The Ligon lintless -2 Short Fiber Mutation Is Located within a Terminal Deletion of Chromosome 18 in Cotton.

Extreme elongation distinguishes about one-fourth of cotton (Gossypium sp.) seed epidermal cells as "lint" fibers, useful for the textile industry, from "fuzz" fibers (<5 mm). Ligon lintless-2 (Li 2 ), a dominant mutation that results in no lint fiber but normal fuzz fiber, offers insight into pathways and mechanisms that differentiate spinnable cotton from its progenitors. A genetic map developed using 1,545 F2 plants showed that marker CISP15 was 0.4 cM from Li 2 , and "dominant" simple sequence repeat (SSR) markers (i.e. with null alleles in the Li 2 genotype) SSR7 and SSR18 showed complete linkage with Li 2 Nonrandom distribution of markers with null alleles suggests that the Li 2 phenotype results from a 176- to 221-kb deletion of the terminal region of chromosome 18 that may have been masked in prior pooled-sample mapping strategies. The deletion includes 10 genes with putative roles in fiber development. Two Glycosyltransferase Family 1 genes showed striking expression differences during elongation of wild-type versus Li 2 fiber, and virus-induced silencing of these genes in the wild type induced Li 2 -like phenotypes. Further, at least 7 of the 10 putative fiber development genes in the deletion region showed higher expression in the wild type than in Li 2 mutants during fiber development stages, suggesting coordinated regulation of processes in cell wall development and cell elongation, consistent with the hypothesis that some fiber-related quantitative trait loci comprise closely spaced groups of functionally diverse but coordinately regulated genes.

Genetic mapping and transcriptional profiling of phytoremediation and heavy metals responsive genes in sorghum.

Decreasing freshwater supply has led to the consideration of treated wastewater as an alternative source for agricultural irrigation. However, the higher content of heavy metals restricts their use in agricultural purposes. In the current study, a set of 181 SSR markers were used to perform association mapping in a structured sorghum population consisting of 107 accessions evaluated under tap-water and heavy-metals-containing water irrigation conditions. Significant differences between optimal and stressed growing conditions were shown in all evaluated phenotypic traits. Association mapping revealed 14 significant associations between 12 SSR markers and heavy metals stress indices of phenotypic traits (R2 = 11.54-30.85%). BLASTP annotation of QTLs genomic sequences identified 102 gene homologs, of which 19 are known to be implicated in phytoremediation and heavy metals tolerance. All nineteen genes exhibit differential expression patterns in heavy metals tolerant and susceptible sorghum accessions, and their transcriptional levels were highly elevated under heavy metals stress, indicating a possible functional association among these genes and provides strong evidence for their role in phytoremediation and heavy metal stress tolerance. Clustering and expression patterns of pentatricopeptide repeat and zinc finger protein genes suggest crucial roles of those genes in phytoremediation and heavy metals tolerance.

Detection of QTLs for salt tolerance in Asian barley (Hordeum vulgare L.) by association analysis with SNP markers.

Two hundred ninety-six Asian barley (Hordeum vulgare L.) accessions were assessed to detect QTLs underlying salt tolerance by association analysis using a 384 single nucleotide polymorphism (SNP) marker system. The experiment was laid out at the seedling stage in a hydroponic solution under control and 250 mM NaCl solution with three replications of four plants each. Salt tolerance was assessed by leaf injury score (LIS) and salt tolerance indices (STIs) of the number of leaves (NL), shoot length (SL), root length (RL), shoot dry weight (SDW) and root dry weight (RDW). LIS was scored from 1 to 5 according to the severity of necrosis and chlorosis observed on leaves. There was a wide variation in salt tolerance among Asian barley accessions. LIS and STI (SDW) were the most suitable traits for screening salt tolerance. Association was estimated between markers and traits to detect QTLs for LIS and STI (SDW). Seven significant QTLs were located on chromosomes 1H (2 QTLs), 2H (2 QTLs), 3H (1 QTL), 4H (1 QTL) and 5H (1 QTL). Five QTLs were associated with LIS and 2 QTLs with STI (SDW). Two QTLs associated with LIS were newly identified on chromosomes 3H and 4H.

Genetic dissection of morphological variation between cauliflower and a rapid cycling Brassica oleracea line.

To improve resolution to small genomic regions and sensitivity to small-effect loci in the identification of genetic factors conferring the enlarged inflorescence and other traits of cauliflower while also expediting further genetic dissection, 104 near-isogenic introgression lines (NIILs) covering 78.56% of the cauliflower genome, were selected from an advanced backcross population using cauliflower [Brassica oleracea var. botrytis L., mutant for Orange gene (ORG)] as the donor parent and a rapid cycling line (TO1434) as recurrent parent. Subsets of the advanced backcross population and NIILs were planted in the field for 8 seasons, finding 141 marker-trait associations for 15 leaf-, stem-, and flower-traits. Exemplifying the usefulness of these lines, we delineated the previously known flower color gene to a 4.5 MB interval on C3; a gene for small plant size to a 3.4 MB region on C8; and a gene for large plant size and flowering time to a 6.1 MB region on C9. This approach unmasked closely linked QTL alleles with opposing effects (on chr. 8) and revealed both alleles with expected phenotypic effects and effects opposite the parental phenotypes. Selected B. oleracea NIILs with short generation time add new value to widely used research and teaching materials.

Mapping of QTLs underlying flowering time in sorghum [Sorghum bicolor (L.) Moench].

Due to its critical importance in crop yield, the photoperiodic regulation of flowering time is considered an important trait in sorghum breeding programs. In this study, quantitative trait loci for flowering time were detected using an F(2) population derived from a cross between Kikuchi Zairai, a late-flowering cultivar originating from Japan and SC112, an early-flowering cultivar originating from Ethiopia. F(2) plants were grown with their parents under a natural day length and a 12 h day length. Two linkage maps were constructed using 213 simple sequence repeats markers. Nine quantitative trait loci controlling flowering time were identified in F(2) plants grown under a natural day length, whereas 7 QTLs were identified under a 12 h day length. Five QTLs controlling flowering time were shared under both of the day length conditions.

Development of diversity array technology (DArT) markers for assessment of population structure and diversity in Aegilops tauschii.

Aegilops tauschii Coss. is the D-genome donor to hexaploid bread wheat (Triticum aestivum) and is the most promising wild species as a genetic resource for wheat breeding. To study the population structure and diversity of 81 Ae. tauschii accessions collected from various regions of its geographical distribution, the genomic representation of these lines were used to develop a diversity array technology (DArT) marker array. This Ae. tauschii array and a previously developed DArT wheat array were used to scan the genomes of the 81 accessions. Out of 7500 markers (5500 wheat and 2000 Ae. tauschii), 4449 were polymorphic (3776 wheat and 673 Ae. tauschii). Phylogenetic and population structure studies revealed that the accessions could be divided into three groups. The two Ae. tauschii subspecies could also be separately clustered, suggesting that the current taxonomy might be valid. DArT markers are effective to detect very small polymorphisms. The information obtained about Ae. tauschii in the current study could be useful for wheat breeding. In addition, the new DArT array from this Ae. tauschii population is expected to be an effective tool for hexaploid wheat studies.

High-density genetic map and genome-wide association studies of aesthetic traits in Phalaenopsis orchids.

Phalaenopsis spp. represent the most popular orchids worldwide. Both P. equestris and P. aphrodite are the two important breeding parents with the whole genome sequence available. However, marker-trait association is rarely used for floral traits in Phalaenopsis breeding. Here, we analyzed markers associated with aesthetic traits of Phalaenopsis orchids by using genome-wide association study (GWAS) with the F1 population P. Intermedia of 117 progenies derived from the cross between P. aphrodite and P. equestris. A total of 113,517 single nucleotide polymorphisms (SNPs) were identified in P. Intermedia by using genotyping-by-sequencing with the combination of two different restriction enzyme pairs, Hinp1 I/Hae III and Apek I/Hae III. The size-related traits from flowers were negatively related to the color-related traits. The 1191 SNPs from Hinp1 I/ Hae III and 23 simple sequence repeats were used to establish a high-density genetic map of 19 homolog groups for P. equestris. In addition, 10 quantitative trait loci were highly associated with four color-related traits on chromosomes 2, 5 and 9. According to the sequence within the linkage disequilibrium regions, 35 candidate genes were identified and related to anthocyanin biosynthesis. In conclusion, we performed marker-assisted gene identification of aesthetic traits with GWAS in Phalaenopsis orchids.

Identification of small effect quantitative trait loci of plant architectural, flowering, and early maturity traits in reciprocal interspecific introgression population in cotton.

Plant architecture, flowering time and maturity traits are important determinants of yield and fiber quality of cotton. Genetic dissection of loci determining these yield and quality components is complicated by numerous loci with alleles conferring small differences. Therefore, mapping populations segregating for smaller numbers and sizes of introgressed segments is expected to facilitate dissection of these complex quantitative traits. At an advanced stage in the development of reciprocal advanced backcross populations from crosses between elite Gossypium hirsutum cultivar 'Acala Maxxa' (GH) and G. barbadense 'Pima S6' (GB), we undertook mapping of plant architectural traits, flowering time and maturity. A total of 284 BC4F1 and BC4F2 progeny rows, 120 in GH and 164 in GB background, were evaluated for phenotype, with only 4 and 3 (of 7) traits showing significant differences among progenies. Genotyping by sequencing yielded 3,186 and 3,026 SNPs, respectively, that revealed a total of 27 QTLs in GH background and 22 in GB, for plant height, days to flowering, residual flowering at maturity and maturity. More than of 90% QTLs identified in both backgrounds had small effects (%PV < 10), supporting the merit of this population structure to reduce background noise and small effect QTLs. Germplasm developed in this study may serve as potential pre-breeding material to develop improved cotton cultivars.

Genome structure and evolutionary history of frankincense producing Boswellia sacra.

Boswellia sacra Flueck (family Burseraceae) tree is wounded to produce frankincense. We report its de novo assembled genome (667.8 Mb) comprising 18,564 high-confidence protein-encoding genes. Comparing conserved single-copy genes across eudicots suggest >97% gene space assembly of B. sacra genome. Evolutionary history shows B. sacra gene-duplications derived from recent paralogous events and retained from ancient hexaploidy shared with other eudicots. The genome indicated a major expansion of Gypsy retroelements in last 2 million years. The B. sacra genetic diversity showed four clades intermixed with a primary genotype-dominating most resin-productive trees. Further, the stem transcriptome revealed that wounding concurrently activates phytohormones signaling, cell wall fortification, and resin terpenoid biosynthesis pathways leading to the synthesis of boswellic acid-a key chemotaxonomic marker of Boswellia. The sequence datasets reported here will serve as a foundation to investigate the genetic determinants of frankincense and other resin-producing species in Burseraceae.

Genetic analysis of QTLs controlling allelopathic characteristics in sorghum.

Mature sorghum herbage is known to contain several water-soluble secondary metabolites (allelochemicals). In this study, we investigated quantitative trait loci (QTLs) associated with allelochemical characteristics in sorghum using linkage mapping and linkage disequilibrium (LD)-based association mapping. A sorghum diversity research set (SDRS) of 107 accessions was used in LD mapping whereas, F2:3 lines derived from a cross between Japanese and African landraces were used in linkage mapping. The QTLs were further confirmed by positional (targeted) association mapping with Q+K model. The inhibitory effect of water-soluble extracts (WSE) was tested on germination and root length of lettuce seedlings in four concentrations (25%, 50%, 75% and 100%). A Significant range of variations was observed among genotypes in both types of mapping populations (P < 0.05). A total of 181 simple sequence repeats (SSRs) derived from antecedently reported map have been used for genotyping of SDRS. A genetic linkage map of 151 sorghum SSR markers was also developed on 134 F2 individuals. The total map length was 1359.3 cM, with an average distance of 8.2 cM between adjacent markers. LD mapping identified three QTLs for inhibition effect on germination and seven QTLs for root length of lettuce seedlings. Whereas, a total of six QTLs for inhibition of germination and ten QTLs for root length were detected in linkage mapping approach. The percent phenotypic variation explained by individual QTL ranged from 6.9% to 27.3% in SDRS and 9.9% to 35.6% in F2:3 lines. Regional association analysis identified four QTLs, three of them are common in other methods too. No QTL was identified in the region where major gene for sorgoleone (SOR1) has been cloned previously on chromosome 5.

Comparative Chloroplast Genomics of Endangered Euphorbia Species: Insights into Hotspot Divergence, Repetitive Sequence Variation, and Phylogeny.

Euphorbia is one of the largest genera in the Euphorbiaceae family, comprising 2000 species possessing commercial, medicinal, and ornamental importance. However, there are very little data available on their molecular phylogeny and genomics, and uncertainties still exist at a taxonomic level. Herein, we sequence the complete chloroplast (cp) genomes of two species, E. larica and E. smithii, of the genus Euphorbia through next-generation sequencing and perform a comparative analysis with nine related genomes in the family. The results revealed that the cp genomes had similar quadripartite structure, gene content, and genome organization with previously reported genomes from the same family. The size of cp genomes ranged from 162,172 to 162,358 bp with 132 and 133 genes, 8 rRNAs, 39 tRNA in E. smithii and E. larica, respectively. The numbers of protein-coding genes were 85 and 86, with each containing 19 introns. The four-junction regions were studied and results reveal that rps19 was present at JLB (large single copy region and inverted repeat b junction) in E. larica where its complete presence was located in the IRb (inverted repeat b) region in E. smithii. The sequence comparison revealed that highly divergent regions in rpoC1, rpocB, ycf3, clpP, petD, ycf1, and ndhF of the cp genomes might provide better understanding of phylogenetic inferences in the Euphorbiaceae and order Malpighiales. Phylogenetic analyses of this study illustrate sister clades of E. smithii with E. tricullii and these species form a monophyletic clade with E. larica. The current study might help us to understand the genome architecture, genetic diversity among populations, and evolutionary depiction in the genera.

First complete chloroplast genomics and comparative phylogenetic analysis of Commiphora gileadensis and C. foliacea: Myrrh producing trees.

Commiphora gileadensis and C. foliacea (family Burseraceae) are pantropical in nature and known for producing fragrant resin (myrrh). Both the tree species are economically and medicinally important however, least genomic understanding is available for this genus. Herein, we report the complete chloroplast genome sequences of C. gileadensis and C. foliacea and comparative analysis with related species (C. wightii and Boswellia sacra). A modified chloroplast DNA extraction method was adopted, followed with next generation sequencing, detailed bioinformatics and PCR analyses. The results revealed that the cp genome sizes of C. gileadensis and C. foliacea, are 160,268 and 160,249 bp, respectively, with classic quadripartite structures that comprises of inverted repeat's pair. Overall, the organization of these cp genomes, GC contents, gene order, and codon usage were comparable to other cp genomes in angiosperm. Approximately, 198 and 175 perfect simple sequence repeats were detected in C. gileadensis and C. foliacea genomes, respectively. Similarly, 30 and 25 palindromic, 15 and 25 forward, and 20 and 25 tandem repeats were determined in both the cp genomes, respectively. Comparison of these complete cp genomes with C. wightii and B. sacra revealed significant sequence resemblance and comparatively highest deviation in intergenic spacers. The phylo-genomic comparison showed that C. gileadensis and C. foliacea form a single clade with previously reported C. wightii and B. sacra from family Burseraceae. Current study reports for the first time the cp genomics of species from Commiphora, which could be helpful in understanding genetic diversity and phylogeny of this myrrh producing species.

Alignment of Rutaceae Genomes Reveals Lower Genome Fractionation Level Than Eudicot Genomes Affected by Extra Polyploidization.

Owing to their nutritional and commercial values, the genomes of several citrus plants have been sequenced, and the genome of one close relative in the Rutaceae family, atalantia (Atalantia buxifolia), has also been sequenced. Here, we show a family-level comparative analysis of Rutaceae genomes. By using grape as the outgroup and checking cross-genome gene collinearity, we systematically performed a hierarchical and event-related alignment of Rutaceae genomes, and produced a gene list defining homologous regions based on ancestral polyploidization or speciation. We characterized genome fractionation resulting from gene loss or relocation, and found that erosion of gene collinearity could largely be described by a geometric distribution. Moreover, we found that well-assembled Rutaceae genomes retained significantly more genes (65-82%) than other eudicots affected by recursive polyploidization. Additionally, we showed divergent evolutionary rates among Rutaceae plants, with sweet orange evolving faster than others, and by performing evolutionary rate correction, re-dated major evolutionary events during their evolution. We deduced that the divergence between the Rutaceae family and grape occurred about 81.15-91.74 million years ago (mya), while the split between citrus and atalantia plants occurred <10 mya. In addition, we showed that polyploidization led to a copy number expansion of key gene families contributing to the biosynthesis of vitamin C. Overall, the present effort provides an important comparative genomics resource and lays a foundation to understand the evolution and functional innovation of Rutaceae genomes.

Identification of charcoal rot resistance QTLs in sorghum using association and in silico analyses.

Charcoal rot disease, a root and stem disease caused by the soil-borne fungus Macrophomina phaseolina (Tassi) Goid., is a major biotic stress that limits sorghum productivity worldwide. Charcoal rot resistance-related parameters, e.g., pre-emergence damping-off%, post-emergence damping-off%, charcoal rot disease severity, and plant survival rates, were measured in a structured sorghum population consisting of 107 landraces. Analysis of variance of charcoal rot resistance-related parameters revealed significant variations in the response to M. phaseolina infection within evaluated accessions. Continuous phenotypic variations for resistance-related parameters were observed indicating a quantitative inheritance of resistance. The population was genotyped using 181 simple sequence repeat (SSR) markers. Association analysis identified 13 markers significantly associated with quantitative trait genes (QTLs) conferring resistance to charcoal rot disease with an R2 value ranging between 9.47 to 18.87%, nine of which are environment-specific loci. Several QTL-linked markers are significantly associated with more than one resistance-related parameter, suggesting that those QTLs might contain genes involved in the plant defense response. In silico analysis of four novel major QTLs identified 11 putative gene homologs that could be considered as candidate genes for resistance against charcoal rot disease. Cluster analysis using the genotypic data of 181 SSR markers from 107 sorghum accessions identified 12 main clusters. The results provide a basis for further functional characterization of charcoal rot disease resistance or defense genes in sorghum and for further dissection of their molecular mechanisms.

Genetic Analysis of Gossypium Fiber Quality Traits in Reciprocal Advanced Backcross Populations.

In mapping populations segregating for many loci, the large amount of variation among genotypes often masks small-effect quantitative trait loci (QTL). This problem can be reduced by development of populations with fewer chromosome segments segregating. Here, we report early QTL detection in reciprocal advanced backcross populations from crosses between elite Gossypium hirsutum L. 'Acala Maxxa' (GH) and G. barbadense L. 'Pima S6' (GB). A total of 297 BCF and BCF progeny rows-127 segregating for GB chromosome segments in GH background and 170 segregating for GH chromosome segments in GB background-were evaluated in three environments. Totals of 3186 and 3026 polymorphic single-nucleotide polymorphisms (SNPs) in GH and GB backgrounds, respectively, were identified and used for trait mapping. Small-effect QTL (<10% variance explained) made up 87 and 100% of QTL in GH and GB backgrounds, respectively. In both species, favorable alleles were found with effects being masked or neutralized by unfavorable alleles, with greater scope for improvement of GH than GB by introgressive breeding. A total of three stable QTL-two in GH background for fiber elongation (ELO) and micronaire (MIC) and one in GB background for upper-half mean length (UHM)-were identified in two out of three environments. Curiously, only four QTL-three for UHM and one for ELO-showed the expected opposite effects in reciprocal backgrounds, perhaps reflecting the combined consequences of epistasis, small phenotypic effects, and low coverage of some genomic regions. Along with new information for marker-assisted breeding, this study adds to knowledge that can be used to unravel complex genetic networks governing fiber quality traits.

Advanced Backcross QTL Analysis of Fiber Strength and Fineness in a Cross between Gossypium hirsutum and G. mustelinum.

The molecular genetic basis of cotton fiber strength and fineness in crosses between Gossypium mustelinum and Gossypium hirsutum (Upland cotton) was dissected using 21 BC3F2 and 12 corresponding BC3F2:3 and BC3F2:4 families. The BC3F2 families were genotyped with simple sequence repeat markers from a G. hirsutum by G. mustelinum linkage map, and the three generations of BC3-derived families were phenotyped for fiber strength (STR) and fineness (Micronaire, MIC). A total of 42 quantitative trait loci (QTLs) were identified through one-way analysis of variance, including 15 QTLs for STR and 27 for MIC, with the percentage of variance explained by individual loci averaging 13.86 and 14.06%, respectively. Eighteen of the 42 QTLs were detected at least twice near the same markers in different generations/families or near linked markers in the same family, and 28 of the 42 QTLs were identified in both mixed model-based composite interval mapping and one-way variance analyses. Alleles from G. mustelinum increased STR for eight of 15 and reduced MIC for 15 of 27 QTLs. Significant among-family genotypic effects (P < 0.001) were detected in 13 and 10 loci for STR and MIC respectively, and five loci showed significant (P < 0.001) genotype × family interaction for MIC. These results support the hypothesis that fiber quality improvement for Upland cotton could be realized by introgressing G. mustelinum alleles although complexities due to the different effects of genetic background on introgressed chromatin might be faced. Building on prior work with G. barbadense, G. tomentosum, and G. darwinii, QTL mapping involving introgression of G. mustelinum alleles offers new allelic variation to Upland cotton germplasm.

Genome-wide divergence, haplotype distribution and population demographic histories for Gossypium hirsutum and Gossypium barbadense as revealed by genome-anchored SNPs.

Use of 10,129 singleton SNPs of known genomic location in tetraploid cotton provided unique opportunities to characterize genome-wide diversity among 440 Gossypium hirsutum and 219 G. barbadense cultivars and landrace accessions of widespread origin. Using the SNPs distributed genome-wide, we examined genetic diversity, haplotype distribution and linkage disequilibrium patterns in the G. hirsutum and G. barbadense genomes to clarify population demographic history. Diversity and identity-by-state analyses have revealed little sharing of alleles between the two cultivated allotetraploid genomes, with a few exceptions that indicated sporadic gene flow. We found a high number of new alleles, representing increased nucleotide diversity, on chromosomes 1 and 2 in cultivated G. hirsutum as compared with low nucleotide diversity on these chromosomes in landrace G. hirsutum. In contrast, G. barbadense chromosomes showed negative Tajima's D on several chromosomes for both cultivated and landrace types, which indicate that speciation of G. barbadense itself, might have occurred with relatively narrow genetic diversity. The presence of conserved linkage disequilibrium (LD) blocks and haplotypes between G. hirsutum and G. barbadense provides strong evidence for comparable patterns of evolution in their domestication processes. Our study illustrates the potential use of population genetic techniques to identify genomic regions for domestication.