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Overview of RTB Theme 1 –
Unlocking the value and use potential
of genetic resources
Nicolas Roux, Julie Sardos, Mathieu Rouard, Ismail Rabbi, Flor
Rodriguez, Severin Polreich, Bettina Heider, Ranjana Bhattacharjee,
Xavier Perrier, Angelique d’Hont and Luis Augusto Becerra Lopez-Lavalle
RTB Annual Meeting, Lima Peru, 8 December 2015
Research Objectives
• Population structure analysis
• Assessment of genetic diversity
• Gap analysis
• Variety identification and population structure analysis
• Genetic integrity
• In situ versus ex situ
• Genome sequencing
• Bioinformatics
Genome P
Genome A
(Rodriguez et al, in prep)
S. stenotomum
S. goniocalix
S. phureja
Phylogenetic analysis:
wild and cultivated
potatoes
34
41
99
100
100
100
S. bukasovii
S. candolleanum
S. ambosinum
SNP calling 1:
• Tassel 4 GbS Pipeline
• ref. genome:
PGSC_DM_v4.03
• allele freq 0.05-0.95
• <5% missing data in SNP
• <10% missing data in Taxa
• 13,150 SNP
• 260 Taxa
SNP calling 2:
• CIP Pipeline
• ref. genome:
PGSC_DM_v4.03
• allele freq 0.05-0.95
• <5% missing data in SNP
• <10% missing data in Taxa
• 5,241 SNP
• 274 Taxa
Phylogenetic analysis:
• RAxML
• GTR+CAT model
• 1,000 bootstraps
Wild
potatoes
2x cultivated potatoes
Assessment of genetic diversity
384 D. alata germplasm genotyped
with 34 SSR markers
A total of 847 alleles recorded
across 26 SSRs with mean number
of alleles ranging from 10.6
(CIRAD) to 7.5 (IITA)
IITA INRA CIRAD CTCRI
Group 1
Group 2
Group 1
Group 3
Group 3 Group 4
Group 4
Group 5
Group 5
Group 6
Group 7
Arnau et al (2015). SSR-based genetic diversity in
different collection of Dioscorea alata (under
review)
Indicators INR
A
CIRAD CTCRI IITA
N
At
Am
129
217
9.0
83
255
10.6
82
194
8.1
90
181
7.5
Gap analysis
Priorities for gap filling and conservation by country
Potato
• 32 species (43.8%) are High Priority
Species (HPS) and need to be
prioritized for further collecting
• Castañeda-Álvarez et al. 2015 DOI:
10.1371/journal.pone.0122599 potato
Sweetpotato
• 11 out of 14 (79%) species are
High Priority Species (HPS for
further collecting
• Khoury C.K. et al. 2015 doi:
10.3389/fpls.2015.00251
Variety identification
Individual ancestry estimates to determine contribution of ancestral clones to new varieties
Hierarchical clustering dendrogram to determine identical clones.
Variety names often do not match
specific genotypes
182 different names recorded for the entire sample collection and here only names that
occurred > 9 times are represented.
I"
II"
III"
IX"
V"
VI"VII"
VIII" X" XI" KOTEE"AMPENKYENE"
TUAKA"
AFIA0KOFIE"
ESIABAYAA"
BOSOMENSIA"
ABENWOHA"
DEBOR"
BANKYE0"KOKOO"
ANKRA"
Determination of genetically unique varieties in 2 Peruvian diversity hotspots
Baseline characterization
223 76 15
Huancavelica (n =658) Apurímac (n=190)
36.2% of total samples
unique
47.9%
of total samples unique
a. Potato landrace samples (658 from Huancavelica and 190 from Apurimac, Peru) were collected and molecularly analyzed in 2013 and 2014. b. Genetic fingerprinting with 12 polymorphic SSR microsatellite was carried out and 115 alleles
identified in total.
c. The number of unique cultivars and pure duplicates within the genetically characterized (sub)populations was determined.
Geospatial coverage of in situ/ex situ diversity
In situ Ex situ
(baseline
data)
(passport data
CIP
Genebank)
No. different
varieties
No. varieties
registered
Dept: Apurimac
Prov: Cotabambas
Distrito: Haquira* 91 8
Dept: Huancavelica
Prov: Huancavelica
and Acobamba
Districts: Yauli and
Paucara
238
15
• Diversity hotspots geo-spatially
underrepresented at CIP’s genebank.
• Genetic analyses accomplished by 61%
in Huancavelica resulted in 238
different varieties.
total number of varieties from the
respective districts maintained ex situ =
15!
• Former studies of de Haan et al. (2010)
indicated high genotypic diversity and
allelic variation that are site specific.
• Comparative analyses of material
maintained in hotspots and kept in CIP’s
gene bank will help to determine
evolutionary dynamics in situ and
improve the genetic and geo-spatial
coverage of the ex situ collection.
Tetraploid
Diploid
Tri-Pentaploid
Highest
richness Endemism
I. Hotspot identification
MACRO MESO MICRO
Indicators
• Endemism
• Richness
• Coexistence wild relatives
• Diversity of cultural and agro-
ecological conditions
II. Richness-Altitude
Indicators
• Number varieties
• Altitude (masl)
IV. Red List
Indicators
• Abundance
• Frequency
Findings:
• Major diversity between 3800-4300 masl.
Timeline comparison indicates that this altitudinal belt
ascended >250 m during the last 50 years.
• On average >60% of observed varieties are grown by
<5% of farm households. diversity is highly
scattered within the communities
• >70% of varieties show a very low abundance and
frequency in farmers fields. repeated measuring
will reveal how threatened they are.
III. Horizontal dynamics
(crop rotation)
Indicators
• Crop rotation
pattern
Women and Conservation - GENDER
• Women play a central role in in situ
conservation of RTB crops, through
utilizing them for nutrition, health
hazards as well as marketing and
exchange of crops and genetic
material.
• Enhancing their livelihood is essential
to establishing a sustainable, long-term
in situ conservation system for locally
adapted landraces and associated crop
wild relatives.
• Important gender dimensions of in situ
management involve seed selection,
farmer food choices and preference
traits (drivers of conservation), child
nutrition, folk taxonomy and medicinal
uses and feminized conservation,
where temporal or permanent male
migration occurs (Andes, West Africa).
Nieves Mamani, 41 years old conservationist farmer
conserving 235 different potato landraces , from the
Community Cariquina Grande, La Paz Department, Bolivia.
(Photo credit: Victor Iriarte )
a wide phenotypic diversity
but little or no genetic diversity with usual
molecular markers
235,425 polymorphic SNPs of 29 African plantains
and ‘plantain-like’
read number: deviation % from plantain consensus
by chromosome, for each accession
interpretation in chromosome rearrangements
Is a high density SNP coverage of the
genome able to reveal genetic
variations within somaclonal
subgroups?
Clonal differentiation
French Plantain False Horn Plantain Horn Plantain
1) Are these variations due to the
accumulation of mutations over time
(vegetatively propagated) ?
2) Are these variants induced by in
vitro culture?
In 2015, complementary analysis
performed on the accessions only ever
in vivo (provided by IITA Onne station)
Nzumoigne
Ihitisim
Obubit
Ihitisim
11
10
9
8
7
6
5
4
3
2
1
Obino
l’Ewai
Ihitisim
Niangafelo
Agbagba
Nzumoigne
Ntanga
Ihitisim - ITC0121 (analysis 2014)
Ihitisim - ITC0121 (analysis 2015)
Ihitisim – Ni1 (analysis 2015) collection in vivo IITA
Ihitisim was introduced at ITC in 1986 from IITA
Variant genome
Control genome
Chromosome 3
Number of Reads between 10 and 450
8931 SNP s
True-to-type Off-type
Technical Guidelines for
the Safe Movement of
Musa Germplasm –
Published in December
2015
BSV workshop, Montpellier, May 2015
Genome reference sequences
Contribution to the production of Musa genome reference sequences for three
additional M. acuminata subspecies : spp. banksii (Banksii), burmannicoides
(Calcutta4) and zebrina (Maia oa)
spp. malaccensis (DH pahang)
Close to 88% of the assembly anchor to chromosome
(D’hont et al. 2012) and an improved version (Martin et al., BMC genomics, accepted).
zebrina banksii
malaccensis
burmannicoides
Deep re-sequencing (BGI), pan genome and LSV analysis in progress
A
A A A
17-19 August 2015, Bioversity Montpellier, France
January 2015, Ithaca
Bioinformatics: Community of practice
Conclusions
• Benefits of whole genome sequencing
• Synergies between crops and centres
• Importance of data sharing
• Interoperability between databases
• Logistical issues
• Legal issues
• Budget cuts
• In some centres, need for more interactions between
GB-CRP and RTB-CRP
THANK YOU
Augusto Becerra Lopez-
Lavalle
Claudia Perea
Clair Hershey
Joe Tohme
and colleagues
Nicolas Roux
Julie Sardos
Rachel Chase
Mathieu Rouard
and colleagues
Angelique D’Hont
Jean Pierre Horry
Xavier Perrier
Frank Christophe Baurens
and colleagues
Dave Ellis
Flor Rodriguez
Severin Polreich
Bettina Heider
Merideth Bonierbale
and colleagues
Ranjana Bhattacharjee
Tessema Gezahegn
Ismail Rabbi
Rony Swennen
and colleagues