Genetic variation conflicts with morphological boundaries in Iberian L. species Jordan Bilsborrow | John David | Kálmán Könyves | Alastair Culham

Introduction Results

Daffodils are a common garden , and therefore • There is no separation into the key species of the N. minor group economically important to horticulture. Accurate identification using matk, however there is geographical structure (fig. 2a). and classification of garden relies upon an accurate • Microsatellite data reveals low levels of diversity (eMLG=9.99; . However, the taxonomy of Narcissus is largely H =0.33; H =0.45), and interbreeding amongst populations unresolved. Classifications using morphology and cytology have o e (F =0.29). resulted in 16 to 150 recognised taxa. Recent molecular studies1 st conflict with the morphological boundaries. Prevalent natural • There is admixture between samples which fall into clusters 2 and hybridisation further compounds the complexity of the genus2. 4 (fig. 2b), with no separation of the morphologically distinct N. cyclamineus. The closely related N. jacetanus and N. provincialis, fall This project focusses on the Narcissus minor group (fig. 1), part of into clusters 1 and 3 respectively. sect. Pseudonarcissus, to reveal natural levels of variation and

population structure across the northern Iberian Peninsula.

N. N. cyclamineus N. jacetanus N. asturiensis N.

minor

L.

Fern. Casas Fern.

( Jord

DC. a .) Pugsley.)

Figure 1 Four key taxa belonging to the Narcissus minor group.

Aims:

1. To explore the natural variation of the Narcissus minor group b (sect. Pseudonarcissus); using DNA sequencing and microsatellites. Figure 2 (a) Geographic distribution of matK haplotypes for Narcissus minor group taxa. (b) Geographic distribution of DAPC clusters. 2. To utilise NGS for chloroplast genomics and future marker development.

Methods

605 samples were collected from 53 populations, under permit, across the distribution range. DNA was extracted from silica stored leaf material using a modified CTAB protocol.

DNA sequencing: Figure 3 Draft chloroplast genome of • Chloroplast DNA regions (matK, ndhF, ycf1) were sequenced for Conclusions Narcissus poeticus. either all samples, or representatives of each population. The results reveal the genetic complexity in section Pseudonarcissus. • TCS haplotype networks were constructed from the sequence DNA sequencing and microsatellite data reveal conflicting results, dataset. which in turn conflicts with morphological taxonomic boundaries, also 3 DNA fingerprinting: found in Scilla . Whole chloroplast genomes (fig. 3), produced in this project will be key to mining a robust marker set, for a comprehensive • Eleven microsatellite loci were amplified for all samples. reclassification in the future. Robust markers from nuclear and cpDNA • PCR products were sized against Liz500 ladder using capillary genomes can be used to understand prevalent hybridisation. electrophoresis (ABI 3730cl DNA analyzer). • Bin ranges for the amplified loci were scored in Geneious 11.0.3 Contact information microsatellite plugin. • RNG herbarium, School of Biological Sciences, Whiteknights, Reading, RG6 6AS. • Email: [email protected] | @j_Bilsborrow • Genetic diversity estimates were analysed in R, package ‘poppr’,

and DAPC analysis undertaken in ‘adegenet’. STRUCTURE was References 1. Marques, I. et al. (2017) Taxon 66 (4): 832-854; 2. Blanchard, J. (1990) Narcissus a guide to wild daffodils; 3. Speta, F. (1979) Webbia 34: 419-431 used alongside DAPC to assess support.