An overview of some work on the DNA, genomes, chromosomes and genes in saffron crocus, Crocus sativus, from our lab, with mention of other work with whole genome assemblies from several countries, and analysis of repetitive DNA components by in situ hybridization.

1. The DNA & genomes of saffron
crocus: opportunities for
improvement of sustainability
Pat Heslop-Harrison X/Twitter: @PatHH1
Institute for Environmental Futures,
University of Leicester, UK
phh4@le.ac.uk phh@molcyt.com
Crocus project
Farah Badakshi, Nauf Alsayaid,
Sharaf Eldin, John Bailey
Special thanks to Profs Maria Tsimidou & AR Koocheki
www.Crocusbank.org partners, esp. Prof Jose Antonio Fernandez

2. Biology of Crocus sativus:
its genes and DNA
Where did saffron come from? Where is the crop going?
What are its wild relatives? How was it domesticated?
How much diversity is there?
What are yield, quality, and sustainability constraints?
How can we improve it? (quality, yield, labour, climate change …)

3. Key features of the genome of
Crocus sativus
Genus Crocus (100 species, Iridaceae, monocot)
Triploid with 2n = 3x = 24 chromosomes (3 sets of 8)
Not naturally occurring species. Sterile.
Large genome size – 4,000 Mbp each genome,
Each with c. 30,000 genes
(so three genomes each larger than human)

5. Key features of the genome of
Crocus sativus
Fluorescent in situ
hybridization with
CT/GACA probes
Dispersed repetitive DNA

6. 45S rDNA: 12 sites
C. thomasii genomic probes
highlighting repetitive DNA

7. Groups of
three similar
but not
identical
chromosomes

9. Meiosis in triploid Crocus cultivars
Saffron and the garden Crocus Golden Yellow are both sterile triploids.
In situ hybridization to Golden
Yellow) mitotic metaphases shows
four pairs of chromosomes of
C. flavus origin (2n=8) labelled
red with genomic DNA from C.
flavus, and six chromosomes of
C. angustifolius origin (2n=12)
labelled green with genomic
DNA from C. angustifolius
Golden Yellow, also named Dutch or
Mammoth Yellow;
see Orgaard, HH et al. 1997

10. ‘Golden Yellow’ triploid 2n=3x=14
C. flavus 2n=2x=8 (8 yellow) C. angustifolius 2n=2x=12 (6 green)
Orgaard, Jacobsen & HH
‘Stellaris’ hybrid diploid 2n=2x=10
C. flavus 2n=2x=8 (4 green)
C. angustifolius 2n=2x=12 (6 blue)

12. Metaphase I in triploid Golden Yellow
Four bivalents C. flavus (2n=2x=8) origin
Six univalents from C. angustifolius (2n=2x=12)

13. Origins of Crocus sativus
Hypotheses:
From C. cartwrightianus by autotriploidy
C. cartwrightianus crossed with a related
species involving unreduced gamete
F1 hybrid between 2 species crossed with
3rd species with an unreduced gamete
1 2 3 1 2 3 1 2 3
5 III + 3 II + 3 I
8 III ?
John Bailey

14. SH genomic in
u hybridization
es not
ferentiate
nomes
Crocus sativus

15. Crocus sativus Pachytene
DAPI
Synaptonemal
complex (SC)
spreading
Silver staining

16. Meiotic prophase in triploid Saffron (C. sativus)
ASY-1 antibodies (Franklin and Armstrong) show the progression of lateral element formation and pairing

17. (b) Centromere depolarisation and SC formation during Zygotene
Interphase Leptotene Zygotene Late Zygotene
Telomere
bouquet
Homologue chromosome pairs Centromeres ZYP1
Early Zygotene
1 2 3
Subtelomeric synapsis Interstitial alignment Interstitial elongation
(a) Centromere, telomere and chromosome arm dynamics in meiotic prophase I.
Sepsi et al. Plant Journal 2017

18. Variable IRAP patterns in 11 Crocus species,
including intra-specific variation (19-21 and 22-24)
In this gel, 8 is most similar to saffron (5)
C. sativus C. hadri C. cart
C. pallasii

19. 200 saffron accessions
from 15 countries
150 Crocus spp. 250 accessions
12 countries
Curator: Marcelino de los Mozos (following speaker)

21. Identical patterns in IRAP gel with
diverse Saffron accessions (not 18, of garden-origin)

23. Gene(ATPsynthase) diversity
RED are Saffron Crocus sativus sequences
Three clades, all representing multiple
species

24. g
ATP synthase subunit delta families
Ziwei Wang, Qing Lui, Mathieu Rouard et al.
T2T Ensete glaucum genome assembly
BioRXiv/Gigascience

26. Crocus cartwrightianus
close diploid
Taxonomically in Series Crocus
Genus Crocus, Iridaceae, Monocotyledons
Wild Crocus accessions show high levels of diversity with
several DNA marker systems
Current data points to C. cartwrightianus (2 genomes) and one
of the C. pallasii subspecies as the ancestors of C. sativus but
no very close collection found yet

27. “My saffron is the best in the world”
Relatively unusual with savory & sweet; hot &
cold; water, milk & alcoholic drinks
Saffron:
The taste of luxury, unlocking and elevate flavours
• It is the best! My comment “Use saffron for your local dish”
• Add saffron for regional taste eg India: gravy (‘curry’) and rice; Spain:
paella; Greece: seafood, pilaf, tea/alcohol, sweets; Iran: rice (Tahdig),
tea, sweets

30. By Monsieur Paul Chappellier 1899 reporting work from 1844 onwards
(known to father of partner in Crocusbank, Jean Marie Thiercelin,
seventh generation of saffron and spice company, www.thiercelin1809.com,
“for the Saffron, there is only known a single and unique species;
for ages it has not produced a single variety”, writing that he was
importing bulbs Naples, Athens, Austria, Spain, Kashmir and China.
Advertised for seeds with money prize from 1850 to 1853.
“Peculiarities of structure of the monstrous flowers”
He reports a hybrid with “mania for stigmatisation … teratological phenomeon has
… practical and agricultural side … I try to transform the floral organs into stigmas”
… “so far, these hybrids have had but a botanical and theoretical value, but … it is
reasonable to expect a slight effort to be made in order to arrive at an improved
type … render veritable service to Saffron cultivators”

33. Biology of Crocus sativus
Where did it come from?
What are its wild relatives?
How much diversity is there?
What are constraints?
How can we improve it?
Challenges that are central to our research programme
The PanGenome, genome evolution (structural, copy
number), re-evolution of the species, polyploidy, using
biodiversity …

34. The DNA & genomes of saffron
crocus: opportunities for
improvement of sustainability
Pat Heslop-Harrison X/Twitter: @PatHH1
Institute for Environmental Futures,
University of Leicester, UK
phh4@le.ac.uk phh@molcyt.com
Crocus project
Farah Badakshi, Nauf Alsayaid,
Sharaf Eldin, John Bailey
Special thanks to Profs Maria Tsimidou & AR Koocheki
www.Crocusbank.org partners, esp. Prof Jose Antonio Fernandez

37. Retroelement Markers
Retrotransposon LTR
LTR
Retrotransposon LTR
LTR
Retrotransposon
LTR LTR
Retrotransposon LTR
LTR
Insertion
IRAP – InterRetroelement PCR
Retrotransposon LTR
LTR
Retrotransposon
LTR LTR

38. How does chromosome number change?
How does the genome get so large?
Qing Liu - Avena genome evolution from grass ancestor
• Remarkable and uniform expansion of genome