A high-continuity and annotated tomato reference genome.

BACKGROUND: Genetic and functional genomics studies require a high-quality genome assembly. Tomato (Solanum lycopersicum), an important horticultural crop, is an ideal model species for the study of fruit development. RESULTS: Here, we assembled an updated reference genome of S. lycopersicum cv. Heinz 1706 that was 799.09 Mb in length, containing 34,384 predicted protein-coding genes and 65.66% repetitive sequences. By comparing the genomes of S. lycopersicum and S. pimpinellifolium LA2093, we found a large number of genomic fragments probably associated with human selection, which may have had crucial roles in the domestication of tomato. We also used a recombinant inbred line (RIL) population to generate a high-density genetic map with high resolution and accuracy. Using these resources, we identified a number of candidate genes that were likely to be related to important agronomic traits in tomato. CONCLUSION: Our results offer opportunities for understanding the evolution of the tomato genome and will facilitate the study of genetic mechanisms in tomato biology.

Mapping QTLs for 1000-grain weight and genes controlling hull type using SNP marker in Tartary buckwheat (Fagopyrum tataricum).

BACKGROUND: Tartary buckwheat (Fagopyrum tataricum), an important pseudocereal crop, has high economic value due to its nutritional and medicinal properties. However, dehulling of Tartary buckwheat is difficult owing to its thick and tough hull, which has greatly limited the development of the Tartary buckwheat processing industry. The construction of high-resolution genetic maps serves as a basis for identifying quantitative trait loci (QTLs) and qualitative trait genes for agronomic traits. In this study, a recombinant inbred lines (XJ-RILs) population derived from a cross between the easily dehulled Rice-Tartary type and Tartary buckwheat type was genotyped using restriction site-associated DNA (RAD) sequencing to construct a high-density SNP genetic map. Furthermore, QTLs for 1000-grain weight (TGW) and genes controlling hull type were mapped in multiple environments. RESULTS: In total, 4151 bin markers comprising 122,185 SNPs were used to construct the genetic linkage map. The map consisted of 8 linkage groups and covered 1444.15 cM, with an average distance of 0.35 cM between adjacent bin markers. Nine QTLs for TGW were detected and distributed on four loci on chromosome 1 and 4. A major locus detected in all three trials was mapped in 38.2-39.8 cM region on chromosome 1, with an LOD score of 18.1-37.0, and explained for 23.6-47.5% of the phenotypic variation. The genes controlling hull type were mapped to chromosome 1 between marker Block330 and Block331, which was closely followed by the major locus for TGW. The expression levels of the seven candidate genes controlling hull type present in the region between Block330 and Block336 was low during grain development, and no significant difference was observed between the parental lines. Six non-synonymous coding SNPs were found between the two parents in the region. CONCLUSIONS: We constructed a high-density SNP genetic map for the first time in Tartary buckwheat. The mapped major loci controlling TGW and hull type will be valuable for gene cloning and revealing the mechanism underlying grain development and easy dehulling, and marker-assisted selection in Tartary buckwheat.

Genetic mapping of male sterility and pollen fertility QTLs in triticale with sterilizing Triticum timopheevii cytoplasm.

Cytoplasmic male sterility (CMS) phenomenon is widely exploited in commercial hybrid seed production in economically important crop species, including rye, wheat, maize, rice, sorghum, cotton, sugar beets, and many vegetables. Although some commercial successes, little is known about QTLs responsible for the trait in case of triticale with sterilizing Triticum timopheevii (Tt) cytoplasm. Recombinant inbred line (RIL) F6 mapping population encompassing 182 individuals derived from the cross of individual plants representing the HT352 line and cv Borwo was employed for genetic map construction using SNP markers and identification of QTLs conferring pollen sterility in triticale with CMS Tt. The phenotypes of the F1 lines resulting from crossing of the HT352 (Tt) with HT352 (maintainer) × Borwo were determined by assessing the number of the F2 seeds per spike. A genetic map with 21 linkage groups encompasses 29,737 markers and spanned over the distance of 2549 cM. Composite (CIM) and multiple (MIM) interval mappings delivered comparable results. Single QTLs mapped to the 1A, 1B, 2A, 2R, 3B, 3R, 4B, and 5B chromosomes, whereas the 5R and 6B chromosomes shared 3 and 2 QTLs, respectively. The QTLs with the highest LOD score mapped to the 5R, 3R, 1B, and 4B chromosomes; however, the QRft-5R.3 has the highest explained variance of the trait.

Genotyping by sequencing for SNP marker development in onion.

Onion (Allium cepa) is not highly tractable for development of molecular markers due to its large (16 gigabases per 1C) nuclear genome. Single nucleotide polymorphisms (SNPs) are useful for genetic characterization and marker-aided selection of onion because of codominance and common occurrence in elite germplasm. We completed genotyping by sequencing (GBS) to identify SNPs in onion using 46 F2 plants, parents of the F2 plants (Ailsa Craig 43 and Brigham Yellow Globe 15-23), two doubled haploid (DH) lines (DH2107 and DH2110), and plants from 94 accessions in the USDA National Plant Germplasm System (NPGS). SNPs were called using the TASSEL 3.0 Universal Network Enabled Analysis (UNEAK) bioinformatics pipeline. Sequences from the F2 and DH plants were used to construct a pseudo-reference genome against which genotypes from all accessions were scored. Quality filters were used to identify a set of 284 high quality SNPs, which were placed onto an existing genetic map for the F2 family. Accessions showed a moderate level of diversity (mean He = 0.341) and evidence of inbreeding (mean F = 0.592). GBS is promising for SNP discovery in onion, although lack of a reference genome required extensive custom scripts for bioinformatics analyses to identify high quality markers.

Identification and Mapping of Stable QTLs for Seed Oil and Protein Content in Soybean [Glycine max (L.) Merr.].

This research aimed to identify stable quantitative trait loci (QTL) associated with oil and protein content in soybean. A population of 196 recombinant inbred lines (RILs) derived from Huachun 2 × Wayao was used to evaluate these target traits. A high-density genetic linkage map was constructed by using high-throughput genome-wide sequencing technology, which contained 3413 recombination bin markers and spanned 5400.4 cM with an average distance of 1.58 cM between markers. Eighteen stable QTLs controlling oil and protein content were detected. Among them, qOil-11-1 was identified for the first time as a novel QTL, while qOil-5-1, qPro-10-1, and qPro-14-1 were strong and stable QTLs with high log-likelihood (LOD) values. Sixteen differentially expressed genes (DEGs) within these four QTLs were shown to be highly expressed during seed development based on RNA sequencing (RNA-seq) data analysis. Our results may contribute toward gene mining and marker-assisted selection (MAS) for breeding a high-quality soybean in the future.

A SNP-Based High-Density Genetic Map of Leaf and Fruit Related Quantitative Trait Loci in Wolfberry (Lycium Linn.).

Wolfberry (Lycium Linn. 2n = 24) fruit, Gouqizi, is a perennial shrub, traditional food and medicinal plant resource in China. Leaf and fruit related characteristics are economically important traits that are the focus for genetic improvement, but few studies into the molecular genetics of this crop have been reported to date. Here, an F1 population (302 individuals) derived from a cross between "NO.1 Ningqi" (Lycium barbarum L.) and "Chinese gouqi" (Lycium chinese Mill.) was constructed. We recorded fruit weight, longitude, diameter and index along with leaf length, width and index for three consecutive years from 2015 to 2017. Based on this population and these phenotypic data, we constructed the first high-density genetic map of Lycium using specific length amplified fragment sequencing (SLAF-seq) and analyzed quantitative trait loci (QTLs). The map contains 6733 single nucleotide polymorphisms and 12 linkage groups (LG) with a total map distance of 1702.45 cM and an average map distance of 0.253 cM. A total of 55 QTLs were mapped for more than 2 years, of which 18 stable QTLs for fruit index on LG 11, spanning an interval of 73.492-90.945 cM, were detected. qFI11-15 for fruit index was an impressive QTL with logarithm of odds (LOD) and proportion of variance explained (PEV) values reaching 11.07 and 19.7%, respectively. The QTLs on LG 11 were gathered tightly, having an average interval of less than 1 cM per QTL, suggesting that there might be a cluster region controlling fruit index. Remarkably, qLI10-2 and qLI11-2 for leaf index were detectable for 3 years. These results give novel insight into the genetic control of leaf and fruit related traits in Lycium and provide robust support for undertaking further positional cloning studies and implementing marker-assisted selection in seedlings.

Genetic imprints of domestication for disease resistance, oil quality, and yield component traits in groundnut (Arachis hypogaea L.).

Ploidy difference between wild Arachis species and cultivated genotypes hinder transfer of useful alleles for agronomically important traits. To overcome this genetic barrier, two synthetic tetraploids, viz., ISATGR 1212 (A. duranensis ICG 8123 × A. ipaensis ICG 8206) and ISATGR 265-5A (A. kempff-mercadoi ICG 8164 × A. hoehnei ICG 8190), were used to generate two advanced backcross (AB) populations. The AB-populations, namely, AB-pop1 (ICGV 91114 × ISATGR 1212) and AB-pop2, (ICGV 87846 × ISATGR 265-5A) were genotyped with DArT and SSR markers. Genetic maps were constructed for AB-pop1 and AB-pop2 populations with 258 loci (1415.7 cM map length and map density of 5.5 cM/loci) and 1043 loci (1500.8 cM map length with map density of 1.4 cM/loci), respectively. Genetic analysis identified large number of wild segments in the population and provided a good source of diversity in these populations. Phenotyping of these two populations identified several introgression lines with good agronomic, oil quality, and disease resistance traits. Quantitative trait locus (QTL) analysis showed that the wild genomic segments contributed favourable alleles for foliar disease resistance while cultivated genomic segments mostly contributed favourable alleles for oil quality and yield component traits. These populations, after achieving higher stability, will be useful resource for genetic mapping and QTL discovery for wild species segments in addition to using population progenies in breeding program for diversifying the gene pool of cultivated groundnut.

High-density SNP linkage map construction and QTL mapping for flavonoid-related traits in a tea plant (Camellia sinensis) using 2b-RAD sequencing.

BACKGROUND: Flavonoids are important components that confer upon tea plants a unique flavour and health functions. However, the traditional breeding method for selecting a cultivar with a high or unique flavonoid content is time consuming and labour intensive. High-density genetic map construction associated with quantitative trait locus (QTL) mapping provides an effective way to facilitate trait improvement in plant breeding. In this study, an F1 population (LJ43×BHZ) was genotyped using 2b-restriction site-associated DNA (2b-RAD) sequencing to obtain massive single nucleotide polymorphism (SNP) markers to construct a high-density genetic map for a tea plant. Furthermore, QTLs related to flavonoids were identified using our new genetic map. RESULTS: A total of 13,446 polymorphic SNP markers were developed using 2b-RAD sequencing, and 4,463 of these markers were available for constructing the genetic linkage map. A 1,678.52-cM high-density map at an average interval of 0.40 cM with 4,217 markers, including 427 frameset simple sequence repeats (SSRs) and 3,800 novel SNPs, mapped into 15 linkage groups was successfully constructed. After QTL analysis, a total of 27 QTLs related to flavonoids or caffeine content (CAF) were mapped to 8 different linkage groups, LG01, LG03, LG06, LG08, LG10, LG11, LG12, and LG13, with an LOD from 3.14 to 39.54, constituting 7.5% to 42.8% of the phenotypic variation. CONCLUSIONS: To our knowledge, the highest density genetic map ever reported was constructed since the largest mapping population of tea plants was adopted in present study. Moreover, novel QTLs related to flavonoids and CAF were identified based on the new high-density genetic map. In addition, two markers were located in candidate genes that may be involved in flavonoid metabolism. The present study provides valuable information for gene discovery, marker-assisted selection breeding and map-based cloning for functional genes that are related to flavonoid content in tea plants.

[Screening of different AFLP fragments between near-isogenic lines of male sterile and fertile Salvia miltiorrhiza and their comparison analysis].

There is distinctive advantage of using male sterile lines to breed new cultivar and produce hybrids, when compared with general breeding method on yield and quality. In our previous work, near-isogenic lines (NILs) of male sterile and fertile Salvia miltiorrhiza have been obtained through continuous hybridization in many years. In this investigation, 378 primer combination were screened by using AFLP and BSA technique, in which 26 markers amplified from seven primers were found to tightly link to male sterile gene. Based on these markers, two linkage genetic maps were constructed. A 2 027,2 028 bp fragment was amplifed from NILs of fertile and sterile S. miltiorrhiza, respectively, using genome walking technique and previous E11/M4-208 marker as template. Four base mutations were found in intron when comparing both fragments. Among all different markers between NILs of male sterile and fertile S. miltiorrhiza, four was found to have 100% identities to chromosome 1, 3 and 5 of Arabidopsis, namely, E01/M09-418, E05/M13-308, E05/M04-750 and E01/M01-204. The E01/M09-418 marker was very close to male sterile gene of S. miltiorrhiza with distance of 2.1 cM, which also had 100% identities to male sterile gene MS2 in Arabidopsis. Both were distributed in chromosome 3 of Arabidopsis. The 2 028 bp fragment also had 100% identities to MS2 gene. Another E05/M04-750 marker that had 100% identities to chromosome 5 of Arabidopsis was found to have high identities to POP085-M05 gene of poplars and low affinity calcium antiporter CAX2 of Arabidopsis with very low E-value. The constructed genetic map and differential fragments with potential functions found in this study provide a solid foundation to lock male sterile genes in S. miltiorrhiza genome and to discover their functions.

Construction of reference chromosome-scale pseudomolecules for potato: integrating the potato genome with genetic and physical maps.

The genome of potato, a major global food crop, was recently sequenced. The work presented here details the integration of the potato reference genome (DM) with a new sequence-tagged site marker-based linkage map and other physical and genetic maps of potato and the closely related species tomato. Primary anchoring of the DM genome assembly was accomplished by the use of a diploid segregating population, which was genotyped with several types of molecular genetic markers to construct a new ~936 cM linkage map comprising 2469 marker loci. In silico anchoring approaches used genetic and physical maps from the diploid potato genotype RH89-039-16 (RH) and tomato. This combined approach has allowed 951 superscaffolds to be ordered into pseudomolecules corresponding to the 12 potato chromosomes. These pseudomolecules represent 674 Mb (~93%) of the 723 Mb genome assembly and 37,482 (~96%) of the 39,031 predicted genes. The superscaffold order and orientation within the pseudomolecules are closely collinear with independently constructed high density linkage maps. Comparisons between marker distribution and physical location reveal regions of greater and lesser recombination, as well as regions exhibiting significant segregation distortion. The work presented here has led to a greatly improved ordering of the potato reference genome superscaffolds into chromosomal "pseudomolecules".