Genetic Mapping of Millions of SNPs in Safflower (Carthamus tinctorius L.) via Whole-Genome Resequencing.

Accurate assembly of complete genomes is facilitated by very high density genetic maps. We performed low-coverage, whole-genome shotgun sequencing on 96 F6 recombinant inbred lines (RILs) of a cross between safflower (Carthamus tinctorius L.) and its wild progenitor (C. palaestinus Eig). We also produced a draft genome assembly of C. tinctorius covering 866 million bp (∼two-thirds) of the expected 1.35 Gbp genome after sequencing a single, short insert library to ∼21 × depth. Sequence reads from the RILs were mapped to this genome assembly to facilitate SNP identification, and the resulting polymorphisms were used to construct a genetic map. The resulting map included 2,008,196 genetically located SNPs in 1178 unique positions. A total of 57,270 scaffolds, each containing five or more mapped SNPs, were anchored to the map. This resulted in the assignment of sequence covering 14% of the expected genome length to a genetic position. Comparison of this safflower map to genetic maps of sunflower and lettuce revealed numerous chromosomal rearrangements, and the resulting patterns were consistent with a whole-genome duplication event in the lineage leading to sunflower. This sequence-based genetic map provides a powerful tool for the assembly of a low-cost draft genome of safflower, and the same general approach is expected to work for other species.

Gene Networks in Plant Biology: Approaches in Reconstruction and Analysis.

Even though vast amounts of genome-wide gene expression data have become available in plants, it remains a challenge to effectively mine this information for the discovery of genes and gene networks, for instance those that control agronomically important traits. These networks reflect potential interactions among genes and, therefore, can lead to a systematic understanding of the molecular mechanisms underlying targeted biological processes. We discuss methods to analyze gene networks using gene expression data, specifically focusing on four common statistical approaches used to reconstruct networks: correlation, feature selection in supervised learning, probabilistic graphical model, and meta-prediction. In addition, we discuss the effective use of these methods for acquiring an in-depth understanding of biological systems in plants.

Single Nucleotide Polymorphism Identification in Polyploids: A Review, Example, and Recommendations.

Understanding the relationship between genotype and phenotype is a major biological question and being able to predict phenotypes based on molecular genotypes is integral to molecular breeding. Whole-genome duplications have shaped the history of all flowering plants and present challenges to elucidating the relationship between genotype and phenotype, especially in neopolyploid species. Although single nucleotide polymorphisms (SNPs) have become popular tools for genetic mapping, discovery and application of SNPs in polyploids has been difficult. Here, we summarize common experimental approaches to SNP calling, highlighting recent polyploid successes. To examine the impact of software choice on these analyses, we called SNPs among five peanut genotypes using different alignment programs (BWA-mem and Bowtie 2) and variant callers (SAMtools, GATK, and Freebayes). Alignments produced by Bowtie 2 and BWA-mem and analyzed in SAMtools shared 24.5% concordant SNPs, and SAMtools, GATK, and Freebayes shared 1.4% concordant SNPs. A subsequent analysis of simulated Brassica napus chromosome 1A and 1C genotypes demonstrated that, of the three software programs, SAMtools performed with the highest sensitivity and specificity on Bowtie 2 alignments. These results, however, are likely to vary among species, and we therefore propose a series of best practices for SNP calling in polyploids.

Genetic diversity in Carthamus tinctorius (Asteraceae; safflower), an underutilized oilseed crop.

UNLABELLED: • PREMISE OF THE STUDY: Underutilized crops are potentially valuable resources for meeting increasing food demands. Safflower, an oilseed crop, is an example of one such underutilized crop that thrives in moisture-limited areas. Characterization of the genetic diversity maintained within the gene pools of underutilized crops such as safflower is an important step in their further development.• METHODS: A total of 190 safflower individuals, including 134 USDA accessions, 48 breeding lines from two private North American safflower breeding companies, and eight wild safflower individuals, were genotyped using 133 single nucleotide polymorphism (SNP) markers. We then used the resulting data to assess the amount and distribution of genetic diversity within and among these collections of safflower.• KEY RESULTS: Although just a modest reduction in gene diversity was observed in the commercial breeding lines (relative to the other safflower groupings), safflower domestication was accompanied by a significant decrease in allelic richness. Further, our results suggest that most safflower breeding lines originated from a single pool of diversity within the Old World safflower germplasm.• CONCLUSIONS: Taken together, our results suggest that both the safflower germplasm collection and related, wild species harbor previously undocumented genetic diversity that could help fuel future improvement efforts. Paired with analyses of functional diversity, the molecular resources described herein will be thus be useful in the continued development of safflower as an oilseed crop.

Genetic analysis of safflower domestication.

BACKGROUND: Safflower (Carthamus tinctorius L.) is an oilseed crop in the Compositae (a.k.a. Asteraceae) that is valued for its oils rich in unsaturated fatty acids. Here, we present an analysis of the genetic architecture of safflower domestication and compare our findings to those from sunflower (Helianthus annuus L.), an independently domesticated oilseed crop within the same family.We mapped quantitative trait loci (QTL) underlying 24 domestication-related traits in progeny from a cross between safflower and its wild progenitor, Carthamus palaestinus Eig. Also, we compared QTL positions in safflower against those that have been previously identified in cultivated x wild sunflower crosses to identify instances of colocalization. RESULTS: We mapped 61 QTL, the vast majority of which (59) exhibited minor or moderate phenotypic effects. The two large-effect QTL corresponded to one each for flower color and leaf spininess. A total of 14 safflower QTL colocalized with previously reported sunflower QTL for the same traits. Of these, QTL for three traits (days to flower, achene length, and number of selfed seed) had cultivar alleles that conferred effects in the same direction in both species. CONCLUSIONS: As has been observed in sunflower, and unlike many other crops, our results suggest that the genetics of safflower domestication is quite complex. Moreover, our comparative mapping results indicate that safflower and sunflower exhibit numerous instances of QTL colocalization, suggesting that parallel trait transitions during domestication may have been driven, at least in part, by parallel genotypic evolution at some of the same underlying genes.

Mitochondrial heteroplasmy and paternal leakage in natural populations of Silene vulgaris, a gynodioecious plant.

It is currently thought that most angiosperms transmit their mitochondrial genomes maternally. Maternal transmission limits opportunities for genetic heterogeneity (heteroplasmy) of the mitochondrial genome within individuals. Recent studies of the gynodioecious species Silene vulgaris and Silene acaulis, however, document both direct and indirect evidence of mitochondrial heteroplasmy, suggesting that the mitochondrial genome is at times transmitted via paternal leakage. This heteroplasmy allows the generation of multi-locus recombinants, as documented in recent studies of both species. A prior study that employed quantitative PCR (q-PCR) on a limited sample provided direct evidence of heteroplasmy in the mitochondrial gene atp1 in S. vulgaris. Here, we apply the q-PCR methods to a much larger sample and extend them to incorporate the study of an additional atp1 haplotype along with two other haplotypes of the mitochondrial gene cox1 to evaluate the origin, extent, and transmission of mitochondrial genome heteroplasmy in S. vulgaris. We first calibrate our q-PCR methods experimentally and then use them to quantify heteroplasmy in 408 S. vulgaris individuals sampled from 22 natural populations located in Virginia, New York, and Tennessee. Sixty-one individuals exhibit heteroplasmy, including five that exhibited the joint heteroplasmy at both loci that is a prerequisite for effective recombination. The heteroplasmic individuals were distributed among 18 of the populations studied, demonstrating that heteroplasmy is a widespread phenomenon in this species. Further, we compare mother and offspring from 71 families to determine the rate of heteroplasmy gained and lost via paternal leakage and vegetative sorting across generations. Of 17 sibships exhibiting cox1 heteroplasmy and 14 sibships exhibiting atp1 heteroplasmy, more than half of the observations of heteroplasmy are generated via paternal leakage at the time of fertilization, with the rest being inherited from a heteroplasmic mother. Moreover, we show that the average paternal contribution during paternal leakage is about 12%. These findings are surprising, given that the current understanding of gynodioecy assumes that mitochondrial cytoplasmic male sterility elements are strictly maternally inherited. Knowledge of the dynamics of mitochondrial populations within individuals plays an important role in understanding the evolution of gynodioecy, and we discuss our findings within this context.