The expert consultation on the use of crop wild relatives for pre-breeding in potato was a workshop organized by the Global Crop Diversity Trust in collaboration with CIP and took place from the 22nd – 24th of February 2012.

1. Do potatoes have a single evolutionary history?
Flor Rodríguez
February 22nd, 2012
Lima, Peru

2. Previous phylogenetic studies
plastid DNA restriction site
S. bulbocastanum
Clade 2 S. cardiophyllum subsp. cardiophyllum
S. cardiophyllum subsp. ehrenbergii
Clade 3
all ser. Piurana
Clade 4 Remaining SA species
NA & CA polyploid species
and S. verrucosum
Clade 1 Most US, Mexican & Central American diploids
Outgroup (S. palustre)
Hosaka et al., 1984; Spooner & Sytsma, 1992;
al.,
Castillo & Spooner, 1997; Rodríguez & Spooner, 1997

3. Previous phylogenetic studies
DNA sequences, WAXY
900 bp
1200 characters
Ser. Longipedicellata c1+2 c4b
Ser. Conicibaccata c3 c4a
Iopetala Group :
S. schenkii & iopetalum c3 c4a c4b
S. hougasii c3 c4b
S. demisum c4a c4b
Rodriguez et al., 2008
al.,

4. Previous phylogenetic studies
DNA sequences
nitrate reductase
1.00
S. lycopersicum
S. acaule 1
1.00 S. acaule 2
0.59 S. demisum 1
0.97 S. demisum 2
S. raphanifolium 2
1200 bp,
0.78 S. raphanifolium 1
0.77 0.99 S. agrimonifolium 1
1.00 S. colombianum 1
0.74 S longiconicum 1
0.78 S. violaceimarmoratum
1500 characters
S. microdontum
0.71 S. lignicaule
1 S. berthaultii
0.69 S. tarijense
0.95 S. infundibuliforme 1
S. sparsipilum 1
0.98
0.98
0.96
S. brevicaule 1
S. infundibuliforme 2
S. chacoense
Clade 4
Genome A
1.00 S. gandarillasii
S. boliviense
S. megistacrolobum
1.00 S. brachycarpum 1
0.85 S. S. verrucosum
hjertingii 1
1.00 S. stoloniferum 1 Ser. Longipedicellata c1+2 c4b
Ser. Conicibaccata c3 c4a
S. demisum 3
1.00 S. demisum 4
S. fendleri 1
S. schenkii 1
0.98
0.96
0.96
1 S. demisum 5
S. demisum 6
S. fendleri 2
Iopetala Group:
S. iopetalum c3 c4b
0.83 S. schenkii 2
1 S. oplosence A1
1 S. oplosence A2
S. oplosence A3
0.94
0.52
S. brevicaule 2
S. leptophyes
S. sparsipilum 2
S. schenckii c1+2 c3 c4b
0.96 S. oplosence A4
1.00 S. oplocense B1
S. oplosence B2
S. demissum c4a c4b
1 S. oplosence A5
0.99 S. oplosence A6
0.91 S. oplosence B3
0.98 S. oplosence B4
0.76 S. stenophyllidium
0.69 S cardiophyllum
S. bulbocastanum
0.97 0.98 S. cardiophyllum subsp.ehrenbergii 1
S. cardiophyllum subsp.ehrenbergii 2
0.99 S. fendleri 3
S. fendleri 4 Diploid species
1.00 S. hjertingii 2
0.93 S. stoloniferum 2
S. stoloniferum 3
S. stoloniferum 4
Clade 1+2 Polyploid species ser. Longipedicellata
Polyploid species ser. Connicibaccata
Genome B
0.75 S. jamesii
0.74 S. pinnatisectum
1.00 0.68 1.00 S. schenkii 3
S. schenkii 4 Polyploid species Acaulia Group
S.trifidum
S. clarum
0.98
1.00
S. morelliforme Polyploid species Iopetalum Group
1.00 S. lesteri
S. polyadenium
S. agrimonifolium 2
Polyploid species ser. Piurana
0.79 S. agrimonifolium 3
0.68
S. agrimonifolium 4
S. brachycarpum 2
Polyploid species ser. Tuberosa
1.00 S. colombianum 2
0.56 S. colombianum 3
72 S longiconicum 2
0.66
Clade 3
1 S. tuquerrense 1
S. tuquerrense 2
0.95 S. andreanum
1 S. paucijugum 1
0.96 0.95 S. paucijugum 2
S. schenkii 5
S. schenkii 6
Piurana Clade
Genome C or D or P
0.57 0.59
S. albornozii
0.97 S. paucijugum 3
0.57 1 S. paucijugum 4
S. tuquerrense 3
S. tuquerrense 4
S. immite
Rodríguez & Spooner, 2009

5. Objectives
 Identify COS appropriate for wild potato
phylogenetic studies
 Generate a molecular data set to reconstruct the
phylogeny of diploid and polyploid wild potatoes

6. COS selection criteria
 Chromosome coverage
 Single-copy in potato
 Sequence amplification length
 Intron and exon content

7. Predominant history …. and others too
12 COSII, 10,886 total characters
The predominant history The other two different histories
History 1 History 3 History 2
with 12COS or 6COS with 1COS with 3COS

8. Another way to see alternative
evolutionary histories
Bayesian concordance analysis (12 COSII)
α = 1, 10 and infinite
The numbers supporting the branches are concordance
values indicating the proportion of the genes supporting
these branches

9. Conclusions
 COS are useful for potato phylogenetic studies
 Intron contents more than 60% are the best to investigate
relationships among closely related species
 The total evidence approach produce a completely resolved
potato phylogeny
 The “prior agreement” approach and “concordance
analysis” identified two more histories in potato
 Additional studies using more accessions and more
nuclear orthologs will show and bring up more details of
potato evolution

10. Challenges
 Sequence more COS/genes
 Identify COS appropriate for wild potato phylogenetic
studies
 Identify all alleles of a genotype

11. Domestication-related loci
 Summary of domestication-related loci with putative
conservation in Solanaceae reported in 15
publications
 At least 3 common markers among maps were used
to determine co-linearity with Tomato-EXPEN2000
map
 171 COSII close to them: 130 of them are single
copy in potato and 76 in potato and tomato

12. Domestication-related loci
 25 traits :

13. Identifying all alleles of a genotype
 Cloning was the traditional approach for uncovering
allelic variants
 Cloning issues: PCR recombination and
heteroduplex fixation
 Asymmetric PCR SSCP

14. Current research
 the number of potato diploid species in each main
clade was incresed up to 70 species
 28 new COS were selected for having a single band
in agarose gel
 direct sequencing and asymmetric-PCR-SSCP

15. Preliminary results
blb
bst
76
ehr
ehr.2
75 cph
Clade 1+2
trf
cph.2
jam
pnt 24 COS
pld
les
clr
24,844 characters
clr.2
mrl
cmm
blv
blv.2
69 mga
mga.2
ber
* tar
tar.2
ifd
* ifd.2
lph
lph.2
71 brc
* ver
spl
spl2 Clade 4
gnd
78 78 gnd.2
chc
75
chc.2
ktz
ktz.2
vio
lxs
79 lmb
lmb.2
54 igl
rap
rap.2
buk
buk.2
chm
* chm.2
pcs
chl p
65 chm B
chq B
jlc
79
dcm
abz
Clade 3
60 adr
mtp
72 imt
68 imt.2
mcq
smp
hcr
hcb
hcb.2
crc
crc.2
etb
pls
dul
50 changes

16. What does COS tell on the evolution of
wild potato polyploid species?
 6 COS sequenced
 11 polyploid species, 2-10 accessions per
species, in total 44 accessions
 37 diploid accessions of 30 species

17. Results in polyploid species
Number of
Species accessions/species Clade 1+2 Clade 3 Clade 4
examined
Precise origin not identified, but at S. lignicaule; S. candolleanum and
Solanum acaule 3/3
least 1 allele with BS=0.96 S. raphanifolium clade
S. candolleanum and S.
3/4 S. hypacrarthrum
raphanifolium clade
S. albicans
S. chilliasense and S. piurae clade; S. candolleanum and S.
1/4
S. chomatophilum raphanifolium clade
S. andreanum and S. albornozii S. laxissimum, S. limbaniense, and
S. agrimonifolium 2/2
clade S. violaceimarmoratum clade
S. colombianum 2/2
S. andreanum and S. albornozii S. laxissimum, S. limbaniense, and
2/2
S. moscopanum clade; S. chomatophilum S. violaceimarmoratum clade
Precise origin not identified, but at S. berthaultii; S. verrucosum; S.
6/7 least 1 allele with BS>0.96 for all candolleanum and S.
accessions raphanifolium clade
S. demissum
S. berthaultii, S. verrucosum, S.
Precise origin not identified, but at
1/7 S. trifidum candolleanum and S.
least 1 allele with BS=0.95
raphanifolium clade

18. Results in polyploid species
Number of
Species accessions/species Clade 1+2 Clade 3 Clade 4
examined
S. stenophyllidium; S. S. andreanum and S.
S. hougasii 5/5 S. berthaultii, S. verrucosum
trifidum albornozii clade
S. berthaultii, S. verrucosum; S. laxissimum,
S. andreanum and S.
S. iopetalum 7/7 S. limbaniense, and S. violaceimarmoratum
albornozii clade
clade
S. polyadenium; S. S. andreanum and S.
5/6 S. berthaultii; S. verrucosum
stenophyllidium; S. trifidum albornozii clade
S. schenckii
1/6 S. stenophyllidium S. berthaultii; S. verrucosum
7/10
S. stenophyllidium S. berthaultii; S. verrucosum
S. stoloniferum
2/10 S. chomatophilum S. candolleanum and S. raphanifolium clade
1/10 S. andreanum and S.
S. polyadenium, S. trifidum S. berthaultii; S. verrucosum
albornozii clade
S. hjertingii 6/6 S. stenophyllidium S. berthaultii; S. verrucosum

19. Conclusions
 Phylogenetic results are often incongruent among different
phylogenetic markers
 COS are useful for potato phylogenetic studies
 There are much diversity and genomic complexity that was
known within traditional recognized polyploid species
 At least 4 polyploid species have multiple origins and allele
losses
 Additional studies using more accessions and more nuclear
orthologs will show and bring up more details of potato
evolution

20. Conclusions
 Results could be useful for designing crossing strategies to
incorporate wild species germplasm into cultivated potato
 To use wild species effectively in potato breeding programs,
it is important to consider both genomes and endosperm
balance number
 The identification of the genomes that have contributed to
allopolyploid species may provide a strategy for their use in
bridging crosses

21. Acknowledgements
Dr. David Spooner
People in Spooner’s lab
Dr. David Baum and Cecile Ane
University of Wisconsin Plant Breeding and Plant
Genetics Program
US Potato genebank