Backcross Families Research Articles

A genome-guided strategy for climate resilience in American chestnut restoration populations

American chestnut (Castanea dentata) is a deciduous tree species of eastern North America that was decimated by the introduction of the chestnut blight fungus (Cryphonectria parasitica) in the early 20th century. Although millions of American chestnuts survive as root collar sprouts, these trees rarely reproduce. Thus, the species is considered functionally extinct. American chestnuts with improved blight resistance have been developed through interspecific hybridization followed by conspecific backcrossing, and by genetic engineering. Incorporating adaptive genomic diversity into these backcross families and transgenic lines is important for restoring the species across broad climatic gradients. To develop sampling recommendations for ex situ conservation of wild adaptive genetic variation, we coupled whole-genome resequencing of 384 stump sprouts with genotype-environment association analyses and found that the species range can be subdivided into three seed zones characterized by relatively homogeneous adaptive allele frequencies. We estimated that 21 to 29 trees per seed zone will need to be conserved to capture most extant adaptive diversity. We also resequenced the genomes of 269 backcross trees to understand the extent to which the breeding program has already captured wild adaptive diversity, and to estimate optimal reintroduction sites for specific families on the basis of their adaptive portfolio and future climate projections. Taken together, these results inform the development of an ex situ germplasm conservation and breeding plan to target blight-resistant breeding populations to specific environments and provides a blueprint for developing restoration plans for other imperiled tree species.

Marker assisted breeding of Sub1 introgressed rice (Oryza sativa L.) lines and identification of stable variety MTU 1232 suitable for flood prone ecosystem

The Development of flood-tolerant rice varieties is a prerequisite for climate resilience in flood-prone areas. The present study aimed to develop a stable, high, yielding, and tolerant rice variety against flash floods and stagnant flooding across multiple environments. Sub1A was incorporated into a popular rice variety MTU 1075 using Swarna-sub1 as a donor to generate BC3F5 families. Sub1BC2 was used as a foreground marker for selection, a proxy for the Sub1A gene. RM23865 and RM464 on Chromosome 9 were used as recombinant markers. Backcross families from the BC3F2 generation were evaluated under two weeks of flash floods 15 days after transplanting. This was followed by stagnant flooding and survived BC3F4 families were used for background selection using a 50K high-density SNP chip. The nine best families identified were included in the field trial evaluation under eight environments. Consequently, MTU Rice 1232 was identified as a high-yielding, flood-tolerant rice variety using Additive Main effects and Multiplicative Interaction stability analysis. The MTU Rice 1232 ranked first by the Best linear unbiased prediction (BLUP) based ranking parameters and 4th based on stability parameters ranking. This flood-tolerant rice variety can tolerate both flash floods and stagnant flooding and possesses an 80% survival rate. It has a yield potential of 3792 Kg ha-1 under severe floods and 6000 Kg ha-1 under normal conditions.

A study of pod constriction in a peanut population with mixed wild and cultivated genetics

AbstractWild relatives comprise a diverse genetic source from which to introduce beneficial alleles for peanut breeding. However, when using wild species for breeding, unadapted agronomic characteristics are often co‐inherited along with favorable traits. Previously, an advanced backcross population was developed from a cross between Arachis hypogaea and the wild‐derived induced allotetraploid (Arachis batizocoi × Arachis stenosperma)4x to incorporate nematode resistance genes into elite peanut. We observed additional wild alleles unintentionally inherited throughout the genome, possibly causing variation in several pod and seed characteristics. To identify the wild introgressions controlling the variation in pod constriction (PC), a critical trait for peanut's acceptability and marketability, we performed an association analysis using 419 SNPs and phenotypic data of 72 BC3F2/3 families. An introgression on the top of chromosome B08 (pod constriction‐quantitative trait locus [PC‐QTL], 3.7 Mbp) derived from A. batizocoi was found to be controlling 47.3% of the PC variation. To confirm the PC‐QTL, six BC3F4 families harboring the wild loci in a homozygous or heterozygous state were selected and used for segregation analysis. All progenies from the backcross families carrying the homozygous allele had severely constricted pods, whereas progenies from families with the heterozygous allele were segregated for PC. Finally, analyses using end‐point genotyping with four linked SNP assays confirmed the association of the PC‐QTL with PC. The present work studied PC and developed DNA markers to assist the selection against constricted pods while retaining the new and beneficial wild‐derived genes.

Validation of an alternative small stem assay for blight resistance in backcross hybrid chestnuts (Castanea spp.) and recommendations for its expanded use.

We evaluated an alternative small stem assay (AltSSA) for blight resistance in backcross hybrid chestnuts (Castanea dentata/mollissima). Whereas standard small stem assays (SSAs) are done by inoculating small incisions in stems, in our AltSSA, 4 to 5 mm stems are cut off, and the exposed (living) stem tips are inoculated with discs of Cryphonectria parasitica inoculum and temporarily covered with plastic sleeves. Intended primarily for forward selection, this method was designed to be easy to implement, to consistently induce cankering, and to better enable seedling recovery via the development of lateral shoots from the lower stem. After 90+ days, cankers are evaluated and removed, and seedlings are prepared for out-planting. Previous results showed that AltSSAs performed at least as well as a common SSA method in distinguishing resistant and susceptible types. In this follow-up analysis of 35 lines of backcross seedlings studied in 2020 and 2021, we show that mean orange zone canker length (OZCL) and a multifactor PCA-based blight resistance index gave results consistent with predictions derived from the blight resistance phenotype (two methods) and percent American chestnut ancestry of the parents of each line. As expected, based upon the apparent polygenic inheritance of blight resistance in backcross chestnuts, mean OZCL of backcross families ranged from intermediate (F1-level) to low (American chestnut-level). Consistent with prior results, canker production was near 100%, survivorship after out-planting was very high, and post-inoculation deaths were apparently unrelated to the stem tip inoculations (e.g., unrelated basal cankering). Altogether, these results suggest that the AltSSA is a viable method for early detection of relative blight resistance in seedlings and may enable a reduction in the numbers of trees out-planted and placed under care for long-term evaluation and breeding.

Genetic architectures of postmating isolation and morphology of two highly diverged rockfishes (genus Sebastes).

Postmating isolation is thought to be an important driver of the late stages of speciation. However, relatively little is empirically known about the process compared with other isolating mechanisms that drive the early stages of speciation, especially in non-model organisms. We characterized the genetic architecture of postmating isolation between 2 rockfishes, Sebastes schlegelii and S. trivittatus, whose reproductive isolation is complete. We examined transmission ratio distortion (TRD) patterns of genetic markers in 2 reciprocal backcross populations. Markers showing either of the 2 types of TRD was widespread across the genome, with some of the distorted markers forming extensive clusters around the recombination coldspots. These suggest that the postmating isolation effectively prevents gene flow across the genome and the recombination landscape contributes to the genetic architecture. Comparisons between 2 backcross families and 2 developmental stages showed little similarity in the distorted markers, suggesting asymmetry and stage specificity of the isolation. This may be due to hybrid incompatibility involving maternal factors or extrinsic selection. The lack of sex-ratio distortion in the mapping families suggested that Haldane's rule in terms of hybrid inviability does not hold. Additionally, quantitative trait loci (QTL) mapping detected significant QTLs for sex and the morphological traits relevant to speciation and convergence of rockfishes, including body coloration. Genes in the melanocortin system, including agouti-signaling protein 1 (asip1) and melanocortin 1 receptor (mc1r), might underlie the horizontal and vertical color patterns on the body, respectively. These findings constitute an essential step toward a comprehensive understanding of speciation and morphological diversification of rockfishes.

Convergent shifts in soil fungal communities associated with Fagaceae reforestation in the Southern Appalachian Mountains

Forests of the Southern Appalachian Mountains were historically dominated by hardwood species within the family Fagaceae, e.g., American chestnut (Castanea dentata) and white oak (Quercus alba) among others. Due to numerous biotic and abiotic stressors, including pathogen pressure, populations of many Fagaceae have been greatly reduced, or functionally eliminated as is the case with American chestnut. This has led to reforestation efforts designed to increase American chestnut and white oak populations in this region, but success has been minimal. Since soil fungal communities are crucial to plant health, understanding how reforestation efforts alter soil fungal communities will inform reforestation efforts. Here, using nursery-reared bareroot seedlings of American chestnut, Chinese chestnut, 3rd generation backcross chestnut hybrids (BC3F3), and white oaks outplanted at a locally xeric site in the Southern Appalachian Mountains, we investigated if and how these species and backcross families within species differentially impact soil fungal communities. We demonstrate that after three years of growth, plant-associated soil fungal communities change similarly among different Fagaceae and are distinct from pre-planting soil communities. Interestingly, we observed differential shifts in fungal functional guilds among the Fagaceae species, although these were rather minimal. Taken together, the largely convergent shifts in soil communities across Fagaceae species suggest that these tree species may have similar impacts on soils and/or share similar communities that become enriched in closely related fungal species. The similarity in shared mutualist fungal communities suggests that companion planting or reforestation of genetically disease resistant American chestnut adjacent to establishing white oak trees might enhance survival and growth of both species at xeric sites.

Population Genomics of Nonrecessive Resistance to Bt Toxin Cry1Ac in Helicoverpa armigera From Northern China.

Transgenic crops that produce insecticidal proteins from Bacillus thuringiensis (Bt) have provided control of some key pests since 1996. However, the evolution of resistance by pests reduces the benefits of Bt crops. Resistance to Bt crops that is not recessively inherited is especially challenging to manage. Here we analyzed nonrecessive resistance to Bt toxin Cry1Ac in eight field populations of Helicoverpa armigera sampled in 2018 from northern China, where this global pest has been exposed to Cry1Ac in Bt cotton since 1997. Bioassays revealed 7.5% of field-derived larvae were resistant to Cry1Ac of which 87% had at least one allele conferring nonrecessive resistance. To analyze this nonrecessive resistance, we developed and applied a variant of a genomic mapping approach called quantitative trait locus (QTL)-seq. This analysis identified a region on chromosome 10 associated with nonrecessive resistance to Cry1Ac in all 21 backcross families derived from field-collected moths. Individual sequencing revealed that all 21 field-collected resistant grandparents of the backcross families had a previously identified dominant point mutation in the tetraspanin gene HaTSPAN1 that occurs in the region of chromosome 10 identified by QTL-seq. QTL-seq also revealed a region on chromosome 26 associated with nonrecessive resistance in at most 14% of the backcross families. Overall, the results imply the point mutation in HaTSPAN1 is the primary genetic basis of nonrecessive resistance to Cry1Ac in field populations of H. armigera from northern China. Moreover, because nonrecessive resistance is predominant, tracking the frequency of this point mutation could facilitate resistance monitoring in the region.

Evaluation of morphologically important traits of deep water rice from advanced backcross families

Local germplasm of deep water rice possesses low yield productivity. Our studies showed that grain yield in Laxmi Digha can be increased significantly through backcross program with the high yielding rice variety Binadhan-18. Advanced backcross progenies (Laxmi Digha x Binadhan-18) exhibited higher values for most of the yield and yield-related traits analyzed in comparison with the parent. Number of tillers plant-1 and filled grains panicles-1 are positively correlated to yield and those resulted in higher grain yield in backcross families. In S103, S104, S105, S106 and S111 families, yield increase ranges from 52.97 to 84.1% when compared to Laxmi Digha parent. Five families, viz., S103, S104, S105, S106 and S111, were identified through yield increase analysis as most promising genotypes for cultivar development and processes variation in special characteristics viz. aerial root number and Knee capacity. The backcross families will be used to develop a deep water rice variety and efficient source of genetic diversity for future breeding programs.

Mapping and Validation of Alectra vogelii Resistance in the Cowpea Landrace B301

Cowpea is the most important food legume in West and Central Africa and a valuable economic commodity in the region. Among the major biotic constraints to cowpea production are root parasitic weeds of which Alectra vogelii (Benth.) is of increasing importance. The cowpea landrace B301 was previously identified as a source of Alectra resistance, but neither the genes nor genomic loci conferring this resistance have been mapped. Therefore, to map and identify genetic markers linked to Alectra resistance for use in the molecular improvement of cowpea, we developed an F2 population from a cross of the susceptible variety 524B with B301. The population was phenotyped for resistance to A. vogelii and genotyped with a cowpea high density single nucleotide polymorphism (SNP) microarray. Putative resistance loci were mapped in F2 populations by categorical trait–multiple interval mapping and validated by selective genotyping. Selective genotyping indicated that the resistance loci on Vu04 (Rav1) and Vu11 (Rav2) were significantly associated with resistance (p ≤ 0.01). Using marker assisted backcrossing, the two resistance loci were introgressed independently into the susceptible 524B genetic background. Phenotyping and genotyping of the segregating backcross families delineated Rav1 to a 10 cM on chromosome 4 and Rav2 to a 6.7 cM interval in chromosome 11. These two loci are desirable for breeding Alectra resistant cowpea varieties due to their simple inheritance and ability to independently confer complete immunity to the parasite.

ZmCCT haplotype H5 improves yield, stalk-rot resistance, and drought tolerance in maize.

The ZmCCT locus underlies both stalk-rot resistance and photoperiod sensitivity in maize (Zea mays L.). We previously introduced nine resistant ZmCCT haplotypes into seven elite but susceptible maize inbred lines (containing the haplotype H1) to generate 63 backcross families. Here, we continued backcrossing, followed by selfing, to develop 63 near-isogenic lines (NILs). We evaluated 22 of these NILs for stalk-rot resistance and flowering time under long-day conditions. Lines harboring the haplotype H5 outperformed the others, steadily reducing disease severity, while showing less photoperiod sensitivity. To demonstrate the value of haplotype H5 for maize production, we selected two pairs of NILs, 83B28H1/83B28H5 and A5302H1/A5302H5, and generated F1 hybrids with the same genetic backgrounds but different ZmCCT alleles: 83B28H1 × A5302H1, 83B28H1 × A5302H5, 83B28H5 × A5302H1, and 83B28H5 × A5302H5. We performed field trials to investigate yield/yield-related traits, stalk-rot resistance, flowering time, and drought/salt tolerance in these four hybrids. 83B28H5 × A5302H1 performed the best, with significantly improved yield, stalk-rot resistance, and drought tolerance compared to the control (83B28H1 × A5302H1). Therefore, the ZmCCT haplotype H5 has great value for breeding maize varieties with high yield potential, stalk-rot resistance, and drought tolerance.

Inheritance of Compact Growth Habit, and Investigation of Linkage to Weeping Architecture and Purple Leaf Color in Eastern Redbud (Cercis canadensis L.)

Eastern redbud (Cercis canadensis L.) is a commonly used small landscape tree. Compact growth, purple leaf color, and weeping architecture are three popular ornamental phenotypes. Inheritances of weeping architecture and purple leaves have been reported previously. Inheritance of compact growth habit and its genetic linkage with the weeping and purple leaf genes have not been reported. In the present research, the inheritance of compact growth derived from ‘Ace of Hearts’ was explored in the F1, F2, and reciprocal backcross families resulting from the controlled hybridization of ‘Ruby Falls’ (normal growth/weeping architecture/purple leaf) × ‘Ace of Hearts’ (compact growth/nonweeping architecture/green leaf). All 27 F1 individuals were nonweeping, green-leaved, and noncompact. A total of 572 F2 progeny were obtained, and subsequent analysis of segregation revealed a single recessive gene controlled compact growth habit. Analysis of reciprocal backcross families confirmed this result as well. Weeping architecture and purple leaf color were also controlled by single recessive genes, confirming findings presented in previous studies in another redbud family. No linkage between the three genes was detected. This research is the first to report the inheritance of compact growth in eastern redbud and confirms independent assortment between the compact, purple leaf, and weeping genes.

Identification of Pmel17 for golden skin color using linkage mapping in Mozambique tilapia

Fish color is an important trait involved in many biological functions and related to commercial value of fish. However, our understanding of the molecular mechanisms underlying color variations in fish species is limited. In this study, we found 12 natural golden mutants in a Mozambique tilapia population. Analysis of the inheritance pattern of the golden and blackish skin colors in nine backcross families revealed that the golden phenotype is a recessive mutant determined by a single locus and follows simple Mendelian inheritance. Using linkage mapping, fine mapping and analyses of gene expressions, we uncovered that the Pmel17 gene was responsible for golden and black skin colors. Further sequence analysis of the gene discovered that an insertion in 3′ UTR of Pmel17 was fully associated with the golden skin color in 1835 tilapias. Expression analysis showed that Pmel17 with the insertion reduced the transcripts of Pmel17. These results indicate that the insertion in 3′ UTR of Pmel17 is responsible for the golden skin color in tilapia. The insertion can be used as a reliable DNA marker to accelerate the breeding for golden color.

Association mapping analysis of oil palm interspecific hybrid populations and predicting phenotypic values via machine learning algorithms

AbstractThe genotyping‐by‐sequencing (GBS) approach was applied to genotype selected interspecific hybrid (F1) and backcross (BC2) families of Elaeis oleifera and Elaeis guineensis. Genome‐wide linkage disequilibrium (LD) was estimated at 150‐kb pairwise distance for r2 values of 0.17 and 0.42 for F1 and BC2, respectively. Single marker‐trait association analysis identified 47 markers associated with five fatty acid composition (FAC) traits (C16:0, C18:0, C18:1, C18:2 and iodine value [IV]) in F1, and 12 significant markers linked to oleic acid (C18:1) and vegetative traits (petiole width and mean leaf width) in BC. Within the QTL region associated with FAC traits, we identified key candidate genes influencing fatty acid synthesis. We implemented two machine learning algorithms, namely random forest and gradient boosting, to evaluate the ability of significant markers in predicting phenotype values. We also demonstrated the contribution of different marker combinations on trait values via prediction trees. This is the first attempt to evaluate the predictive ability of a combination of markers associated with traits identified from association mapping analysis in oil palm populations.

Identification of Distant Regulatory Elements Using Expression Quantitative Trait Loci Mapping for Heat-Responsive Genes in Oysters.

Many marine ectotherms, especially those inhabiting highly variable intertidal zones, develop high phenotypic plasticity in response to rapid climate change by modulating gene expression levels. Herein, we examined the regulatory architecture of heat-responsive gene expression plasticity in oysters using expression quantitative trait loci (eQTL) analysis. Using a backcross family of Crassostrea gigas and its sister species Crassostrea angulata under acute stress, 56 distant regulatory regions accounting for 6–26.6% of the gene expression variation were identified for 19 heat-responsive genes. In total, 831 genes and 164 single nucleotide polymorphisms (SNPs) that could potentially regulate expression of the target genes were screened in the eQTL region. The association between three SNPs and the corresponding target genes was verified in an independent family. Specifically, Marker13973 was identified for heat shock protein (HSP) family A member 9 (HspA9). Ribosomal protein L10a (RPL10A) was detected approximately 2 kb downstream of the distant regulatory SNP. Further, Marker14346-48 and Marker14346-85 were in complete linkage disequilibrium and identified for autophagy-related gene 7 (ATG7). Nuclear respiratory factor 1 (NRF1) was detected approximately 3 kb upstream of the two SNPs. These results suggested regulatory relationships between RPL10A and HSPA9 and between NRF1 and ATG7. Our findings indicate that distant regulatory mutations play an important role in the regulation of gene expression plasticity by altering upstream regulatory factors in response to heat stress. The identified eQTLs provide candidate biomarkers for predicting the persistence of oysters under future climate change scenarios.

Prospecting on Passiflora backcross families: implications for breeding aiming at CABMV resistance

Passion fruit woodiness disease induced by Cowpea aphid-borne mosaic virus (CABMV) is currently the most harmful viral disease affecting passion fruit crops. Given the economic importance of this fruit crop in Brazil, it is essential that passion fruit breeding programs invest in developing resistant and productive cultivars. The goals of this study were to estimate genetic parameters of agronomic and resistance traits via restricted maximum likelihood (REML) and select individuals found to be resistant to CABMV and agronomically superior, in order to provide the basis for further studies on passion fruit breeding, with the aim to develop cultivars resistant to CABMV. The study was conducted in randomized block design, with four replicates. Three full-sib families from the recombination of RC1 parents were evaluated. Based on the severity data, the area below the mean disease progress curve (AUDPCM) was estimated. Five traits relating to fruit yield and quality were also assessed. The components of variance and genetic values were estimated by the REML/best linear unbiased prediction (BLUP) mixed model methodology. A difference in the severity of CABMV symptoms was recorded among the 119 genotypes evaluated. AUDPCM values varied from 750 to 1252.5 for individuals 202 and 161, respectively. Family 1 distinguished itself with the largest number of individuals selected for resistance; for agronomic traits, families 1 and 2 contributed most of the selected genotypes. Genotypes 167, 24, 34, 23 and 202 achieved the lowest AUDPCM values, being selected for most of the agronomic traits; these results indicate the potential of these genotypes to be chosen and recombined for the advancement of generations in breeding programs aimed at developing resistance to CABMV. The strategy of recombining resistant genotypes from RC1 populations appears to be promising for breeding for resistance to CABMV, of which the heritability is complex.

A 50-kb deletion disrupting the RSPO2 gene is associated with tetradysmelia in Holstein Friesian cattle

BackgroundTetradysmelia is a rare genetic disorder that is characterized by an extremely severe reduction of all limb parts distal of the scapula and pelvic girdle. We studied a Holstein Friesian backcross family with 24 offspring, among which six calves displayed autosomal recessive tetradysmelia. In order to identify the genetic basis of the disorder, we genotyped three affected calves, five dams and nine unaffected siblings using a Bovine Illumina 50 k BeadChip and sequenced the whole genome of the sire.ResultsPathological examination of four tetradysmelia cases revealed a uniform and severe dysmelia of all limbs. Applying a homozygosity mapping approach, we identified a homozygous region of 10.54 Mb on chromosome 14 (Bos taurus BTA14). Only calves that were diagnosed with tetradysmelia shared a distinct homozygous haplotype for this region. We sequenced the whole genome of the cases’ sire and searched for heterozygous single nucleotide polymorphisms (SNPs) and small variants on BTA14 that were uniquely present in the sire and absent from 3102 control whole-genome sequences of the 1000 Bull Genomes Project, but none were identified in the 10.54-Mb candidate region on BTA14. Therefore, we subsequently performed a more comprehensive analysis by also considering structural variants and detected a 50-kb deletion in the targeted chromosomal region that was in the heterozygous state in the cases’ sire. Using PCR, we confirmed that this detected deletion segregated perfectly within the family with tetradysmelia. The deletion spanned three exons of the bovine R-spondin 2 (RSPO2) gene, which encode three domains of the respective protein. R-spondin 2 is a secreted ligand of leucine-rich repeats containing G protein-coupled receptors that enhance Wnt signalling and is involved in a broad range of developmental processes during embryogenesis.ConclusionsWe identified a 50-kb deletion on BTA14 that disrupts the coding sequence of the RSPO2 gene and is associated with bovine tetradysmelia. To our knowledge, this is the first reported candidate causal mutation for tetradysmelia in a large animal model. Since signalling pathways involved in limb development are conserved across species, the observed inherited defect may serve as a model to further elucidate fundamental pathways of limb development.

Field Performance of Backcross (BC1) Blueberry Hybrids of Vaccinium padifolium (Section Hemimyrtillus) with V. corymbosum/V. angustifolium

There is ongoing interest in transferring new characteristics into commercial blueberry from other blueberry species. Vaccinium padifolium is a species distantly related to commercial blueberry that has traits of notable value to conventional blueberry development. Among these traits are upright structure, strong growth, abundant flowering and fruiting, superior self-fertility, fruit-cluster structure suited to mechanical harvesting, and repeat/continuous flowering. Previously produced F1 hybrids of V. padifolium × V. corymbosum were used in crosses with a variety of conventional blueberry selections to generate 13 backcross (BC1) families. The backcross families were evaluated under field conditions to determine their performance and to guide further use of this germplasm. The offspring varied considerably, but most families produced several individuals of acceptable commercial quality. The recovery of V. padifolium characteristics varied. Many plants bore evidence of V. padifolium plant structure, but none showed indications thus far of continuous or repeat flowering. The best selections from these families have been saved and will be intermated to generate the next cycle of this material. These clones will also be crossed to hybrid combinations of V. arctostaphylos and V. cylindraceum to further diversify and recombine this germplasm.

Genome-wide association studies for resistance to viral nervous necrosis in three populations of European sea bass (Dicentrarchus labrax) using a novel 57k SNP array DlabChip

Viral Nervous Necrosis (VNN) is a major threat for the European sea bass (Dicentrarchus labrax) aquaculture industry. The improvement of disease resistance through selective breeding is a promising option to reduce outbreaks. With the development of high-throughput genotyping technologies, identification of genomic regions involved in the resistance could improve the efficiency of selective breeding. The aim of this study was to identify quantitative trait loci (QTL) involved in VNN resistance and to quantify their effect.Four experimental backcross families comprising 378, 454, 291 and 211 individuals and two commercial populations A and B comprising 1027 and 1042 individuals obtained from partial factorial crosses (59♂ x 20♀ for pop A; 39♂ x 14♀ for pop B) were submitted to a redspotted grouper nervous necrosis virus (RGNNV) challenge by bath. A high-density single nucleotide polymorphism (SNP) chip panel was designed to develop the ThermoFisher Axiom™ 57k SNP DlabChip, which was used for genotyping all individuals and building a high quality linkage map. In the backcross families, composite interval mapping was performed on 30,917, 23,592, 30,656 and 31,490 markers, respectively. In the commercial populations, 40,263 markers in pop A and 41,166 markers in pop B were used to perform genome-wide association studies (GWAS) using a GBLUP and a BayesCπ approach.One QTL was identified on chromosome LG12 in three of the four experimental backcross families, and one additional QTL on LG8 was detected in only one family. In commercial populations, QTL mapping revealed a total of seven QTLs, among which the previously mentioned QTL on LG12 was detected in both. This QTL, which was mapped to an interval of 3.45 cM, explained 9.21% of the total genetic variance in pop A, while other identified QTLs individually explained less than 1% of the total genetic variance.The identification of QTL regions involved in VNN resistance in European sea bass, with one having a strong effect, should have a great impact on the aquaculture industry. Future work could focus on the fine mapping of the causal mutation present on LG12 using whole genome sequencing.

Segregation of Flower Color and Eyespot in Althea

Althea (Hibiscus syriacus) is an ornamental shrub prized for its winterhardiness, flower colors, and unique flower forms, including single-flowered and double-flowered types. Although floral traits are most important for breeders of althea, little is known about their segregation patterns. The objective of this study was to determine segregation patterns in flower color, including eyespot, among hybrid seedlings of elite taxa. Over 4 years, more than 3100 flowering seedlings were produced for observation of F1, F2, and backcross families. For each plant, data were collected including presence of eyespot and petal body color (CIEL*a*b*) using a colorimeter. Recessive testcrosses and χ2 analyses were performed among three taxa (‘Buddha Belly’, ‘Diana’, and White Chiffon®), and between this recessive group and a suite of colorful taxa. Self-pollination and intercrosses within homozygous dominant and homozygous recessive groups further confirmed their genotypes. Based on these results, we propose that eyespot is controlled by a single gene called spotless, named for the recessive allele that results in a complete elimination of color in flowers. Crosses that resulted in seedlings that all produced eyespots were observed to segregate for color in the petal body. Of these, one group produced white to blush pink petals, which was recessive to full color. Recessive testcrosses and χ2 analyses were performed among nine taxa exhibiting eyespots with white to blush petal bodies, and between taxa with full-color petal bodies. These testcrosses resulted in a putative homozygous dominant group composed mostly of blue and dark pink taxa, whereas the heterozygous group was composed mostly of pink taxa. Spotless taxa were also added to these two groups, suggesting an epistatic interaction with the spotless allele. Based on these results, we propose that petal body color is controlled by a single gene called geisha, named for the recessive allele that produces white to blush-pink petal bodies and dark red eyespot. This trait exhibits incomplete dominance and is under epistatic control by spotless. Geisha-type flowers lack pigment in the petal body, or exhibit a blush pink, likely produced by low levels of cyanidin, peonidin, and pelargonidin. The interaction and segregation of these two genes was confirmed in F1, F2, and backcross families from two crosses: Lil’ Kim™ × Blue Chiffon™ and Fiji™ × White Chiffon®. This study on segregation of flower color in H. syriacus contributes substantial and useful information on inheritance of color and will facilitate targeted breeding to improve this vibrant ornamental shrub.

Genome survey and high-resolution backcross genetic linkage map construction of the ridgetail white prawn Exopalaemon carinicauda applications to QTL mapping of growth traits

BackgroundHigh-resolution genetic linkage map is critical for QTL mapping, genome sequence assembly and marker-assisted selection in aquaculture species. The ridgetail white prawn Exopalaemon carinicauda is one of the most economic shrimp species naturally distributed in the coasts of eastern China and western Korea. However, quite limited genomics and genetics information have been exploited for genetic improvement of economic traits in this species.ResultsIn the present study, we conducted genome survey and constructed high-resolution genetic linkage maps of the ridgetail white prawn with reciprocal-cross mapping family genotyped using next-generation sequencing approaches. The estimated genome size was 9.33 Gb with a heterozygosity of 0.26% and a repeat sequence ratio of 76.62%. 65,772 protein-coding genes were identified by genome annotation. A total of 10,384 SNPs were used to high-throughput genotyping and assigned to 45 linkage groups (LGs) from reciprocal backcross families of E. carinicauda, and the average marker distances were 0.73 cM and 0.55 cM, respectively. Based on the high-resolution linkage map, twenty-three QTLs related to five growth traits were detected. All QTLs could explain 8.8–15.7% of the total growth-traits variation.ConclusionsThe genome size of E. carinicauda was estimated more accurately by genome survey analysis, which revealed basic genomic architecture. The first high-resolution backcross genetic linkage map and QTLs related to growth traits will provide important information for QTL fine mapping, genome assembly and genetic improvement of E. carinicauda and other palaemon shrimps.