Spring flowering habit in field pennycress (Thlaspi arvense) has arisen multiple independent times

Abstract

Field pennycress (Thlaspi arvense L.) is currently being developed as a new cold-tolerant oilseed crop. In natural populations, pennycress, like many Brassicaceae relatives, can exhibit either a winter or spring annual phenotype. Pennycress is a diploid relative of Arabidopsis thaliana, a model species that has been used to study many adaptive phenotypes, including flowering time and developmental timing. In Arabidopsis and other Brassicaceae species, mutations in negative regulators of flowering, including FLOWERING LOCUS C and FRIGIDA can cause the transition to a spring annual habit. The genetics underlying the difference between spring and winter annual pennycress lines are currently unknown. Here, we report the identification of four natural alleles of FLC in pennycress that confer a spring annual growth habit identified through whole genome sequencing, cosegregation analyses, and comparative genomics. The global distribution of these spring annual alleles of FLC suggests that the spring annual growth habit has arisen on several independent occasions. The two spring annual FLC alleles present in European accessions were only identified in North American accessions collected in southern Montana, which indicates accessions harboring these two alleles were introduced to North America, likely after pennycress became a widespread species on the continent. These findings provide new information on the natural history of the introduction and spread of spring annual pennycress accessions from Europe into North America. At the molecular level, these findings are important for the ongoing development of pennycress as a winter annual crop. An enhanced understanding of the regulation of flowering in this species should allow for the fine-tuning of flowering in commercial varieties.

Keywords: Thlaspi arvense; flowering locus C; pennycress; spring annual; whole genome sequencing; winter annual.

Figure 1

Segregation analysis confirms that flc‐A allele confers spring annual phenotype. (a) Cogrown MN108SA and MN111 pennycress plants. Insert shows immediate internode elongation in the MN108SA spring annual plant. An F2 population was derived from a cross between MN108SA × MN111 and the segregation ratio of spring to winter annual phenotypes was determined. Whole genome sequencing of the parent plants from each background was also conducted. (b) Gene model of pennycress FLOWERING LOCUS C (FLC). Gray arrow below the first exon indicates the position of the amplicon used to genotype the SNP shown in panel C. (c) Nucleotide and predicted peptide sequence of the 5′ end of the pennycress FLC gene in winter annual MN106, winter annual MN111, and the spring annual accession MN108SA. The flc‐A mutation in MN108SA is highlighted in yellow

Figure 2

Identification of flc‐B and flc‐C alleles of FLC. (a) Expanded gene model of pennycress FLC region, including the upstream TaUFC gene model. Locations of the flc‐B and flc‐C alleles are shown below the gene model. (b) Agarose gel of the diagnostic polymerase chain reaction (PCR) results testing for the flc‐B allele across the shown pennycress accessions. The wild type MN106 allele generates a 2,088 bp amplicon, where as the flc‐B allele produces a 1,632 bp amplicon (Supporting Information Figure S1). (c) Agarose gel of the diagnostic polymerase chain reaction (PCR) results testing for the flc‐C allele across the shown pennycress accessions. Amplification in the top row confirms the presence of the deletion, whereas amplification in the second row confirms absence of the deletion. The sequences of primers used are shown in Supporting Information Table S2, and primer locations and amplicon strategy is shown in Supporting Information Figure S2. Results for accession PI650286 (Groitzsch, Saxony, Germany) are not shown, but was confirmed to have the flc‐C allele. (d) Nucleotide and predicted peptide sequence of FLC in MN106 and PI633414 (Wachstedt, Germany), with the flc‐D allele highlighted in yellow. (e) Global distribution of the four spring annual alleles of FLC in pennycress accessions tested in this study (listed in Supporting Information Table S3)