Download Table 1

The dispensable chromosome of Leptosphaeria maculans shelters an effector gene
conferring avirulence towards Brassica rapa
Marie Hélène Balesdent1*, Isabelle Fudal1, Bénédicte Ollivier1, Pascal Bally1, Jonathan
Grandaubert1, Frédérique Eber2, Anne-Marie-Chèvre2, Martine Leflon3, Thierry Rouxel1
1
INRA, UR1290 BIOGER, Avenue Lucien Brétignières, BP 01, F-78850 Thiverval-Grignon,
France
2
INRA, UMR1349 IGEPP, BP35327, F-35653 Le Rheu cedex, France
3
CETIOM, Avenue Lucien Brétignières, BP 01, F-78850 Thiverval-Grignon, France
Supporting information Figs S1S3, Tables S1S6, Methods S1
Methods S1 Inoculation tests: modelling the loss of the dispensable chromosome in
Leptosphaeria maculans populations
Table S1 PCR primers used in this study
Table S2 Analysis of the progeny of crosses between a resistant B. rapa plant (2n=20) and
the susceptible B. napus (2n=38) variety Darmor
Table S3 Analysis of the progeny of crosses between a resistant B. rapa plant (2n=20) and
the susceptible B. napus (2n=38) variety Eurol
Table S4 List and characteristics of SC22 predicted genes
Table S5 List and characteristics of SC22 predicted proteins
Table S6 Interaction phenotypes of wild-type or transformed Leptosphaeria maculans
isolates on a Brassica differential set
Fig. S1 Percentage of plants with different chromosome numbers assessed by flow cytometry
in the progeny of B. rapa x B. napus (AAC) hybrids crossed to their recurrent parents Darmor
or Eurol
Fig. S2 Interaction phenotypes of wild type and transformed Leptosphaeria maculans isolates
with AvrLm11 candidate genes
Fig. S3 Evolution of the frequency of the dispensable chromosome in field populations of
Leptosphaeria maculans under different fitness cost hypotheses
Methods S1
Inoculation tests. Isolates were inoculated on cotyledons of 2 weeks-old seedlings. Each half
cotyledon was punctured with a needle and a drop of 10µL of a 107 spores ml-1 was deposited
on each hole. Plantlets were incubated in a growth chamber at 16 °C (night)-24 °C (day) with
a 12 h photoperiod. Symptoms were scored 14 to 21 days after inoculation using the semiquantitative IMASCORE rating scale comprising 6 classes (IC) (Balesdent et al., 2001). IC1
is the typical hypersensitive response (HR), IC2 includes larger (1.5 to 3 mm) dark necrotic
lesions, IC3 corresponds to lesion without any pycnidia, that may or may not show tissue
collapse as in IC4 to IC6, but that is always sharply delimited by a dark necrotic margin that
may extend within the lesion. IC4-IC6 are characterized by gray-green tissue collapse without
a darkened margin, whose size increases with time, and showing no sporulation (IC4), a few
pycnidia (IC5), or profuse sporulation (IC6). IC1 to IC3 are resistance responses, whereas IC4
to IC6 are susceptibility symptoms. At least 10 seedlings (10 replicates) were inoculated for
each isolate-plant genotype combination. The result of the interaction was evaluated by
calculating either the mean disease scoring, or the percentage of resistance (percentage of
IC1-3) or susceptibility (percentage of IC4-6) symptoms.
Modelling the loss of the dispensable chromosome in Leptosphaeria maculans
populations. We used a simple model in which it is assumed that there is no migration in the
local populations. The population of year i is composed of isolates issued from ascospores
generated by random mating between individuals composing the population of year i-1. The
frequencies of isolates with (MC+) and without (MC-) are fi and (1-fi) respectively.
Probabilities of crosses between two MC+ isolates, two MC- isolates, or one MC+and one MCisolates therefore are fi2, 2fi(1-fi) and (1-fi)2, respectively. MC-x MC- crosses give rise to only
MC- isolates (Leclair et al., 1996). It is hypothesized that MC+ x MC- crosses generate 50%
MC+ and 50% MC- isolates while MC+ x MC+ crosses generate MC- isolates with a mutation
rate, termed G, found as G=0.048 in our study. If considering that the loss of the MC has no
fitness cost (i.e. the proportion of MC- isolates involved in mating at the end of the infectious
cycle is identical to that present in the primary inoculum), the frequency of MC+ isolates in
year i+1 will be:
(1)
which can be simplified as (1)
fi+1=(1-G)fi2 + 0.5x2fi(1-fi) + 0x(1-fi)2
fi+1= fi(1-G fi)
If considering that the loss of the MC has a fitness cost, this can be globally assimilated to a
lowered proportion of MC- isolates involved in mating at the end of the infection cycle,
whatever the biological parameter(s) responsible for this fitness cost. From the (1-fi) MCisolates constituting the primary inoculum in year i, only ρ(1-fi) will therefore contribute to
sexual mating producing inoculum for year i+1, with 0 ρ1. Similarly MC+ isolates
contributing to mating will be fi’= 1- ρ(1-fi). Under this hypothesis the proportion of MC+
isolates at the beginning of year i+1 will be calculated by replacing fi by fi’ in (1) as follows
(2)
fi+1= fi’(1-Gfi’) = (1- ρ(1-fi))(1-G(1- ρ(1-fi)))
This equation was used to construct the curves of evolution with time of the frequency of the
MC in natural populations starting with an initial frequency of 0.955 (estimated from the
2000-2001 sampling), G=0.048 and ρ values ranging from 0 (MC- isolates are unable to
complete the life cycle) to 1 (no fitness cost linked to the loss of the MC). As shown in Fig.
S3, only a strong fitness cost linked to the loss of the MC can explain the maintenance of the
MC in such high proportions during a 10 years time interval.
Supporting Table S1 PCR primers used in this study
Primer name
Sequence
Location on
L. maculans genomea
Min22.0_Ub
ATAGGTCTACCTTAAATATATAATAGC
SC22_100123..100149
Min22.0_L
CAAGACCTATAGATAGTATCCTTTTC
SC22_100620..100645 (c)
Min22.1_U
ACAAAGGCGACGAAACAGG
SC22_428258..428258 (c)
Min22.1_L
AGAAAGATTGGCTTCGTTGC
SC22_427877.. 427896
Min22.4_U
CAAGATGCGCAAGATGACAG
SC22_661552.. 661571
Min22.4_L
CTTGCCCAATCTGGGATTCT
SC22_661713..661732 (c)
Min22.6_U
CGACTCGTCCATTCTTCCAT
SC22_712428..712447
Min22.6_L
GGAGGAAACCGAGCAGAGTA
SC22_712589..712608 (c)
Min16987_U
GCTTACCTCTCTTGTAAGGGTA
SC22_294720..294741(c)
Min16987_L
TGCAAATAGGGTAAATAGGTAAGC
SC22_294545..294568
Min16163_U
TTTGAAGCCTACTACCACTGAAGA
SC22_553588..553611 (c)
Min16163_L
CTGCTGTTGGAGGCAAAGTC
SC22_553538..553557
P119130_U
CGTGAGGTCTCTGAAGAAGC
SC22_436773..436792 (c)
P119130_L
GGACAGAGAAGCTCGACACG
SC22_436574..436593 (c)
P119250_U
CAGGGGTAAAGTTGGTGGTG
SC22_609109..609128
P119250_L
GCCTTGGCTTTATCCTCCTC
SC22_609410..609429 (c)
P119260_U
ATGCCCAACACTCAAACTCC
SC22_624709..624728
P119260_L
GCAGCATAGTCAGCGAATTG
SC22_624994..625013 (c)
P119270.1_U
ACTATCACGGGCGATGATCT
SC22_629390..629409
P119270.1_L
CCCACTTCCATATATCTGTAATTCTA
SC22_629671..629696 (c)
MatU
TGGCGAATTAAGGGATTGCTG
SC20_556096..556116 (c)
MatL1
CTCGATGCAATGTACTTGGAGC
NA
MatL2
CGGAGGTGAAGTTGAAGCCG
SC20_555675..555694
AvrLm11-U2 (uP119060_U)
TGCGTTTCTTGCTTCCTATATTT
SC22_294427..294449 (c)
AvrLm11-L (uP119060_L)
CAAGTTGGATCTTTCTCATTCG
SC22_294090..294111
AvrLm11-5UTRU
GCGTTTCTTGCTTCCTATATTTTCTGC
SC22_294422..294448 (c)
AvrLm11-5UTRnestU
GCAAACTGGAAGGGGCAGTAGGCTG
SC22_294329..294353 (c)
AvrLm11-3UTRU
GCAAGTTGGATCTTTCTCATTCGC
SC22_294089..294112
AvrLm11-3UTRnestU
CGTGTGTACTCCTTCCGTTACGACC
SC22_294234..294258
AvrLm11U1
GCCGCTTAGCTACACTTCGC
SC22_294983..295002 (c)
AvrLm11-XhoL
GAGAGACTCGAGCCCTCTATATGCGTTCCTTAG
SC22_293518..293538
a
Location (in bp) along Super-Contig 22 (SC22) or 20 (SC20); (c) indicates a reverse orientation
of the primer; NA, not applicable (the primer is located on the opposite Mating-Type idiomorph
to that of the sequenced isolate).
b
Markers in bold are those used to check the complete deletion of SC22 in field isolates lacking
AvrLm11.
Supporting Table S2 Analysis of the progeny of crosses between a resistant B. rapa plant
(2n=20) and the susceptible B. napus (2n=38) variety Darmor
Cross
analyseda
Resistance
segregationb
R
F1
BC1
BC2
BC3
a
S
χ²
(p
value)
Meiotic stability in selected resistant plants
Selected
plant #
1g 0
49 69 3.39
1
(0.066) 2
3
51 23 10.59
1
(0.001) 2
3
4
5
6
19 9 3.57
1
(0.059) 2
3
4
5
6
7
8
2nc PMCd Average meiotic behavioure
%
Cells
with
19IIf
38
38
38
38
38
38
38
38
39
38
38
38
38
38
38
38
39
14.3
0
25**
55
65**
25
25
25
66.7**
42.9
45
63**
43.5
54.5
55.6
-
21
22
20
20
20
20
20
20
20
21
21
20
20
23
22
9
6
1.90I+17.38II+0.38III+0.05IV
3.36I+16.54II+0.27III+0.18IV
1.75I+17.6II+0.15III+0.15IV
0.75I+18.4II+0.15III
0.6I+18.55II+0.1III
1.45I+18.2II+0.05III
1.35I+18.2II+0.15III
0.95I+17.5II+0.55III+0.1IV
2.3I+18.35II
0.76I+18.62II
0.8I+18.05II+0.24III+0.1IV
1.1I+18.35II+0.05IV
0.4I+18.5II+0.15IV
0.78I+18.26II+0.17IV
0.82I+18.46II+0.09III
0.89I+18.56II
1.17I+18.67II+0.17III
F1 is the hybrid between one B. rapa resistant plant and the susceptible B. napus cv Darmor.
BC1, BC2 and BC3 were obtained by crossing one resistant plant from the previous
generation showing 2n=38 and the best meiotic behaviour with the susceptible recurrent B.
napus cv.
b
R, number of resistant plants; S, number of susceptible plants toward L. maculans isolates
with AvrLm11 following cotyledon inoculation tests. χ² and (p value) are given for the
expected 50:50 R:S segregation.
c
Number of chromosomes of the selected plant as estimated by flux cytometry.
d
Number of Pollen Mother Cells (PMC) observed for each selected resistant plant.
e
Univalents (I), bivalents (II), trivalents (III) and quadrivalents (IV) observed at metaphase 1.
f
** The plants with the highest percentage of 19 bivalents (II) were selected for the next BC
generation.
g
for this interspecific cross, very few seeds were obtained and only one plant germinated and
could be phenotyped for resistance.
Supporting Table S3 Analysis of the progeny of crosses between a resistant B. rapa plant
(2n=20) and the susceptible B. napus (2n=38) variety Eurol
Cross
analyseda
Resistance
segregationb
R
F1
BC1
BC2
BC3
a
S
χ²
(p
value)
10 2
48 73 5.16
(0.023)
Meiotic stability in selected resistant plants
Selected
plant #
1
2
3
19 62 22.83
1
-6
(2 10 ) 2
3
4
5
6
7
8
8 25 8.76
1
(0.0031) 2
3
4
2nc PMCd Average meiotic behavioure
%
Cells
with
19IIf
38
38
38
38
38
38
38
38
38
38
38
38
38
38
38
0
20**
0
50
55
45
50
65
40
61.9**
47.6
60
50**
65**
55.6
20
20
20
22
20
20
20
20
20
21
21
20
20
20
18
3.80I + 16.60II + 0.25IV
1.15I+17.80II+0.35III+0.05IV
1.00I+16.65II+0.3III+0.70IV
0.64I+18.23II+0.18III+0.09IV
0.8I+18.45II+0.1III
0.55I+18II+0.35III+0.1IV
0.85I+18.4II+0.05III+0.05IV
0.2I+18.4II+0.25IV
1I+17.85II+0.3III+0.1IV
0.71I+18.43II+0.14III
0.9I+18.34II+0.14III
0.5I+18.45II+0.15IV
0.2I+18.4II+0.25IV
0.6I+18.3II+0.2IV
1.11+18.45II
F1 is the hybrid between one B. rapa resistant plant and the susceptible B. napus cv Eurol.
BC1, BC2 and BC3 were obtained by crossing one resistant plant from the previous
generation showing 2n=38 and the best meiotic behaviour with the susceptible recurrent B.
napus cv.
b
R, number of resistant plants; S, number of susceptible plants toward L. maculans isolates
with AvrLm11 following cotyledon inoculation tests. χ² and (p value) are given for the
expected 50:50 R:S segregation.
c
Number of chromosomes of the selected plant as estimated by flux cytometry.
d
Number of Pollen Mother Cells (PMC) observed for each selected resistant plant.
e
Univalents (I), bivalents (II), trivalents (III) and quadrivalents (IV) observed at metaphase 1.
f
** The plants with the highest percentage of 19 bivalents (II) were selected for the next BC
generation.
.
Supporting Table S4 List and characteristics of SC22 predicted genes
Location on
SC22a
Isochore
locationb
Feature
lenght
(bp)
TpA/ApTc
lmctg_1587_v2_egn4_Lema_T119030.1
101169..104023
GC1
2855
0.81
1.02
2220
55.45
56.35
lmctg_1587_v2_egn4_Lema_uT119040.1
104915..105094 (c)
GC1
180
1.76
1.00
102
44.12
52.94
lmctg_1587_v2_egn4_Lema_T119050.1
105263..106588 (c)
GC1
1326
1.19
0.96
1326
56.33
61.31
lmctg_1587_v2_egn4_Lema_uT119060.1
294086..294450 (c)
AT2
365
1.61
0.99
288
48.26
54.17
427173..427364
GC2
192
0.81
1.52
192
21.88
54.69
lmctg_1589_v2_egn4_Lema_T119070.1
427482..429663 (c)
GC2
2182
1.31
0.91
2001
54.57
57.72
lmctg_1589_v2_egn4_Lema_uT119080.1
429788..429979 (c)
GC2
192
0.32
1.28
192
63.54
68.75
lmctg_1589_v2_egn4_Lema_T119090.1
430119..431945
GC2
1827
0.63
1.01
1827
63.27
67.00
lmctg_1589_v2_egn4_Lema_T119100.1
432453..433069
GC2
617
0.72
1.00
510
53.14
53.53
lmctg_1590_v2_egn4_Lema_T119110.1
433905..434534
GC2
630
1.26
0.94
357
48.18
44.54
lmctg_1590_v2_egn4_Lema_T119120.1
434982..435968
GC2
987
1.38
0.95
624
50.32
52.40
lmctg_1590_v2_egn4_Lema_T119130.1
436526..436999 (c)
GC2
474
0.97
1.03
321
58.57
52.34
lmctg_1590_v2_egn4_Lema_T119140.1
437248..438727 (c)
GC2
1480
1.13
0.93
1362
53.52
61.01
lmctg_1590_v2_egn4_Lema_T119150.1
439271..440148 (c)
GC2
878
1.30
0.89
807
57.37
66.17
lmctg_1590_v2_egn4_Lema_T119160.1
440524..441130
GC2
607
0.74
1.10
555
50.99
55.68
lmctg_1590_v2_egn4_Lema_T119170.1
441180..441398
GC2
219
1.12
1.17
219
46.58
53.42
lmctg_1590_v2_egn4_Lema_T119180.1
441826..443256
GC2
1431
0.89
1.14
1431
53.67
60.59
lmctg_1590_v2_egn4_Lema_T119190.1
443437..443912
GC2
476
1.54
1.04
345
48.99
55.65
lmctg_1590_v2_egn4_Lema_T119200.1
549658..551406 (c)
GC3
1749
0.95
1.15
1749
50.66
51.97
lmctg_1590_v2_egn4_Lema_T119210.1
552584..553177
GC3
594
0.56
1.32
594
58.42
54.04
lmctg_1590_v2_egn4_Lema_T119220.1
555624..558041
GC3
2418
0.78
1.23
2367
50.61
50.57
lmctg_1590_v2_egn4_Lema_uT119230.1
559439..560239 (c)
GC3
801
1.09
0.96
270
53.33
62.22
lmctg_1590_v2_egn4_Lema_uT119240.1
560674..560970
GC3
297
0.98
0.99
297
52.86
44.44
lmctg_1590_v2_egn4_Lema_T119250.1
606564..610438 (c)
GC4
3875
1.10
1.07
2880
57.05
63.13
lmctg_1590_v2_egn4_Lema_T119260.1
624144..625048 (c)
GC5
905
2.26
0.71
360
55.83
56.67
Gene ID
AT02_SuperContig_22_10_320860_321051
CDS
CpA+TpG
Lenght %GC %GC3d
/ApC+GpT
(bp)
lmctg_1590_v2_egn4_Lema_T119270.1
629387..629707
GC6
321
1.76
1.06
321
50.16
47.66
lmctg_1590_v2_egn4_Lema_uT119280.1
629911..630127
GC6
217
0.89
1.24
129
41.86
39.53
lmctg_1590_v2_egn4_Lema_T119290.1
632100..632856 (c)
GC6
757
1.35
0.84
585
51.45
64.62
lmctg_1591_v2_egn4_Lema_T119300.1
633273..634431 (c)
GC6
1159
0.81
1.10
1098
66.76
78.69
lmctg_1591_v2_egn4_Lema_T119310.1
660397..662577
GC7
2181
1.00
1.05
2055
56.40
60.88
lmctg_1592_v2_egn4_Lema_T119320.1
703529..704293 (c)
GC8
765
1.09
0.99
765
52.81
53.33
lmctg_1592_v2_egn4_Lema_T119330.1
704474..705397
GC8
924
0.85
1.17
924
54.44
61.04
lmctg_1592_v2_egn4_Lema_T119340.1
705880..706644 (c)
GC8
765
0.82
0.95
765
53.59
48.24
lmctg_1592_v2_egn4_Lema_T119350.1
706950..708772
GC8
1823
1.05
1.06
1755
56.58
60.51
lmctg_1592_v2_egn4_Lema_T119360.1
709203..712628 (c)
GC8
3426
0.87
1.18
3243
59.08
61.15
lmctg_1592_v2_egn4_Lema_T119370.1
713190..713936
GC8
747
1.13
0.94
747
55.56
61.04
a
Location (in bp) along SC22 ; (c) indicates a reverse orientation of the gene.
ATi and GCi refers to the AT-rich or GC-equilibrated isochores, as represented in Fig. 1.
c
TpA/ApT and (CpA+TpG)/(ApC+GpT) are two indices used to detect signatures of RIP (Repeat Induced Point) mutations.
d
Percentages of codons with a G or C at the third base position.
b
Supporting Table S5 List and characteristics of SC22 predicted proteins
Expression in
specific condition:d
Protein
lenght
(AA)
Predicted
localization
/targetinga
BLASTb
lmctg_1587_v2_egn4_Lema_P119030.1
739
-
-
Yes
+
+
+
+
lmctg_1587_v2_egn4_Lema_uP119040.1
33
-
-
Yes
-
+
-
-
lmctg_1587_v2_egn4_Lema_P119050.1
441
M
Yes
Yes
+
+
+
+
lmctg_1587_v2_egn4_Lema_uP119060.1
95
S
-
Yes
-
+/-
+
+/-
AT02_SuperContig_22_10_320860_321051
63
S
-
Yes
+
-
-
+
lmctg_1589_v2_egn4_Lema_P119070.1
666
-
Yes
Yes
+
+
+
+
lmctg_1589_v2_egn4_Lema_uP119080.1
63
M
-
Yes
+
-
+
+
lmctg_1589_v2_egn4_Lema_P119090.1
608
M
Weak
Yes
+
+
+
+
lmctg_1589_v2_egn4_Lema_P119100.1
169
-
-
Yes
+
+
+
+
lmctg_1590_v2_egn4_Lema_P119110.1
118
-
-
Yes
+
-
-
-
lmctg_1590_v2_egn4_Lema_P119120.1
207
-
-
Yes
+/-
-
-
+
lmctg_1590_v2_egn4_Lema_P119130.1
106
S
-
Yes
+
+
+
+
lmctg_1590_v2_egn4_Lema_P119140.1
453
TM
Weak
Yes
+
+
+
+
lmctg_1590_v2_egn4_Lema_P119150.1
268
TM
-
Yes
+
+
+
+
lmctg_1590_v2_egn4_Lema_P119160.1
184
-
-
Yes
+
+
+
+
lmctg_1590_v2_egn4_Lema_P119170.1
72
-
-
Yes
+
+
+
+
lmctg_1590_v2_egn4_Lema_P119180.1
476
-
Yes
Yes
+
+
+
+
lmctg_1590_v2_egn4_Lema_P119190.1
114
-
-
No
-
-
-
-
lmctg_1590_v2_egn4_Lema_P119200.1
582
-
-
Yes
+
+
+
+
lmctg_1590_v2_egn4_Lema_P119210.1
197
-
-
Yes
-
-
+
-
lmctg_1590_v2_egn4_Lema_P119220.1
788
-
Weak
Yes
+
+
+
+
lmctg_1590_v2_egn4_Lema_uP119230.1
89
S
-
No
-
-
-
-
lmctg_1590_v2_egn4_Lema_uP119240.1
98
-
-
No
-
+/-
-
-
Protein ID
Micro-array
supportc
in
vitro
3dpi 7dpi
14dpi
lmctg_1590_v2_egn4_Lema_P119250.1
959
S
Yes
Yes
+
+
+
+
lmctg_1590_v2_egn4_Lema_P119260.1
119
M
-
No
-
-
-
-
lmctg_1590_v2_egn4_Lema_P119270.1
106
-
-
No
-
-
-
-
lmctg_1590_v2_egn4_Lema_uP119280.1
42
-
-
No
-
-
-
-
lmctg_1590_v2_egn4_Lema_P119290.1
194
-
-
Yes
+
+
+
+
lmctg_1591_v2_egn4_Lema_P119300.1
365
-
Yes
Yes
-
+
+
-
lmctg_1591_v2_egn4_Lema_P119310.1
684
S
-
Yes
+
+
+
+
lmctg_1592_v2_egn4_Lema_P119320.1
254
-
-
Yes
+
+
+
+
lmctg_1592_v2_egn4_Lema_P119330.1
307
-
-
Yes
+
+
+
+
lmctg_1592_v2_egn4_Lema_P119340.1
254
-
-
Yes
+
-
-
+
lmctg_1592_v2_egn4_Lema_P119350.1
584
M
Yes
Yes
+
+
+
+/-
lmctg_1592_v2_egn4_Lema_P119360.1
1080
-
-
Yes
+
+
+
+
lmctg_1592_v2_egn4_Lema_P119370.1
248
-
-
Yes
+
+
+
+
a
Predictions of subcellular localisation of the protein; S, secretory pathway; M, mitochondrion; TM, trans-membrane domains; -, no specific
localisation predicted.
b
BLAST hits of the proteins against non-redundant data bases (September 2012); Yes, BLAST hit with a p value < e-10; Weak, BLAST hit with
a p value >e-5, -, no BLAST hit.
c
Micro-array data from Rouxel et al., 2011; Yes, gene models with expression levels higher than three times the median of random probe
intensities in at least two of three biological replicates; No, gene models with expression levels not significantly different from random probe
intensities.
d
Micro-array data from Rouxel et al., 2011; +, the gene is expressed in the two technical repeats for the corresponding condition; +/-, the gene is
expressed for only one of two technical repeats; -, the gene is not expressed whatever the technical repeat; in vitro, RNA was extracted from L.
maculans mycelium grown in Fries liquid medium; 3, 7 and 14 dpi, RNA was extracted from oilseed rape cotyledons inoculated with L.
maculans 3, 7 and 14 days post inoculation, respectively.
Supporting Table S6 Interaction phenotypes of wild-type or transformed Leptosphaeria
maculans isolates on a Brassica differential set
Isolateb
B. napus or B. rapa
linea
v23.1.2
(Av5-6-7-8-11)
v23.1.3
(Av1-4-5-6-7-8-11)
Westarc
Bristol
(Rlm2, Rlm9)
Columbus
(Rlm1, Rlm3)
Darmor (Rlm9)
Pixel (Rlm4)
03.22.3.1 (Rlm3)
02.23.3.1 (Rlm7)
02-159-4-1 (Rlm11)
5.0 ± 0d
a
5.0 ± 0
IBCN14 +
AvrLm11
(Av5-6-11)
5.0 ± 0
IBCN14
(Av5-6)
4.9 ± 0.4
5.0 ± 0
4.9 ± 0.3
4.7 ± 0.5
4.4 ± 0.9
5.0 ± 0
5.0 ± 0
5.0 ± 0
5.0 ± 0
1.0 ± 0
1.0 ± 0
1.8 ± 0.4
5.00 ± 0
1.00 ± 0
5.0 ± 0
1.2 ± 0.4
1.3 ± 0.5
5.00 ± 0
5.00 ± 0
4.9 ± 0.45
5.0 ± 0
5.0 ± 0
1.3 ± 0.7
3.9 ± 1.6
4.6 ± 0.5
4.8 ± 0.4
4.6 ± 0.7
4.9 ± 0.2
3.4 ± 1.0
The resistance gene content is given in brackets.
The avirulence allele composition is given in brackets.
c
Only resistance genes in plant genotypes that were susceptible to IBCN14 were assessed.
d
Mean phenotypic score17 days post inoculation for 10 (v23.1.2 and v23.1.3) to 20
(IBCN14, IBCN14+AvrLm11) plants, on a 1 (resistance) to 6 (susceptibility) scale. Scores
> 3 correspond to compatible interactions (green); scores  3 correspond to incompatible
interactions (pink). Values are mean ± 1SD.
b
25
eurol
darmor
20
15
Plants (%)
10
5
0
29 30 31 32 33 34 35 36 37 38
2n
Supporting Fig. S1 Percentage of plants with different chromosome numbers
assessed by flow cytometry in the progeny of B. rapa x B. napus (AAC) hybrids
crossed to their recurrent parents Darmor or Eurol.
120
% of inoculation 100
sites leading to a
80
compatible
interaction
60
phenotype
40
a
20
0
120
100
bIBCN14
v23.1.3
IBCN14
+ uP119060
IBCN14
+ P119130
IBCN14
v23.1.3
IBCN14
+ uP119060
IBCN14
+ P119130
IBCN14
v23.1.3
80
60
40
20
0
120
100
c
80
60
40
20
0
IBCN14
+ uP119060
IBCN14
+ P119130
Supporting Fig. S2 Interaction phenotypes of wild type and transformed Leptosphaeria
maculans isolates with AvrLm11 candidate genes
▲, virulent isolate BCN14; ■, the v23.1.3 avirulent sequenced isolate; Green, 18 isolates
transformed with the candidate AvrLm11 gene Lema_uP11060; blue, 14 isolates transformed
with another predicted gene of SC22, Lema_P119130.1. Values are percentage of inoculation
points corresponding to a susceptibility symptom (Infection classes 4-6) 15 days after
inoculation on a, the susceptible control Westar, b, the Rlm7 line 02.23.3.1 and c, the resistant
B. rapa line 02-159-4-1. Only comlementation of IBCN14 with uP119060.1 restores the
avirulent phenotype of v23.1.3 on the B. rapa line.
ρ=
1.0
Série12
0
Série3
0.2
Série4
0.3
0.9
Série6
0.4
Frequency of the
dispensable
chromosome in 0.8
populations
Série7
0.5
Série8
0.6
Série9
0.7
Série10
0.8
Série11
0.9
1
0.7
Série1
Série5
0.6
0.5
0
1
2
3
4
5
6
7
8
9
10 11 12
Time (Years)
Supporting Fig. S3 Evolution of the frequency of the dispensable chromosome in field
populations of Leptosphaeria maculans under different fitness cost hypotheses.
Each curve corresponds to the theoretical frequency over time under the random mating
hypothesis, an initial frequency as estimated by field samplings, a frequency of loss at meiosis
of 0.048 as observed under controlled condition, and different fitness cost (1-ρ) ranging from
ρ=1 (no fitness cost) to ρ=0 (isolates that have lost the dispensable chromosome do not
contribute to the next sexual cycle). ▲, observed frequencies of the dispensable chromosome
in field populations collected in France from 2000 (year 0) to 2010 (year 10).