cultivar) genome as reference, a total of 45,081 markers were characterized to construct the genetic map, which spanned a genetic distance of 3486.78 cM. Evaluation of a two-dimensional heat map proved the high quality of the genetic map. We utilized these markers to anchor scaffolds and analyzed the ancestral genome origin of linkage groups (LGs). Furthermore, such a high-density genetic map will be applied for quantitative trait locus (QTL) detection, gene localization, genome-wide association studies (GWAS), and marker-assisted breeding in tobacco.

LegumeSSRdb: A Comprehensive Microsatellite Marker Database of Legumes for Germplasm Characterization and Crop Improvement.

Microsatellites, or simple sequence repeats (SSRs), are polymorphic loci that play a major role as molecular markers for genome analysis and plant breeding. The legume SSR database is a webserver which contains simple sequence repeats (SSRs) from genomes of 13 legume species. A total of 3,706,276 SSRs are present in the database, 698,509 of which are genic SSRs, and 3,007,772 are non-genic. This webserver is an integrated tool to perform end-to-end marker selection right from generating SSRs to designing and validating primers, visualizing the results and blasting the genomic sequences at one place without juggling between several resources. The user-friendly web interface allows users to browse SSRs based on the genomic region, chromosome, motif type, repeat motif sequence, frequency of motif, and advanced searches allow users to search based on chromosome location range and length of SSR. Users can give their desired flanking region around repeat and obtain the sequence, they can explore the genes in which the SSRs are present or the genes between which the SSRs are bound design custom primers, and perform in silico validation using PCR. An SSR prediction pipeline is implemented where the user can submit their genomic sequence to generate SSRs. This webserver will be frequently updated with more species, in time. We believe that legumeSSRdb would be a useful resource for marker-assisted selection and mapping quantitative trait loci (QTLs) to practice genomic selection and improve crop health. The database can be freely accessed at http://bioinfo.usu.edu/legumeSSRdb/.

Construction of a high-density genetic linkage map and QTL analysis of morphological traits in an F1 Malusdomestica × Malus baccata hybrid.

Apple is considered the most commonly grown fruit crop in temperate regions that brings great economic profits to fruit growers. Dwarfing rootstocks have been extensively used in apple breeding as well as commercial orchards, but the molecular and genetic basis of scion dwarfing and other morphological traits induced by them is still unclear. At present, we report a genetic map of Malusdomestica × Malus baccata with high density. The F1 population was sequenced by a specific length amplified fragment (SLAF). In the genetic map, 5064 SLAF markers spanning 17 linkage groups (LG) were included. Dwarf-related and other phenotypic traits of the scion were evaluated over a 3-year growth period. Based on quantitative trait loci (QTL) evaluation of plant height and trunk diameter, two QTL clusters were found on LG 11, which exhibited remarkable influences on dwarfing of the scion. In this analysis, QTL DW2, which was previously reported as a locus that controls dwarfing, was confirmed. Moreover, three novel QTLs for total flower number and branching flower number were detected on LG2 and LG4, exhibited the phenotypic variation that has been explained by QTL ranging from 8.80% to 34.80%. The findings of the present study are helpful to find scion dwarfing and other phenotypes induced by rootstock in the apple. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-021-01069-0.

Novel stable QTLs identification for berry quality traits based on high-density genetic linkage map construction in table grape.

BACKGROUND: Aroma, berry firmness and berry shape are three main quality traits in table grape production, and also the important target traits in grapevine breeding. However, the information about their genetic mechanisms is limited, which results in low accuracy and efficiency of quality breeding in grapevine. Mapping and isolation of quantitative trait locus (QTLs) based on the construction of genetic linkage map is a powerful approach to decipher the genetic determinants of complex quantitative traits. RESULTS: In the present work, a final integrated map consisting of 3411 SLAF markers on 19 linkage groups (LGs) with an average distance of 0.98 cM between adjacent markers was generated using the specific length amplified fragment sequencing (SLAF-seq) technique. A total of 9 significant stable QTLs for Muscat flavor, berry firmness and berry shape were identified on two linkage groups among the hybrids analyzed over three consecutive years from 2016 to 2018. Notably, new stable QTLs for berry firmness and berry shape were found on LG 8 respectively for the first time. Based on biological function and expression profiles of candidate genes in the major QTL regions, 3 genes (VIT_08s0007g00440, VIT_08s0040g02740 and VIT_08s0040g02350) related to berry firmness and 3 genes (VIT_08s0032g01110, VIT_08s0032g01150 and VIT_08s0105g00200) linked to berry shape were highlighted. Overexpression of VIT_08s0032g01110 in transgenic Arabidopsis plants caused the change of pod shape. CONCLUSIONS: A new high-density genetic map with total 3411 markers was constructed with SLAF-seq technique, and thus enabled the detection of narrow interval QTLs for relevant traits in grapevine. VIT_08s0007g00440, VIT_08s0040g02740 and VIT_08s0040g02350 were found to be related to berry firmness, while VIT_08s0032g01110, VIT_08s0032g01150 and VIT_08s0105g00200 were linked to berry shape.

High-Quality SNP Linkage Maps Improved QTL Mapping and Genome Assembly in Populus.

With the advances in high-throughput sequencing technologies and the development of new software for extracting single nucleotide polymorphisms (SNPs) across a mapping population, it is possible to construct high-quality genetic maps with thousands of SNPs in outbred forest trees. Two parent-specific linkage maps were constructed with restriction site-associated DNA sequencing data from an F1 hybrid population derived from Populus deltoides and Populus simonii, and applied in QTL mapping and genome assembly. The female P. deltoides map contained 4018 SNPs, which were divided into 19 linkage groups under a wide range of LOD thresholds from 7 to 55. The male P. simonii map showed similar characteristics, consisting of 2097 SNPs, which also belonged to 19 linkage groups under LOD thresholds of 7 to 29. The SNP order of each linkage group was optimal among different ordering results from several available software. Moreover, the linkage maps allowed the detection of 39 QTLs underlying tree height and 47 for diameter at breast height. In addition, the linkage maps improved the anchoring of 689 contigs of P. simonii to chromosomes. The 2 parental genetic maps of Populus are of high quality, especially in terms of SNP data quality, the SNP order within linkage groups, and the perfect match between the number of linkage groups and the karyotype of Populus, as well as the excellent performances in QTL mapping and genome assembly. Both approaches for extracting and ordering SNPs could be applied to other species for constructing high-quality genetic maps.

Partial Resistance Against Erysiphe pisi and E. trifolii Under Different Genetic Control in Lathyrus cicera: Outcomes from a Linkage Mapping Approach.

Powdery mildew infections are among the most severe foliar biotrophic fungal diseases in grain legumes. Several accessions of Lathyrus cicera (chickling pea) show levels of partial resistance to Erysiphe pisi, the causal agent of pea powdery mildew, and to E. trifolii, a powdery mildew pathogen recently confirmed to infect pea and Lathyrus spp. Nevertheless, the underlying L. cicera resistance mechanisms against powdery mildews are poorly understood. To unveil the genetic control of resistance against powdery mildews in L. cicera, a recombinant inbred line population segregating for response to both species was used in resistance linkage analysis. An improved L. cicera genetic linkage map was used in this analysis. The new higher-density linkage map contains 1,468 polymorphic loci mapped on seven major and two minor linkage groups, covering a total of 712.4 cM. The percentage of the leaf area affected by either E. pisi or E. trifolii was recorded in independent screenings of the recombinant inbred line population, identifying a continuous range of resistance-susceptibility responses. Distinct quantitative trait loci (QTLs) for partial resistance against each pathogen were identified, suggesting different genetic bases are involved in the response to E. pisi and E. trifolii in L. cicera. Moreover, through comparative mapping of L. cicera QTL regions with the pea reference genome, candidate genes and pathways involved in resistance against powdery mildews were identified. This study extended the previously available genetic and genomic tools in Lathyrus species, providing clues about diverse powdery mildew resistance mechanisms useful for future resistance breeding of L. cicera and related species.

Improving Nelumbo nucifera genome assemblies using high-resolution genetic maps and BioNano genome mapping reveals ancient chromosome rearrangements.

Genetic and physical maps are powerful tools to anchor fragmented draft genome assemblies generated from next-generation sequencing. Currently, two draft assemblies of Nelumbo nucifera, the genomes of 'China Antique' and 'Chinese Tai-zi', have been released. However, there is presently no information on how the sequences are assembled into chromosomes in N. nucifera. The lack of physical maps and inadequate resolution of available genetic maps hindered the assembly of N. nucifera chromosomes. Here, a linkage map of N. nucifera containing 2371 bin markers [217 577 single nucleotide polymorphisms (SNPs)] was constructed using restriction-site associated DNA sequencing data of 181 F2 individuals and validated by adding 197 simple sequence repeat (SSR) markers. Additionally, a BioNano optical map covering 86.20% of the 'Chinese Tai-zi' genome was constructed. The draft assembly of 'Chinese Tai-zi' was improved based on the BioNano optical map, showing an increase of the scaffold N50 from 0.989 to 1.48 Mb. Using a combination of multiple maps, 97.9% of the scaffolds in the 'Chinese Tai-zi' draft assembly and 97.6% of the scaffolds in the 'China Antique' draft assembly were anchored into pseudo-chromosomes, and the centromere regions along the pseudo-chromosomes were identified. An evolutionary scenario was proposed to reach the modern N. nucifera karyotype from the seven ancestral eudicot chromosomes. The present study provides the highest-resolution linkage map, the optical map and chromosome level genome assemblies for N. nucifera, which are valuable for the breeding and cultivation of N. nucifera and future studies of comparative and evolutionary genomics in angiosperms.