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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
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 …)
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)
Key features of the genome of
Crocus sativus
Fluorescent in situ
hybridization with
CT/GACA probes
Dispersed repetitive DNA
45S rDNA: 12 sites
C. thomasii genomic probes
highlighting repetitive DNA
Groups of
three similar
but not
identical
chromosomes
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
‘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)
Metaphase I in triploid Golden Yellow
Four bivalents C. flavus (2n=2x=8) origin
Six univalents from C. angustifolius (2n=2x=12)
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
SH genomic in
u hybridization
es not
ferentiate
nomes
Crocus sativus
Crocus sativus Pachytene
DAPI
Synaptonemal
complex (SC)
spreading
Silver staining
Meiotic prophase in triploid Saffron (C. sativus)
ASY-1 antibodies (Franklin and Armstrong) show the progression of lateral element formation and pairing
(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
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
200 saffron accessions
from 15 countries
150 Crocus spp. 250 accessions
12 countries
Curator: Marcelino de los Mozos (following speaker)
Identical patterns in IRAP gel with
diverse Saffron accessions (not 18, of garden-origin)
Gene(ATPsynthase) diversity
RED are Saffron Crocus sativus sequences
Three clades, all representing multiple
species
g
ATP synthase subunit delta families
Ziwei Wang, Qing Lui, Mathieu Rouard et al.
T2T Ensete glaucum genome assembly
BioRXiv/Gigascience
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
“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
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”
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 …
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
Retroelement Markers
Retrotransposon LTR
LTR
Retrotransposon LTR
LTR
Retrotransposon
LTR LTR
Retrotransposon LTR
LTR
Insertion
IRAP – InterRetroelement PCR
Retrotransposon LTR
LTR
Retrotransposon
LTR LTR
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

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SaffronCrocusGenomicsThessalonikiOnlineMay2024TalkOnline.pptx

  • 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)
  • 4.
  • 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
  • 8.
  • 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)
  • 11.
  • 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)
  • 20.
  • 21. Identical patterns in IRAP gel with diverse Saffron accessions (not 18, of garden-origin)
  • 22.
  • 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
  • 25.
  • 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
  • 28.
  • 29.
  • 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”
  • 31.
  • 32.
  • 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
  • 35.
  • 36.
  • 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