Three amphioxus reference genomes reveal gene and chromosome evolution of chordates

The slow-evolving invertebrate amphioxus has an irreplaceable role in advancing our understanding into the vertebrate origin and innovations. Here we resolve the nearly complete chromosomal genomes of three amphioxus species, one of which best recapitulates the 17 chordate ancestor linkage groups. We reconstruct the fusions, retention or rearrangements between descendants of whole genome duplications (WGDs), which gave rise to the extant microchromosomes likely existed in the vertebrate ancestor. Similar to vertebrates, the amphioxus genome gradually establishes its 3D chromatin architecture at the onset of zygotic activation, and forms two topologically associated domains at the Hox gene cluster. We find that all three amphioxus species have ZW sex chromosomes with little sequence differentiation, and their putative sex-determining regions are nonhomologous to each other. Our results illuminate the unappreciated interspecific diversity and developmental dynamics of amphioxus genomes, and provide high-quality references for understanding the mechanisms of chordate functional genome evolution.

Development of an SNP Marker Set for Marker-Assisted Backcrossing Using Genotyping-By-Sequencing in Tetraploid Perilla

High-quality molecular markers are essential for marker-assisted selection to accelerate breeding progress. Compared with diploid species, recently diverged polyploid crop species tend to have highly similar homeologous subgenomes, which is expected to limit the development of broadly applicable locus-specific single-nucleotide polymorphism (SNP) assays. Furthermore, it is particularly challenging to make genome-wide marker sets for species that lack a reference genome. Here, we report the development of a genome-wide set of kompetitive allele specific PCR (KASP) markers for marker-assisted recurrent selection (MARS) in the tetraploid minor crop perilla. To find locus-specific SNP markers across the perilla genome, we used genotyping-by-sequencing (GBS) to construct linkage maps of two F2 populations. The two resulting high-resolution linkage maps comprised 2,326 and 2,454 SNP markers that spanned a total genetic distance of 2,133 cM across 16 linkage groups and 2,169 cM across 21 linkage groups, respectively. We then obtained a final genetic map consisting of 22 linkage groups with 1,123 common markers from the two genetic maps. We selected 96 genome-wide markers for MARS and confirmed the accuracy of markers in the two F2 populations using a high-throughput Fluidigm system. We confirmed that 91.8% of the SNP genotyping results from the Fluidigm assay were the same as the results obtained through GBS. These results provide a foundation for marker-assisted backcrossing and the development of new varieties of perilla.

Identification of novel genetic regions associated with resistance to European canker in apple

Resistance to Neonectria ditissima, the fungus causing European canker in apple, was studied in a multiparental population of apple scions using several phenotyping methods. The studied population consists of individuals from multiple families connected through a common pedigree. The degree of disease of each individual in the population was assessed in three experiments: artificial inoculations of detached dormant shoots, potted trees in a glasshouse and in a replicated field experiment. The genetic basis of the differences in disease was studied using a pedigree-based analysis (PBA). Three quantitative trait loci (QTL), on linkage groups (LG) 6, 8 and 10 were identified in more than one of the phenotyping strategies. An additional four QTL, on LG 2, 5, 15 and 16 were only identified in the field experiment. The QTL on LG2 and 16 were further validated in a biparental population. QTL effect sizes were small to moderate with 4.3 to 19 % of variance explained by a single QTL. A subsequent analysis of QTL haplotypes revealed a dynamic response to this disease, in which the estimated effect of a haplotype varied over the field time-points. Two groups of QTL-haplotypes could be distinguished, one that displayed increased effect and one with a constant effect across time-points. These results suggest that there are different modes of control of N. ditissima in the early stages of infection compared to later time-points of disease development. It also shows that multiple QTL will need to be considered to improve resistance to European canker in apple breeding germplasm.

Identification of Candidate Genes Associated With Hypoxia Tolerance in Trachinotus blochii Using Bulked Segregant Analysis and RNA-Seq

Golden Pompano (Trachinotus blochii) has rapidly developed into the one of the main valuable fish species in Chinese marine aquaculture. Due to its rapid growth, active metabolism, and high oxygen consumption, hypoxia will increase its mortality and cause serious economic losses. We constructed two experimental groups of fish with different degrees of tolerance to hypoxia, used BSR-Seq analysis based on genome and genetic linkage groups to locate SNPs and genes that were related to the differences in hypoxia tolerance. The results showed that hypoxia tolerance SNPs of golden pompano may be jointly determined by multiple linkage groups, especially linkage groups 18 and 22. There were 768 and 348 candidate genes located in the candidate regions of the brain and liver, respectively. These genes were mainly involved in anaerobic energy metabolism, stress response, immune response, waste discharge, and cell death. The prostaglandin-endoperoxide synthase 2 (PTGS2) on LG8, which is involved in the metabolism of arachidonic acid, has a G/A nonsynonymous mutation at position 20641628, and the encoded amino acid was changed from hydrophobic aspartic acid to asparaginate. The specific pathway of the RIG-I-like receptor signaling pathway in the liver may mediate the metabolic system and the immune system, linking glucose metabolism with immune regulation. The death of the hypoxia-intolerant group may be due to the accumulation of lactic acid caused by the activation of anaerobic glycolysis during the early stage of hypoxia stress, and the activation of type I interferon was inhibited, which resulted in decreased immunity. Among the genes involved in the RIG-I-like receptor signaling pathway, the CYLD Lysine 63 Deubiquitinase (CYLD) located on LG16 had a G/T nonsynonymous mutation at position 13629651, and the encoded amino acid was changed from alanine acid to valine. The interferon induced with helicase C domain 1 (Ifih1) located on LG18 has a G/C nonsynonymous mutation at position 16153700, and the encoded hydrophilic glycine was changed to hydrophobic alanine. Our findings suggest these SNPs may assist in the molecular breeding of hypoxia-tolerant golden pompano, and speculate that the balance of glucose and lipid metabolism plays a key role in Trachinotus blochii under acute hypoxia.

The genomics of growth and blue spots in a cultured population of Australasian snapper Chrysophrys auratus

<p>Characterizing the genome and understanding how it influences phenotypic variation is a central goal for studies on evolution. The findings of genomic research are applicable to a wide range of human endeavours, including predicting disease risk, supporting selective breeding programmes, and understanding adaptive variation in natural populations. One industry that could particularly benefit from this knowledge is Aquaculture. In recent years, aquaculture production has been increasing to offset the production limits of wild fisheries. Genomics can be used in aquaculture to quantify variation of captive populations, reconstruct pedigrees, and improve the gains from selective breeding programs. The overall goal of this thesis research was to generate a genome-wide genotyping dataset and investigated several key traits for Australasian snapper (Chrysophrys auratus or Pagrus auratus). The findings will be used to establish one of the first genomics-informed New Zealand aquaculture programmes and provide a better understanding of the genotype-phenotype relationships in this teleost species. The first two chapters of this thesis provide a review of the literature and establish the background information and context for the research in subsequent data chapters. A brief overview of genomics, fisheries and aquaculture, and the intersection of these two fields are provided in the Chapter 1. An in-depth quantitative review of 146 Quantitative Trait Loci (QTL) studies in teleost fish was then carried out in Chapter 2. Chapter 3 provides details about the study population and the collection of genotyping data. Genotyping-By-Sequencing (GBS) was used to generate 11K Single Nucleotide Polymorphism (SNP) markers for individuals in the three generation pedigree. Together with phenotypic data the genotyping was used to reconstruct the pedigree, measure inbreeding, and estimate heritability for a range of traits. Parents were identified for 93% of the offspring and successful pedigree reconstruction indicated highly uneven contributions of each parent to the subsequent generations. The average inbreeding level did not change between generations, but significantly different inbreeding levels were observed between offspring from the two founding cohorts and as a result full and half sibling crosses within the group spawning teleost species. Heritability was estimated for a range of traits using both a pedigree relatedness matrix and a genomic relatedness matrix. Chapter 4, uses the genotyping and phenotyping data to generate a linkage map and carry out a scan for quantitative trait loci (QTLs) associated with growth rate. The linkage map reported in this thesis is one of the highest density maps for any Sparidae species at the time of writing. It contained 24 linkage groups, which represent the 24 snapper chromosomes. Growth QTLs were found on three linkage groups and a scan of available genome data identified three candidate growth genes nearby on the linkage groups. Chapter 5, uses the genotyping data and images collected during the study to characterize snappers blue spots and search for QTLs associated with spot numbers. QTLs were found on 12 of the 24 linkage groups, of which one was consistent between two QTL methods applied. A scan of available genome data identified the tyrosinase gene in the middle of the putative QTL region, which is a causal gene for iridophore cell numbers that form blue spots in other fish species. Chapter 6, discuss the implications, future directions, and application of this research to the snapper breeding programme.</p>

The genomics of growth and blue spots in a cultured population of Australasian snapper Chrysophrys auratus

<p>Characterizing the genome and understanding how it influences phenotypic variation is a central goal for studies on evolution. The findings of genomic research are applicable to a wide range of human endeavours, including predicting disease risk, supporting selective breeding programmes, and understanding adaptive variation in natural populations. One industry that could particularly benefit from this knowledge is Aquaculture. In recent years, aquaculture production has been increasing to offset the production limits of wild fisheries. Genomics can be used in aquaculture to quantify variation of captive populations, reconstruct pedigrees, and improve the gains from selective breeding programs. The overall goal of this thesis research was to generate a genome-wide genotyping dataset and investigated several key traits for Australasian snapper (Chrysophrys auratus or Pagrus auratus). The findings will be used to establish one of the first genomics-informed New Zealand aquaculture programmes and provide a better understanding of the genotype-phenotype relationships in this teleost species. The first two chapters of this thesis provide a review of the literature and establish the background information and context for the research in subsequent data chapters. A brief overview of genomics, fisheries and aquaculture, and the intersection of these two fields are provided in the Chapter 1. An in-depth quantitative review of 146 Quantitative Trait Loci (QTL) studies in teleost fish was then carried out in Chapter 2. Chapter 3 provides details about the study population and the collection of genotyping data. Genotyping-By-Sequencing (GBS) was used to generate 11K Single Nucleotide Polymorphism (SNP) markers for individuals in the three generation pedigree. Together with phenotypic data the genotyping was used to reconstruct the pedigree, measure inbreeding, and estimate heritability for a range of traits. Parents were identified for 93% of the offspring and successful pedigree reconstruction indicated highly uneven contributions of each parent to the subsequent generations. The average inbreeding level did not change between generations, but significantly different inbreeding levels were observed between offspring from the two founding cohorts and as a result full and half sibling crosses within the group spawning teleost species. Heritability was estimated for a range of traits using both a pedigree relatedness matrix and a genomic relatedness matrix. Chapter 4, uses the genotyping and phenotyping data to generate a linkage map and carry out a scan for quantitative trait loci (QTLs) associated with growth rate. The linkage map reported in this thesis is one of the highest density maps for any Sparidae species at the time of writing. It contained 24 linkage groups, which represent the 24 snapper chromosomes. Growth QTLs were found on three linkage groups and a scan of available genome data identified three candidate growth genes nearby on the linkage groups. Chapter 5, uses the genotyping data and images collected during the study to characterize snappers blue spots and search for QTLs associated with spot numbers. QTLs were found on 12 of the 24 linkage groups, of which one was consistent between two QTL methods applied. A scan of available genome data identified the tyrosinase gene in the middle of the putative QTL region, which is a causal gene for iridophore cell numbers that form blue spots in other fish species. Chapter 6, discuss the implications, future directions, and application of this research to the snapper breeding programme.</p>

Quantitative Trait Loci and Candidate Genes Associated with Cold-Acclimation and Microdochium nivale Tolerance/Susceptibility in Winter Triticale (x Triticosecale)

Tolerance to pink snow mold caused by Microdochium nivale appears after a cold-hardening period and it is an essential, genotype-dependent, complex quantitative trait for the wintering of triticale (x Triticosecale) and other cereals. Despite long-term studies, a marker for the selection of the tolerant genotypes is still insufficiently recognized. Chlorophyll fluorescence has been reported as a sensitive indicator of stress effects on photosynthesis and can be used to predict plant tolerance. In this study, the genomic regions (QTLs) associated with the level of winter triticale seedlings damage caused by M. nivale infection as well as photosynthesis quantum efficiency and chlorophyll a fluorescence parameters were identified in seedlings of mapping population of 89 doubled haploids lines (DHs) derived from F1 hybrid of cv. ‘Hewo’ and cv. ‘Magnat’ accompanied with the genetic map consisting of 20 linkage groups with a total map length 4997.4 cm. Independent experiments performed in controlled conditions revealed 13 regions identified by a composite interval mapping, located on 7A, 1B, 2B, 6B, 7B, 3R, 5R, and 6R linkage groups and related to the PI, PIABS, TRo/CS, ABS/CS, ABS/CSm, ABS/RC, and Qy values as well as M. nivale tolerance T and susceptibility level P expressed by the seedling damage index. Additionally, candidate genes were in silico identified with the sequence position on wheat (2B and 7B) and rye (5R) chromosomes, where relevant QTL regions were found. The most important candidate genes indicated for M. nivale tolerance of cold-hardened triticale seedlings include those coding: sterol 3-beta-glucosyltransferase UGT80A2-like, transcription factor NAI1-like, and flavonol3-sulfotransferase-like proteins on chromosomes 2B and 5R.

Prediction of the importance of auxiliary traits using computational intelligence and machine learning: A simulation study

The present study evaluated the importance of auxiliary traits of a principal trait based on phenotypic information and previously known genetic structure using computational intelligence and machine learning to develop predictive tools for plant breeding. Data of an F2 population represented by 500 individuals, obtained from a cross between contrasting homozygous parents, were simulated. Phenotypic traits were simulated based on previously established means and heritability estimates (30%, 50%, and 80%); traits were distributed in a genome with 10 linkage groups, considering two alleles per marker. Four different scenarios were considered. For the principal trait, heritability was 50%, and 40 control loci were distributed in five linkage groups. Another phenotypic control trait with the same complexity as the principal trait but without any genetic relationship with it and without pleiotropy or a factorial link between the control loci for both traits was simulated. These traits shared a large number of control loci with the principal trait, but could be distinguished by the differential action of the environment on them, as reflected in heritability estimates (30%, 50%, and 80%). The coefficient of determination were considered to evaluate the proposed methodologies. Multiple regression, computational intelligence, and machine learning were used to predict the importance of the tested traits. Computational intelligence and machine learning were superior in extracting nonlinear information from model inputs and quantifying the relative contributions of phenotypic traits. The R2 values ranged from 44.0% - 83.0% and 79.0% - 94.0%, for computational intelligence and machine learning, respectively. In conclusion, the relative contributions of auxiliary traits in different scenarios in plant breeding programs can be efficiently predicted using computational intelligence and machine learning.

High-resolution genetic map construction and QTL analysis of important fiber traits in kenaf using RAD-seq

Quantitative trait locus (QTL) mapping is a useful method for revealing the mechanism of complex genetic traits and identifying new genomic information to accelerate crop improvement. In the present study, 154 F2:3 strains and their parents were used for restriction site-associated DNA sequencing, single-nucleotide polymorphism (SNP) identification, and genetic map construction. After filtering based on stringent filtering standards, 297.5 Gb of clean data were obtained. Further, 5,191 polymorphic SNP markers were identified from each sample, of which 1,997 polymorphic SNP markers were successfully mapped onto 18 different linkage groups. Six QTLs (QPH, QFBW, QDBW, QFW, QFT, and QFC) were identified based on the genetic map using the multiple QTL mapping (MQM) method, which were then assigned to three linkage groups, LG16, LG8, and LG3. QPH, QFBW, QDBW, and QFW were related to fiber yield, while QFT and QFC were related to fiber quality. This is the first study of its kind to map QTL of fiber yield and fiber quality, which will facilitate further understanding of the molecular genetic basis of these traits. However, there are limitations regarding the utilization of this map because several large gaps remain in some linkage groups. Therefore, additional markers need to be developed to further narrow these regions.

Marker-trait association study for root-related traits in chickpea (Cicer arietinum L.)

<p class="042abstractstekst">Root structure modification can improve important agronomic traits including yield, drought tolerance and nutrient deficiency resistance. The aim of the present study was to investigate the diversity of root traits and to find simple sequence repeat (SSR) markers linked to root traits in chickpea (<em>Cicer arietinum </em>L.). This research was performed using 39 diverse accessions of chickpea. The results showed that there is significant variation in root traits among chickpea genotypes. A total of 26 alleles were detected 26 polymorphic bands were produced by 10 SSR markers in the eight linkage groups (LG). The results indicated that there is substantial variability present in chickpea<strong> </strong>germplasm for root traits.<strong> </strong>By analyzing the population structure, four subpopulations were identified.<strong> </strong>PsAS2, AF016458, 16549 and 19075 SSR markers on LG1, LG3, LG2 and LG1 linkage group respectively were<strong> </strong>associated with root traits<strong>.</strong> The research findings provide valuable information for improving root traits for chickpea breeders.</p>