Publications by authors named "C Robin Buell"

206 Publications

Focus on the biology of plant genomes.

Plant Cell 2021 Feb 2. Epub 2021 Feb 2.

Editor-in-Chief, The Plant Cell, and Donald Danforth Plant Science Center, Olivette, Missouri 63132, USA, and Division of Plant Sciences, University of Missouri, Columbia, Missouri, 65211, USA.

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http://dx.doi.org/10.1093/plcell/koab039DOI Listing
February 2021

Corrigendum to: Genome sequencing of four culinary herbs reveals terpenoid genes underlying chemodiversity in the Nepetoideae.

DNA Res 2020 08;27(4)

Department of Plant Biology, Michigan State University, East Lansing, MI 48824, USA.

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http://dx.doi.org/10.1093/dnares/dsaa025DOI Listing
August 2020

Corrigendum to: Genome sequencing of four culinary herbs reveals terpenoid genes underlying chemodiversity in the Nepetoideae.

DNA Res 2020 08;27(4)

Department of Plant Biology, Michigan State University, East Lansing, MI 48824, USA.

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http://dx.doi.org/10.1093/dnares/dsaa025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7664839PMC
August 2020

Single-parent expression drives dynamic gene expression complementation in maize hybrids.

Plant J 2021 Jan 29;105(1):93-107. Epub 2020 Nov 29.

Department of Agronomy and Plant Genetics, University of Minnesota, Saint Paul, MN, 55108, USA.

Single-parent expression (SPE) is defined as gene expression in only one of the two parents. SPE can arise from differential expression between parental alleles, termed non-presence/absence (non-PAV) SPE, or from the physical absence of a gene in one parent, termed PAV SPE. We used transcriptome data of diverse Zea mays (maize) inbreds and hybrids, including 401 samples from five different tissues, to test for differences between these types of SPE genes. Although commonly observed, SPE is highly genotype and tissue specific. A positive correlation was observed between the genetic distance of the two inbred parents and the number of SPE genes identified. Regulatory analysis showed that PAV SPE and non-PAV SPE genes are mainly regulated by cis effects, with a small fraction under trans regulation. Polymorphic transposable element insertions in promoter sequences contributed to the high level of cis regulation for PAV SPE and non-PAV SPE genes. PAV SPE genes were more frequently expressed in hybrids than non-PAV SPE genes. The expression of parentally silent alleles in hybrids of non-PAV SPE genes was relatively rare but occurred in most hybrids. Non-PAV SPE genes with expression of the silent allele in hybrids are more likely to exhibit above high parent expression level than hybrids that do not express the silent allele, leading to non-additive expression. This study provides a comprehensive understanding of the nature of non-PAV SPE and PAV SPE genes and their roles in gene expression complementation in maize hybrids.
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http://dx.doi.org/10.1111/tpj.15042DOI Listing
January 2021

Root Crown Response to Fungal Root Rot in Middle American × Andean Lines.

Plant Dis 2020 Dec 20;104(12):3135-3142. Epub 2020 Oct 20.

Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI 48824.

Fusarium root rot (FRR) is a global limiter of dry bean ( L.) production. In common bean and other legumes, resistance to FRR is related to both root development and root architecture, providing a breeding strategy for FRR resistance. Here, we describe the relationships between root traits and FRR disease symptoms. Using "shovelomics" techniques, a subset of recombinant inbred lines was phenotyped for root architecture traits and disease symptoms across three Michigan fields, including one field with artificially increased disease pressure. At the early growth stages, stem diameter, basal root number, and distribution of hypocotyl-borne adventitious roots were all significantly related to FRR disease scores. These results demonstrate that root architecture is a component of resistance to FRR in the field at early growth stages (first expanded trifoliate) complementing previous studies that evaluated root traits at later developmental stages (flowering, pod fill, etc.). Correlation matrices of root traits indicate that resistant and susceptible lines have statistically different root systems and show that basal root number is a key feature in resistant root systems while adventitious root distribution is an important feature in susceptible root systems. Based on the results of this study, selection for increased basal root number, increased adventitious root number, and even distribution of adventitious roots in early growth stages (first expanded trifoliate) would positively impact resistance to FRR.
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http://dx.doi.org/10.1094/PDIS-05-20-0956-REDOI Listing
December 2020

Construction of a chromosome-scale long-read reference genome assembly for potato.

Gigascience 2020 Sep;9(9)

Department of Plant Biology, Michigan State University, 612 Wilson Road, East Lansing, MI 48824, USA.

Background: Worldwide, the cultivated potato, Solanum tuberosum L., is the No. 1 vegetable crop and a critical food security crop. The genome sequence of DM1-3 516 R44, a doubled monoploid clone of S. tuberosum Group Phureja, was published in 2011 using a whole-genome shotgun sequencing approach with short-read sequence data. Current advanced sequencing technologies now permit generation of near-complete, high-quality chromosome-scale genome assemblies at minimal cost.

Findings: Here, we present an updated version of the DM1-3 516 R44 genome sequence (v6.1) using Oxford Nanopore Technologies long reads coupled with proximity-by-ligation scaffolding (Hi-C), yielding a chromosome-scale assembly. The new (v6.1) assembly represents 741.6 Mb of sequence (87.8%) of the estimated 844 Mb genome, of which 741.5 Mb is non-gapped with 731.2 Mb anchored to the 12 chromosomes. Use of Oxford Nanopore Technologies full-length complementary DNA sequencing enabled annotation of 32,917 high-confidence protein-coding genes encoding 44,851 gene models that had a significantly improved representation of conserved orthologs compared with the previous annotation. The new assembly has improved contiguity with a 595-fold increase in N50 contig size, 99% reduction in the number of contigs, a 44-fold increase in N50 scaffold size, and an LTR Assembly Index score of 13.56, placing it in the category of reference genome quality. The improved assembly also permitted annotation of the centromeres via alignment to sequencing reads derived from CENH3 nucleosomes.

Conclusions: Access to advanced sequencing technologies and improved software permitted generation of a high-quality, long-read, chromosome-scale assembly and improved annotation dataset for the reference genotype of potato that will facilitate research aimed at improving agronomic traits and understanding genome evolution.
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http://dx.doi.org/10.1093/gigascience/giaa100DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7509475PMC
September 2020

Generation of a chromosome-scale genome assembly of the insect-repellent terpenoid-producing Lamiaceae species, Callicarpa americana.

Gigascience 2020 Sep;9(9)

Department of Plant Biology, Michigan State University, 612 Wilson Road, East Lansing, MI 48824, USA.

Background: Plants exhibit wide chemical diversity due to the production of specialized metabolites that function as pollinator attractants, defensive compounds, and signaling molecules. Lamiaceae (mints) are known for their chemodiversity and have been cultivated for use as culinary herbs, as well as sources of insect repellents, health-promoting compounds, and fragrance.

Findings: We report the chromosome-scale genome assembly of Callicarpa americana L. (American beautyberry), a species within the early-diverging Callicarpoideae clade of Lamiaceae, known for its metallic purple fruits and use as an insect repellent due to its production of terpenoids. Using long-read sequencing and Hi-C scaffolding, we generated a 506.1-Mb assembly spanning 17 pseudomolecules with N50 contig and N50 scaffold sizes of 7.5 and 29.0 Mb, respectively. In all, 32,164 genes were annotated, including 53 candidate terpene synthases and 47 putative clusters of specialized metabolite biosynthetic pathways. Our analyses revealed 3 putative whole-genome duplication events, which, together with local tandem duplications, contributed to gene family expansion of terpene synthases. Kolavenyl diphosphate is a gateway to many of the bioactive terpenoids in C. americana; experimental validation confirmed that CamTPS2 encodes kolavenyl diphosphate synthase. Syntenic analyses with Tectona grandis L. f. (teak), a member of the Tectonoideae clade of Lamiaceae known for exceptionally strong wood resistant to insects, revealed 963 collinear blocks and 21,297 C. americana syntelogs.

Conclusions: Access to the C. americana genome provides a road map for rapid discovery of genes encoding plant-derived agrichemicals and a key resource for understanding the evolution of chemical diversity in Lamiaceae.
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http://dx.doi.org/10.1093/gigascience/giaa093DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7476102PMC
September 2020

Characterization of the Locus Responsible for Floral Anthocyanin Production in Potato.

G3 (Bethesda) 2020 Oct 5;10(10):3871-3879. Epub 2020 Oct 5.

School of Plant and Environmental Sciences, Virginia Tech, Blacksburg VA 24061

Anthocyanins are pigmented secondary metabolites produced via the flavonoid biosynthetic pathway and play important roles in plant stress responses, pollinator attraction, and consumer preference. Using RNA-sequencing analysis of a cross between diploid potato ( L.) lines segregating for flower color, we identified a homolog of the () gene family that encodes a MYB transcription factor, herein termed , as the regulator of anthocyanin production in potato corollas. Transgenic introduction of in white-flowered homozygous doubled-monoploid plants resulted in a recovery of purple flowers. RNA-sequencing revealed the specific anthocyanin biosynthetic genes activated by as well as expression differences in genes within pathways involved in fruit ripening, senescence, and primary metabolism. Closer examination of the locus using genomic sequence analysis revealed a duplication in the locus closely associated with gene expression that is likely attributable to nearby genetic elements. Taken together, this research provides insight into the regulation of anthocyanin biosynthesis in potato while also highlighting how the dynamic nature of the locus may affect expression.
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http://dx.doi.org/10.1534/g3.120.401684DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7534420PMC
October 2020

Genome sequencing of four culinary herbs reveals terpenoid genes underlying chemodiversity in the Nepetoideae.

DNA Res 2020 Jun;27(3)

Department of Plant Biology, Michigan State University, East Lansing, MI 48824, USA.

Species within the mint family, Lamiaceae, are widely used for their culinary, cultural, and medicinal properties due to production of a wide variety of specialized metabolites, especially terpenoids. To further our understanding of genome diversity in the Lamiaceae and to provide a resource for mining biochemical pathways, we generated high-quality genome assemblies of four economically important culinary herbs, namely, sweet basil (Ocimum basilicum L.), sweet marjoram (Origanum majorana L.), oregano (Origanum vulgare L.), and rosemary (Rosmarinus officinalis L.), and characterized their terpenoid diversity through metabolite profiling and genomic analyses. A total 25 monoterpenes and 11 sesquiterpenes were identified in leaf tissue from the 4 species. Genes encoding enzymes responsible for the biosynthesis of precursors for mono- and sesqui-terpene synthases were identified in all four species. Across all 4 species, a total of 235 terpene synthases were identified, ranging from 27 in O. majorana to 137 in the tetraploid O. basilicum. This study provides valuable resources for further investigation of the genetic basis of chemodiversity in these important culinary herbs.
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http://dx.doi.org/10.1093/dnares/dsaa016DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7508350PMC
June 2020

Genome assembly of Chiococca alba uncovers key enzymes involved in the biosynthesis of unusual terpenoids.

DNA Res 2020 Jun;27(3)

Department of Plant Biology.

Chiococca alba (L.) Hitchc. (snowberry), a member of the Rubiaceae, has been used as a folk remedy for a range of health issues including inflammation and rheumatism and produces a wealth of specialized metabolites including terpenes, alkaloids, and flavonoids. We generated a 558 Mb draft genome assembly for snowberry which encodes 28,707 high-confidence genes. Comparative analyses with other angiosperm genomes revealed enrichment in snowberry of lineage-specific genes involved in specialized metabolism. Synteny between snowberry and Coffea canephora Pierre ex A. Froehner (coffee) was evident, including the chromosomal region encoding caffeine biosynthesis in coffee, albeit syntelogs of N-methyltransferase were absent in snowberry. A total of 27 putative terpene synthase genes were identified, including 10 that encode diterpene synthases. Functional validation of a subset of putative terpene synthases revealed that combinations of diterpene synthases yielded access to products of both general and specialized metabolism. Specifically, we identified plausible intermediates in the biosynthesis of merilactone and ribenone, structurally unique antimicrobial diterpene natural products. Access to the C. alba genome will enable additional characterization of biosynthetic pathways responsible for health-promoting compounds in this medicinal species.
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http://dx.doi.org/10.1093/dnares/dsaa013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7433921PMC
June 2020

The evolutionary origins of the cat attractant nepetalactone in catnip.

Sci Adv 2020 May 13;6(20):eaba0721. Epub 2020 May 13.

Department of Natural Product Biosynthesis, Max Planck Institute for Chemical Ecology, D-07745 Jena, Germany.

Catnip or catmint ( spp.) is a flowering plant in the mint family (Lamiaceae) famed for its ability to attract cats. This phenomenon is caused by the compound nepetalactone, a volatile iridoid that also repels insects. Iridoids are present in many Lamiaceae species but were lost in the ancestor of the Nepetoideae, the subfamily containing . Using comparative genomics, ancestral sequence reconstructions, and phylogenetic analyses, we probed the re-emergence of iridoid biosynthesis in . The results of these investigations revealed mechanisms for the loss and subsequent re-evolution of iridoid biosynthesis in the lineage. We present evidence for a chronology of events that led to the formation of nepetalactone biosynthesis and its metabolic gene cluster. This study provides insights into the interplay between enzyme and genome evolution in the origins, loss, and re-emergence of plant chemical diversity.
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http://dx.doi.org/10.1126/sciadv.aba0721DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7220310PMC
May 2020

Multiple QTL Mapping in Autopolyploids: A Random-Effect Model Approach with Application in a Hexaploid Sweetpotato Full-Sib Population.

Genetics 2020 07 5;215(3):579-595. Epub 2020 May 5.

Bioinformatics Research Center, North Carolina State University, Raleigh, North Carolina 27695.

In developing countries, the sweetpotato, (L.) Lam. [Formula: see text], is an important autopolyploid species, both socially and economically. However, quantitative trait loci (QTL) mapping has remained limited due to its genetic complexity. Current fixed-effect models can fit only a single QTL and are generally hard to interpret. Here, we report the use of a random-effect model approach to map multiple QTL based on score statistics in a sweetpotato biparental population ('Beauregard' × 'Tanzania') with 315 full-sibs. Phenotypic data were collected for eight yield component traits in six environments in Peru, and jointly adjusted means were obtained using mixed-effect models. An integrated linkage map consisting of 30,684 markers distributed along 15 linkage groups (LGs) was used to obtain the genotype conditional probabilities of putative QTL at every centiMorgan position. Multiple interval mapping was performed using our R package QTLpoly and detected a total of 13 QTL, ranging from none to four QTL per trait, which explained up to 55% of the total variance. Some regions, such as those on LGs 3 and 15, were consistently detected among root number and yield traits, and provided a basis for candidate gene search. In addition, some QTL were found to affect commercial and noncommercial root traits distinctly. Further best linear unbiased predictions were decomposed into additive allele effects and were used to compute multiple QTL-based breeding values for selection. Together with quantitative genotyping and its appropriate usage in linkage analyses, this QTL mapping methodology will facilitate the use of genomic tools in sweetpotato breeding as well as in other autopolyploids.
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http://dx.doi.org/10.1534/genetics.120.303080DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7337090PMC
July 2020

Gene and genome duplications in the evolution of chemodiversity: perspectives from studies of Lamiaceae.

Curr Opin Plant Biol 2020 06 25;55:74-83. Epub 2020 Apr 25.

Department of Plant Biology, Michigan State University, 612 Wilson Road, East Lansing, MI 48824, USA; Plant Resilience Institute, Michigan State University, 612 Wilson Road, East Lansing, MI 48824, USA; MSU AgBioResearch, Michigan State University, 446 West Circle Drive, East Lansing, MI 48824, USA. Electronic address:

Plants are reservoirs of extreme chemical diversity, yet biosynthetic pathways remain underexplored in the majority of taxa. Access to improved, inexpensive genomic and computational technologies has recently enhanced our understanding of plant specialized metabolism at the biochemical and evolutionary levels including the elucidation of pathways leading to key metabolites. Furthermore, these approaches have provided insights into the mechanisms of chemical evolution, including neofunctionalization and subfunctionalization, structural variation, and modulation of gene expression. The broader utilization of genomic tools across the plant tree of life, and an expansion of genomic resources from multiple accessions within species or populations, will improve our overall understanding of chemodiversity. These data and knowledge will also lead to greater insight into the selective pressures contributing to and maintaining this diversity, which in turn will enable the development of more accurate predictive models of specialized metabolism in plants.
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http://dx.doi.org/10.1016/j.pbi.2020.03.005DOI Listing
June 2020

Quantitative trait loci and differential gene expression analyses reveal the genetic basis for negatively associated β-carotene and starch content in hexaploid sweetpotato [Ipomoea batatas (L.) Lam.].

Theor Appl Genet 2020 Jan 8;133(1):23-36. Epub 2019 Oct 8.

Michigan State University, East Lansing, MI, 48824, USA.

Key Message: β-Carotene content in sweetpotato is associated with the Orange and phytoene synthase genes; due to physical linkage of phytoene synthase with sucrose synthase, β-carotene and starch content are negatively correlated. In populations depending on sweetpotato for food security, starch is an important source of calories, while β-carotene is an important source of provitamin A. The negative association between the two traits contributes to the low nutritional quality of sweetpotato consumed, especially in sub-Saharan Africa. Using a biparental mapping population of 315 F progeny generated from a cross between an orange-fleshed and a non-orange-fleshed sweetpotato variety, we identified two major quantitative trait loci (QTL) on linkage group (LG) three (LG3) and twelve (LG12) affecting starch, β-carotene, and their correlated traits, dry matter and flesh color. Analysis of parental haplotypes indicated that these two regions acted pleiotropically to reduce starch content and increase β-carotene in genotypes carrying the orange-fleshed parental haplotype at the LG3 locus. Phytoene synthase and sucrose synthase, the rate-limiting and linked genes located within the QTL on LG3 involved in the carotenoid and starch biosynthesis, respectively, were differentially expressed in Beauregard versus Tanzania storage roots. The Orange gene, the molecular switch for chromoplast biogenesis, located within the QTL on LG12 while not differentially expressed was expressed in developing roots of the parental genotypes. We conclude that these two QTL regions act together in a cis and trans manner to inhibit starch biosynthesis in amyloplasts and enhance chromoplast biogenesis, carotenoid biosynthesis, and accumulation in orange-fleshed sweetpotato. Understanding the genetic basis of this negative association between starch and β-carotene will inform future sweetpotato breeding strategies targeting sweetpotato for food and nutritional security.
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http://dx.doi.org/10.1007/s00122-019-03437-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6952332PMC
January 2020

Variation and Inheritance of Small RNAs in Maize Inbreds and F1 Hybrids.

Plant Physiol 2020 01 1;182(1):318-331. Epub 2019 Oct 1.

Department of Plant and Microbial Biology, University of Minnesota, St. Paul, Minnesota 55108

Small RNAs (sRNAs) regulate gene expression, play important roles in epigenetic pathways, and are hypothesized to contribute to hybrid vigor in plants. Prior investigations have provided valuable insights into associations between sRNAs and heterosis, often using a single hybrid genotype or tissue, but our understanding of the role of sRNAs and their potential value to plant breeding are limited by an incomplete picture of sRNA variation between diverse genotypes and development stages. Here, we provide a deep exploration of sRNA variation and inheritance among a panel of 108 maize () samples spanning five tissues from eight inbred parents and 12 hybrid genotypes, covering a spectrum of heterotic groups, genetic variation, and levels of heterosis for various traits. We document substantial developmental and genotypic influences on sRNA expression, with varying patterns for 21-nucleotide (nt), 22-nt, and 24-nt sRNAs. We provide a detailed view of the distribution of sRNAs in the maize genome, revealing a complex makeup that also shows developmental plasticity, particularly for 22-nt sRNAs. sRNAs exhibited substantially more variation between inbreds as compared with observed variation for gene expression. In hybrids, we identify locus-specific examples of nonadditive inheritance, mostly characterized as partial or complete dominance, but rarely outside the parental range. However, the global abundance of 21-nt, 22-nt, and 24-nt sRNAs varies very little between inbreds and hybrids, suggesting that hybridization affects sRNA expression principally at specific loci rather than on a global scale. This study provides a valuable resource for understanding the potential role of sRNAs in hybrid vigor.
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http://dx.doi.org/10.1104/pp.19.00817DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6945832PMC
January 2020

Genome-wide Inference of Somatic Translocation Events During Potato Dihaploid Production.

Plant Genome 2019 06;12(2)

Potato ( L.) breeders often use dihaploids, which are 2× progeny derived from 4× autotetraploid parents. Dihaploids can be used in diploid crosses to introduce new genetic material into breeding germplasm that can be integrated into tetraploid breeding through the use of unreduced gametes in 4× by 2× crosses. Dihaploid potatoes are usually produced via pollination by haploid inducer lines known as in vitro pollinators (IVP). In vitro pollinator chromosomes are selectively degraded from initially full hybrid embryos, resulting in 2× seed. During this process, somatic translocation of IVP DNA may occur. In this study, a genome-wide approach was used to identify such events and other chromosome-scale abnormalities in a population of 95 dihaploids derived from a cross between potato cultivar Superior and the haploid inducing line IVP101. Most Superior dihaploids showed translocation rates of <1% at 16,947,718 assayable sites, yet two dihaploids showed translocation rates of 1.86 and 1.60%. Allelic ratios at translocation sites suggested that most translocations occurred in individual cell lineages and were thus not present in all cells of the adult plants. Translocations were enriched in sites associated with high gene expression and H3K4 dimethylation and H4K5 acetylation, suggesting that they tend to occur in regions of open chromatin. The translocations likely result as a consequence of double-stranded break repair in the dihaploid genomes via homologous recombination during which IVP chromosomes are used as templates. Additionally, primary trisomy was observed in eight individuals. As the trisomic chromosomes were derived from Superior, meiotic nondisjunction may be common in potato.
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http://dx.doi.org/10.3835/plantgenome2018.10.0079DOI Listing
June 2019

Multiple Maize Reference Genomes Impact the Identification of Variants by Genome-Wide Association Study in a Diverse Inbred Panel.

Plant Genome 2019 06;12(2)

Use of a single reference genome for genome-wide association studies (GWAS) limits the gene space represented to that of a single accession. This limitation can complicate identification and characterization of genes located within presence-absence variations (PAVs). In this study, we present the draft de novo genome assembly of 'PHJ89', an 'Oh43'-type inbred line of maize ( L.). From three separate reference genome assemblies ('B73', 'PH207', and PHJ89) that represent the predominant germplasm groups of maize, we generated three separate whole-seedling gene expression profiles and single nucleotide polymorphism (SNP) matrices from a panel of 942 diverse inbred lines. We identified 34,447 (B73), 39,672 (PH207), and 37,436 (PHJ89) transcripts that are not present in the respective reference genome assemblies. Genome-wide association studies were conducted in the 942 inbred panel with both the SNP and expression data values to map (SCMV) resistance. Highlighting the impact of alternative reference genomes in gene discovery, the GWAS results for SCMV resistance with expression values as a surrogate measure of PAV resulted in robust detection of the physical location of a known resistance gene when the B73 reference that contains the gene was used, but not the PH207 reference. This study provides the valuable resource of the Oh43-type PHJ89 genome assembly as well as SNP and expression data for 942 individuals generated from three different reference genomes.
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http://dx.doi.org/10.3835/plantgenome2018.09.0069DOI Listing
June 2019

Transcriptomic analysis of sweet potato under dehydration stress identifies candidate genes for drought tolerance.

Plant Direct 2018 Oct 30;2(10):e00092. Epub 2018 Oct 30.

International Potato Center Lima Peru.

Sweet potato ( [L.] Lam.) is an important subsistence crop in Sub-Saharan Africa, yet as for many crops, yield can be severely impacted by drought stress. Understanding the genetic mechanisms that control drought tolerance can facilitate the development of drought-tolerant sweet potato cultivars. Here, we report an expression profiling study using the US-bred cultivar, Beauregard, and a Ugandan landrace, Tanzania, treated with polyethylene glycol (PEG) to simulate drought and sampled at 24 and 48 hr after stress. At each time-point, between 4,000 to 6,000 genes in leaf tissue were differentially expressed in each cultivar. Approximately half of these differentially expressed genes were common between the two cultivars and were enriched for Gene Ontology terms associated with drought response. Three hundred orthologs of drought tolerance genes reported in model species were identified in the reference genome, of which 122 were differentially expressed under at least one experimental condition, constituting a list of drought tolerance candidate genes. A subset of genes was differentially regulated between Beauregard and Tanzania, representing genotype-specific responses to drought stress. The data analyzed and reported here provide a resource for geneticists and breeders toward identifying and utilizing drought tolerance genes in sweet potato.
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http://dx.doi.org/10.1002/pld3.92DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6508841PMC
October 2018

Cold stress induces enhanced chromatin accessibility and bivalent histone modifications H3K4me3 and H3K27me3 of active genes in potato.

Genome Biol 2019 06 17;20(1):123. Epub 2019 Jun 17.

Department of Horticulture, University of Wisconsin-Madison, Madison, WI, 53706, USA.

Background: Cold stress can greatly affect plant growth and development. Plants have developed special systems to respond to and tolerate cold stress. While plant scientists have discovered numerous genes involved in responses to cold stress, few studies have been dedicated to investigation of genome-wide chromatin dynamics induced by cold or other abiotic stresses.

Results: Genomic regions containing active cis-regulatory DNA elements can be identified as DNase I hypersensitive sites (DHSs). We develop high-resolution DHS maps in potato (Solanum tuberosum) using chromatin isolated from tubers stored under room (22 °C) and cold (4 °C) conditions. We find that cold stress induces a large number of DHSs enriched in genic regions which are frequently associated with differential gene expression in response to temperature variation. Surprisingly, active genes show enhanced chromatin accessibility upon cold stress. A large number of active genes in cold-stored tubers are associated with the bivalent H3K4me3-H3K27me3 mark in gene body regions. Interestingly, upregulated genes associated with the bivalent mark are involved in stress response, whereas downregulated genes with the bivalent mark are involved in developmental processes. In addition, we observe that the bivalent mark-associated genes are more accessible than others upon cold stress.

Conclusions: Collectively, our results suggest that cold stress induces enhanced chromatin accessibility and bivalent histone modifications of active genes. We hypothesize that in cold-stored tubers, the bivalent H3K4me3-H3K27me3 mark represents a distinct chromatin environment with greater accessibility, which may facilitate the access of regulatory proteins required for gene upregulation or downregulation in response to cold stress.
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http://dx.doi.org/10.1186/s13059-019-1731-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6580510PMC
June 2019

Overcoming Self-Incompatibility in Diploid Potato Using CRISPR-Cas9.

Front Plant Sci 2019 2;10:376. Epub 2019 Apr 2.

Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI, United States.

Potato breeding can be redirected to a diploid inbred/F1 hybrid variety breeding strategy if self-compatibility can be introduced into diploid germplasm. However, the majority of diploid potato clones ( spp.) possess gametophytic self-incompatibility that is primarily controlled by a single multiallelic locus called the -locus which is composed of tightly linked genes, (-locus RNase) and multiple (-locus F-box proteins), which are expressed in the style and pollen, respectively. Using genes known to function in the Solanaceae gametophytic SI mechanism, we identified alleles with flower-specific expression in two diploid self-incompatible potato lines using genome resequencing data. Consistent with the location of the -locus in potato, we genetically mapped the gene using a segregating population to a region of low recombination within the pericentromere of chromosome 1. To generate self-compatible diploid potato lines, a dual single-guide RNA (sgRNA) strategy was used to target conserved exonic regions of the gene and generate targeted knockouts (KOs) using a Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated protein 9 (Cas9) approach. Self-compatibility was achieved in nine KO T lines which contained bi-allelic and homozygous deletions/insertions in both genotypes, transmitting self compatibility to T progeny. This study demonstrates an efficient approach to achieve stable, consistent self-compatibility through KO for use in diploid potato breeding approaches.
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http://dx.doi.org/10.3389/fpls.2019.00376DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6454193PMC
April 2019

Quantitative Trait Loci for Freezing Tolerance in a Lowland x Upland Switchgrass Population.

Front Plant Sci 2019 29;10:372. Epub 2019 Mar 29.

U.S. Dairy Forage Research Center, United States Department of Agriculture-Agricultural Research Service, Madison, WI, United States.

Low-temperature related abiotic stress is an important factor affecting winter survival in lowland switchgrass when grown in northern latitudes in the United States. A better understanding of the genetic architecture of freezing tolerance in switchgrass will aid the development of lowland switchgrass cultivars with improved winter survival. The objectives of this study were to conduct a freezing tolerance assessment, generate a genetic map using single nucleotide polymorphism (SNP) markers, and identify QTL (quantitative trait loci) associated with freezing tolerance in a lowland × upland switchgrass population. A pseudo-F mapping population was generated from an initial cross between the lowland population Ellsworth and the upland cultivar Summer. The segregating progenies were screened for freezing tolerance in a controlled-environment facility. Two clonal replicates of each genotype were tested at six different treatment temperatures ranging from -15 to -5°C at an interval of 2°C for two time periods. Tiller emergence (days) and tiller number were recorded following the recovery of each genotype with the hypothesis that upland genotype is the source for higher tiller number and early tiller emergence. Survivorship of the pseudo-F population ranged from 89% at -5°C to 5% at -15°C with an average LT of -9.7°C. Genotype had a significant effect on all traits except tiller number at -15°C. A linkage map was constructed from bi-allelic single nucleotide polymorphism markers generated using exome capture sequencing. The final map consisted of 1618 markers and 2626 cM, with an average inter-marker distance of 1.8 cM. Six significant QTL were identified, one each on chromosomes 1K, 5K, 5N, 6K, 6N, and 9K, for the following traits: tiller number, tiller emergence days and LT. A comparative genomics study revealed important freezing tolerance genes/proteins, such as COR47, DREB2B, zinc finger-CCCH, WRKY, GIGANTEA, HSP70, and NRT2, among others that reside within the 1.5 LOD confidence interval of the identified QTL.
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http://dx.doi.org/10.3389/fpls.2019.00372DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6450214PMC
March 2019

Genomic Prediction for Winter Survival of Lowland Switchgrass in the Northern USA.

G3 (Bethesda) 2019 06 5;9(6):1921-1931. Epub 2019 Jun 5.

United States Dairy Forage Research Center, USDA-ARS, Madison, WI 53706.

The lowland ecotype of switchgrass has generated considerable interest because of its higher biomass yield and late flowering characteristics compared to the upland ecotype. However, lowland ecotypes planted in northern latitudes exhibit very low winter survival. Implementation of genomic selection could potentially enhance switchgrass breeding for winter survival by reducing generation time while eliminating the dependence on weather. The objectives of this study were to assess the potential of genomic selection for winter survival in lowland switchgrass by combining multiple populations in the training set and applying the selected model in two independent testing datasets for validation. Marker data were generated using exome capture sequencing. Validation was conducted using (1) indirect indicators of winter adaptation based on geographic and climatic variables of accessions from different source locations and (2) winter survival estimates of the phenotype. The prediction accuracies were significantly higher when the training dataset comprising all populations was used in fivefold cross validation but its application was not useful in the independent validation dataset. Nevertheless, modeling for population heterogeneity improved the prediction accuracy to some extent but the genetic relationship between the training and validation populations was found to be more influential. The predicted winter survival of lowland switchgrass indicated latitudinal and longitudinal variability, with the northeast USA the region for most cold tolerant lowland populations. Our results suggested that GS could provide valuable opportunities for improving winter survival and accelerate the lowland switchgrass breeding programs toward the development of cold tolerant cultivars suitable for northern latitudes.
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http://dx.doi.org/10.1534/g3.119.400094DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6553536PMC
June 2019

Evaluation of Methods to Assess Activity of Engineered Genome-Editing Nucleases in Protoplasts.

Front Plant Sci 2019 8;10:110. Epub 2019 Feb 8.

Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI, United States.

Genome-editing is being implemented in increasing number of plant species using engineered sequence specific nucleases (SSNs) such as Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated systems (CRISPR/Cas9), Transcription activator like effector nucleases (TALENs), and more recently CRISPR/Cas12a. As the tissue culture and regeneration procedures to generate gene-edited events are time consuming, large-scale screening methodologies that rapidly facilitate validation of genome-editing reagents are critical. Plant protoplast cells provide a rapid platform to validate genome-editing reagents. Protoplast transfection with plasmids expressing genome-editing reagents represents an efficient and cost-effective method to screen for activity of genome-editing constructs and resulting targeted mutagenesis. In this study, we compared three existing methods for detection of editing activity, the T7 endonuclease I assay (T7EI), PCR/restriction enzyme (PCR/RE) digestion, and amplicon-sequencing, with an alternative method which involves tagging a double-stranded oligodeoxynucleotide (dsODN) into the SSN-induced double stranded break and detection of on-target activity of gene-editing reagents by PCR and agarose gel electrophoresis. To validate these methods, multiple reagents including TALENs, CRISPR/Cas9 and Cas9 variants, eCas9(1.1) (enhanced specificity) and Cas9-HF1 (high-fidelity1) were engineered for targeted mutagenesis of (), () and their paralogs in potato. While all methods detected editing activity, the PCR detection of dsODN integration provided the most straightforward and easiest method to assess on-target activity of the SSN as well as a method for initial qualitative evaluation of the functionality of genome-editing constructs. Quantitative data on mutagenesis frequencies obtained by amplicon-sequencing of revealed that the mutagenesis frequency of CRISPR/Cas9 reagents is better than TALENs. Context-based choice of method for evaluation of gene-editing reagents in protoplast systems, along with advantages and limitations associated with each method, are discussed.
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http://dx.doi.org/10.3389/fpls.2019.00110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6376315PMC
February 2019

Haplotype-phased genome and evolution of phytonutrient pathways of tetraploid blueberry.

Gigascience 2019 03;8(3)

Department of Horticulture, Michigan State University, 1066 Bogue Street, East Lansing, MI, 48824, USA.

Background: Highbush blueberry (Vaccinium corymbosum) has long been consumed for its unique flavor and composition of health-promoting phytonutrients. However, breeding efforts to improve fruit quality in blueberry have been greatly hampered by the lack of adequate genomic resources and a limited understanding of the underlying genetics encoding key traits. The genome of highbush blueberry has been particularly challenging to assemble due, in large part, to its polyploid nature and genome size.

Findings: Here, we present a chromosome-scale and haplotype-phased genome assembly of the cultivar "Draper," which has the highest antioxidant levels among a diversity panel of 71 cultivars and 13 wild Vaccinium species. We leveraged this genome, combined with gene expression and metabolite data measured across fruit development, to identify candidate genes involved in the biosynthesis of important phytonutrients among other metabolites associated with superior fruit quality. Genome-wide analyses revealed that both polyploidy and tandem gene duplications modified various pathways involved in the biosynthesis of key phytonutrients. Furthermore, gene expression analyses hint at the presence of a spatial-temporal specific dominantly expressed subgenome including during fruit development.

Conclusions: These findings and the reference genome will serve as a valuable resource to guide future genome-enabled breeding of important agronomic traits in highbush blueberry.
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http://dx.doi.org/10.1093/gigascience/giz012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6423372PMC
March 2019

A chromosomal-scale genome assembly of Tectona grandis reveals the importance of tandem gene duplication and enables discovery of genes in natural product biosynthetic pathways.

Gigascience 2019 03;8(3)

Department of Plant Biology, Michigan State University, 612 Wilson Road, East Lansing, MI 48824, USA.

Background: Teak, a member of the Lamiaceae family, produces one of the most expensive hardwoods in the world. High demand coupled with deforestation have caused a decrease in natural teak forests, and future supplies will be reliant on teak plantations. Hence, selection of teak tree varieties for clonal propagation with superior growth performance is of great importance, and access to high-quality genetic and genomic resources can accelerate the selection process by identifying genes underlying desired traits.

Findings: To facilitate teak research and variety improvement, we generated a highly contiguous, chromosomal-scale genome assembly using high-coverage Pacific Biosciences long reads coupled with high-throughput chromatin conformation capture. Of the 18 teak chromosomes, we generated 17 near-complete pseudomolecules with one chromosome present as two chromosome arm scaffolds. Genome annotation yielded 31,168 genes encoding 46,826 gene models, of which, 39,930 and 41,155 had Pfam domain and expression evidence, respectively. We identified 14 clusters of tandem-duplicated terpene synthases (TPSs), genes central to the biosynthesis of terpenes, which are involved in plant defense and pollinator attraction. Transcriptome analysis revealed 10 TPSs highly expressed in woody tissues, of which, 8 were in tandem, revealing the importance of resolving tandemly duplicated genes and the quality of the assembly and annotation. We also validated the enzymatic activity of four TPSs to demonstrate the function of key TPSs.

Conclusions: In summary, this high-quality chromosomal-scale assembly and functional annotation of the teak genome will facilitate the discovery of candidate genes related to traits critical for sustainable production of teak and for anti-insecticidal natural products.
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http://dx.doi.org/10.1093/gigascience/giz005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6394206PMC
March 2019

Genome Editing in Potato with CRISPR/Cas9.

Methods Mol Biol 2019 ;1917:183-201

Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI, USA.

Cultivated potato, Solanum tuberosum Group Tuberosum L. (2n = 4x = 48) is a heterozygous tetraploid crop that is clonally propagated, thereby resulting in identical genotypes. Due to the lack of sexual reproduction and its concomitant segregation of alleles, genetic engineering is an efficient way of introducing crop improvement traits in potato. In recent years, genome-editing via the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9 system for targeted genome modifications has emerged as the most powerful method due to the ease in designing and construction of gene-specific single guide RNA (sgRNA) vectors. These sgRNA vectors are easily reprogrammable to direct Streptococcus pyogenes Cas9 (SpCas9) to generate double stranded breaks (DSBs) in the target genomes that are then repaired by the cell via the error-prone non-homologous end-joining (NHEJ) pathway or by precise homologous recombination (HR) pathway. CRISPR/Cas9 technology has been successfully implemented in potato for targeted mutagenesis to generate knockout mutations (by means of NHEJ) as well as gene targeting to edit an endogenous gene (by HR). In this chapter, we describe procedures for designing sgRNAs, protocols to clone sgRNAs for CRISPR/Cas9 constructs to generate knockouts, design of donor repair templates and use geminivirus replicons (GVRs) to facilitate gene-editing by HR in potato. We also describe tissue culture procedures in potato for Agrobacterium-mediated transformation to generate gene-edited events along with their molecular characterization.
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http://dx.doi.org/10.1007/978-1-4939-8991-1_14DOI Listing
June 2019

An updated gene atlas for maize reveals organ-specific and stress-induced genes.

Plant J 2019 03 22;97(6):1154-1167. Epub 2019 Jan 22.

Department of Plant Biology, Michigan State University, East Lansing, MI, 48824, USA.

Maize (Zea mays L.), a model species for genetic studies, is one of the two most important crop species worldwide. The genome sequence of the reference genotype, B73, representative of the stiff stalk heterotic group was recently updated (AGPv4) using long-read sequencing and optical mapping technology. To facilitate the use of AGPv4 and to enable functional genomic studies and association of genotype with phenotype, we determined expression abundances for replicated mRNA-sequencing datasets from 79 tissues and five abiotic/biotic stress treatments revealing 36 207 expressed genes. Characterization of the B73 transcriptome across six organs revealed 4154 organ-specific and 7704 differentially expressed (DE) genes following stress treatment. Gene co-expression network analyses revealed 12 modules associated with distinct biological processes containing 13 590 genes providing a resource for further association of gene function based on co-expression patterns. Presence-absence variants (PAVs) previously identified using whole genome resequencing data from 61 additional inbred lines were enriched in organ-specific and stress-induced DE genes suggesting that PAVs may function in phenological variation and adaptation to environment. Relative to core genes conserved across the 62 profiled inbreds, PAVs have lower expression abundances which are correlated with their frequency of dispersion across inbreds and on average have significantly fewer co-expression network connections suggesting that a subset of PAVs may be on an evolutionary path to pseudogenization. To facilitate use by the community, we developed the Maize Genomics Resource website (maize.plantbiology.msu.edu) for viewing and data-mining these resources and deployed two new views on the maize electronic Fluorescent Pictograph Browser (bar.utoronto.ca/efp_maize).
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http://dx.doi.org/10.1111/tpj.14184DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6850026PMC
March 2019

Candidate Variants for Additive and Interactive Effects on Bioenergy Traits in Switchgrass ( L.) Identified by Genome-Wide Association Analyses.

Plant Genome 2018 11;11(3)

Switchgrass ( L.) is a promising herbaceous energy crop, but further gains in biomass yield and quality must be achieved to enable a viable bioenergy industry. Developing DNA markers can contribute to such progress, but depiction of genetic bases should be reliable, involving simple additive marker effects and also interactions with genetic backgrounds (e.g., ecotypes) or synergies with other markers. We analyzed plant height, C content, N content, and mineral concentration in a diverse panel consisting of 512 genotypes of upland and lowland ecotypes. We performed association analyses based on exome capture sequencing and tested 439,170 markers for marginal effects, 83,290 markers for marker × ecotype interactions, and up to 311,445 marker pairs for pairwise interactions. Analyses of pairwise interactions focused on subsets of marker pairs preselected on the basis of marginal marker effects, gene ontology annotation, and pairwise marker associations. Our tests identified 12 significant effects. Homology and gene expression information corroborated seven effects and indicated plausible causal pathways: flowering time and lignin synthesis for plant height; plant growth and senescence for C content and mineral concentration. Four pairwise interactions were detected, including three interactions preselected on the basis of pairwise marker correlations. Furthermore, a marker × ecotype interaction and a pairwise interaction were confirmed in an independent switchgrass panel. Our analyses identified reliable candidate variants for important bioenergy traits. Moreover, they exemplified the importance of interactive effects for depicting genetic bases and illustrated the usefulness of preselecting marker pairs for identifying pairwise marker interactions in association studies.
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http://dx.doi.org/10.3835/plantgenome2018.01.0002DOI Listing
November 2018

Genome Editing for Crop Improvement - Applications in Clonally Propagated Polyploids With a Focus on Potato ( L.).

Front Plant Sci 2018 13;9:1607. Epub 2018 Nov 13.

Department of Plant, Soil and Microbial Sciences, Michigan State University, East Lansing, MI, United States.

Genome-editing has revolutionized biology. When coupled with a recently streamlined regulatory process by the U.S. Department of Agriculture and the potential to generate transgene-free varieties, genome-editing provides a new avenue for crop improvement. For heterozygous, polyploid and vegetatively propagated crops such as cultivated potato, Group Tuberosum L., genome-editing presents tremendous opportunities for trait improvement. In potato, traits such as improved resistance to cold-induced sweetening, processing efficiency, herbicide tolerance, modified starch quality and self-incompatibility have been targeted utilizing CRISPR/Cas9 and TALEN reagents in diploid and tetraploid clones. However, limited progress has been made in other such crops including sweetpotato, strawberry, grapes, citrus, banana etc., In this review we summarize the developments in genome-editing platforms, delivery mechanisms applicable to plants and then discuss the recent developments in regulation of genome-edited crops in the United States and The European Union. Next, we provide insight into the challenges of genome-editing in clonally propagated polyploid crops, their current status for trait improvement with future prospects focused on potato, a global food security crop.
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http://dx.doi.org/10.3389/fpls.2018.01607DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6243044PMC
November 2018

A common genetic mechanism underlies morphological diversity in fruits and other plant organs.

Nat Commun 2018 11 9;9(1):4734. Epub 2018 Nov 9.

Department of Horticulture and Crop Science, The Ohio State University, 1680 Madison Ave, Wooster, OH, 44691, USA.

Shapes of edible plant organs vary dramatically among and within crop plants. To explain and ultimately employ this variation towards crop improvement, we determined the genetic, molecular and cellular bases of fruit shape diversity in tomato. Through positional cloning, protein interaction studies, and genome editing, we report that OVATE Family Proteins and TONNEAU1 Recruiting Motif proteins regulate cell division patterns in ovary development to alter final fruit shape. The physical interactions between the members of these two families are necessary for dynamic relocalization of the protein complexes to different cellular compartments when expressed in tobacco leaf cells. Together with data from other domesticated crops and model plant species, the protein interaction studies provide possible mechanistic insights into the regulation of morphological variation in plants and a framework that may apply to organ growth in all plant species.
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http://dx.doi.org/10.1038/s41467-018-07216-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6226536PMC
November 2018