Tragopogon (Asteraceae) is an excellent natural system for studies of recent polyploidy. Development of an efficient CRISPR/Cas9-based genome editing platform in Tragopogon will facilitate novel studies of the genetic consequences of polyploidy. Here, we report our initial results of developing CRISPR/Cas9 in Tragopogon. We have established a feasible tissue culture and transformation protocol for Tragopogon. Through protoplast transient assays, use of the TragCRISPR system (i.e. the CRISPR/Cas9 system adapted for Tragopogon) was capable of introducing site-specific mutations in Tragopogon protoplasts. Agrobacterium-mediated transformation with Cas9-sgRNA constructs targeting the phytoene desaturase gene (TraPDS) was implemented in this model polyploid system. Sequencing of PCR amplicons from the target regions indicated simultaneous mutations of two alleles and four alleles of TraPDS in albino shoots from Tragopogon porrifolius (2x) and Tragopogon mirus (4x), respectively. The average proportions of successfully transformed calli with the albino phenotype were 87% and 78% in the diploid and polyploid, respectively. This appears to be the first demonstration of CRISPR/Cas9-based genome editing in any naturally formed neopolyploid system. Although a more efficient tissue culture system should be developed in Tragopogon, application of a robust CRISPR/Cas9 system will permit unique studies of biased fractionation, the gene-balance hypothesis and cytonuclear interactions in polyploids. In addition, the CRISPR/Cas9 platform enables investigations of those genes involved in phenotypic changes in polyploids and will also facilitate novel functional biology studies in Asteraceae. Our workflow provides a guide for applying CRISPR/Cas9 to other nongenetic model plant systems.

Although major advances have been made during the past 20 years in our understanding of the genetic and genomic consequences of polyploidy, our knowledge of polyploidy and the proteome is in its infancy. One of our goals is to stimulate additional study, particularly broad-scale proteomic analyses of polyploids and their progenitors. Although it may be too early to generalize regarding the extent to which transcriptomic data are predictive of the proteome of polyploids, it is clear that the proteome does not always reflect the transcriptome. Despite limited data, important observations on the proteomes of polyploids are emerging. In some cases, proteomic profiles show qualitatively and/or quantitatively non-additive patterns, and proteomic novelty has been observed. Allopolyploids generally combine the parental contributions, but there is evidence of parental dominance of one contributing genome in some allopolyploids. Autopolyploids are typically qualitatively identical to but quantitatively different from their parents. There is also evidence of parental legacy at the proteomic level. Proteomes clearly provide insights into the consequences of genomic merger and doubling beyond what is obtained from genomic and/or transcriptomic data. Translating proteomic changes in polyploids to differences in morphology and physiology remains the holy grail of polyploidy--this daunting task of linking genotype to proteome to phenotype should emerge as a focus of polyploidy research in the next decade. This article is part of a Special Issue entitled: Plant Proteomics--a bridge between fundamental processes and crop production, edited by Dr. Hans-Peter Mock.