Table S1 Mean, range and standard deviation of the quantitative traits measured in parent genotypes Denmark and DGI007 and their F2 progeny
Trait a
(mapping population 92S05)
Denmark
DGI007
Progeny
Range
Mean (SD)
Range
Mean (SD)
Range
Mean (SD)
σ2
FC (% of dry matter.)
0 - 0.05
0.02 (0.025)
0.15 - 0.4
0.25 (0.077)
0 - 0.8
0.1 (0.151)
0.023
GC (% of d.m.)
0.9 - 2.35
1.62 (0.373)
0.65 - 1.55
1.27 (0.259)
0.5 - 2.7
1.53 (0.369)
0.136
BAC (% of d.m.)
0.65 - 1.05
0.85 (0.111)
0.65 - 1.05
0.82 (0.094)
0.55 - 1.3
0.83 (0.127)
0.016
DF (days) b
132 - 144
139 (3.6)
88 - 103
97 (4.9)
94 - 162
116 (13.9)
193
SH (% hard seed)c
0 - 23
14.2 (7.36)
13 - 59
39.5 (13.1)
0 - 62
22.9 (15.3)
235
PC (1 – 9) d
1
-
7-8
7.69 (0.461)
1-8
4.48 (2.6)
6.74
PS (1 – 9) e
4-6
4.23 (0.576)
1
-
1-5
2.42 (1.09)
1.18
HS (1 – 9) f
1-2
1.15 (0.361)
5-6
5.31 (0.461)
1-6
4.1 (1.23)
1.51
HP (1 – 9) f
a
1
-
4-6
5.08 (0.474)
b
Days to first flowering from an early May sowing in Perth, Australia.
Percent hard seed after 16 weeks in a diurnally fluctuating 15/60 oC temperature cabinet.
d
1-9 rating scale, from no pigmentation (1) to whole calyx tube pigmented (9).
e
f
3.35 (1.26)
Formononetin (FC), genistein (GC), biochanin A (BAC), days to first flowering (DF), pigmentation of calyces (PC) and stipules (PS), hairiness of stipules (HS)
and peduncles (HP), and hardseededness (SH).
c
1-5
1-9 rating scale from no pigmentation (1) to whole stipule pigmented (9).
1-9 rating scale from glabrous (1) to strongly hairy (9).
.
1.59
Table S2 Mean, range and standard deviation of the quantitative traits measured in parent genotypes Woogenellup and Daliak and their F2 progeny
(mapping population 92S80)
Trait a
Woogenellup
Daliak
Progeny
Range
Mean (SD)
Range
Mean (SD)
Range
Mean (SD)
σ2
FC (% of dry matter.)
0.25 - 0.95
0.55 (0.24)
0.25 - 0.95
0.66 (0.199)
0.05 - 1.75
0.63 (0.352)
0.124
GC (% of d.m.)
1.1 - 1.95
1.71 (0.22)
0.45 - 0.95
0.69 (0.317)
0.3 - 2.15
1.1 (0.289)
0.083
BAC (% of d.m.)
0.55 - 0.85
0.73 (0.08)
0.3 - 0.55
0.44 (0.085)
0.3 - 1.8
0.73 (0.214)
0.046
DF (days) b
122 - 133
126 (2.6)
96 - 108
102 (3.4)
110 - 156
126 (8)
64.3
SH (% hard seed)c
0 - 40
11.6 (12.1)
29 - 60
48.55 (9.04)
0 - 66 (12.5)
18.66 (14)
195
PC (1 – 9) d
1
-
8-9
8.92 (0.276)
1-9
6.2 (3.02)
9.15
HS (1 – 9) e
1
-
3-5
4.5 (0.645)
1 – 6 (1.72)
1.94 (1.69)
2.87
a
Formononetin (FC), genistein (GC), biochanin A (BAC), days to first flowering (DF), pigmentation of calyces (PC) and stipules (PS), hairiness of stipules
(HS) and peduncles (HP), and hardseededness (SH).
b
Days to first flowering from an early May sowing in Perth, Australia.
c
Percent hard seed after 16 weeks in a diurnally fluctuating 15/60 oC temperature cabinet.
d
1-9 rating scale, from no pigmentation (1) to whole calyx tube pigmented (9).
e
1-9 rating scale, from glabrous (1) to strongly hairy (9).
Table S3 Summary of SSR marker polymorphisms between T. subterraneum populations 92S05 and 92S80
Marker name
RCS
NMSa
1 438
92S05
92S80
Source
NPMb
NALc
PRd (%)
NPM
NAL
PR (%)
Species
Sequences
Reference
298
395
20.6
365
438
25.4
Trifolium
EST &
(Sato et al., 2005) & Isobe et al.
pratense
genomic
(unpublished observations)
TPSSR
26
9
9
34.6
8
8
30.8
T. pratense
genomic
(Kölliker et al., 2001)
WCS
1 091
172
188
15.8
212
222
19.4
T. repens
EST
Isobe et al. (unpublished
observations)
ats
30
4
9
13.3
6
11
20
T.repens
genomic
(Barrett et al., 2004)
prs
21
5
8
23.8
4
7
19
T. repens
EST
(Barrett et al., 2004)
AI, AJ, AL,
36
6
6
16.7
12
12
33.3
AW, BE, BF,
Medicago
EST
(Sledge et al., 2005)
BAC
(Sledge et al., 2005)
EST
(Julier et al., 2003)
truncatula
BG, BI, MTBA
MT1
2
2
2
100
1
1
50
M.
truncatula
MTIC
2
0
0
0
0
0
0
M.
truncatula
Total
a
2 646
496
Number of markers screened
b
Number of polymorphic markers
c
Number of amplified loci
617
18.7
608
699
22
d
Polymorphic ratio
Table S4 PCR programs used in segregation analysis in the 92S05 population and their
corresponding amplified markers
PCR program
Details
Amplified
markers
SSR Flo1
5 min of initial denaturation cycle followed by first 30 cycles
RCS, WCS
of 94°C for 30 s, 52°C for 45 s and 72°C for 90 s, then 8
cycles of 94°C for 30 s with annealing temperature of 53°C
for 45 s and 72°C for 90 s followed by a final extension for
10 min at 72°C (Bandelj et al., 2004) .
SSR Flo2
5 min of initial denaturation cycle followed by first 15 cycles
RCS, WCS, ats,
of 94°C for 30 s, 53°C for 45 s and 72°C for 100 s, with
prs
0.33°C decrease in annealing temperature per cycle, then
30 cycles of 94°C for 30 s with constant annealing
temperature (53°C) for 45 s and 72°C for 90 s followed by a
final extension for 10 min at 72°C.
SSR Flo3
5 min of initial denaturation cycle followed by first 21 cycles
RCS, WCS, ats
of 94°C for 30 s, 53°C for 45 s and 72°C for 90 s, with 0.33°C
decrease in annealing temperature per cycle, then an
extension of 72°C for 5 min, after that another 30 cycles of
94°C for 30 s with constant annealing temperature (53°C)
for 45 s and 72°C for 90 s followed by a final extension for
10 min at 72°C.
SSR Flo4
5 min of initial denaturation cycle followed by first 30 cycles
RCS, WCS
of 94°C for 30 s, 51°C for 45 s and 72°C for 90 s, then an
extension of 72°C for 5 min, after that another 8 cycles of
10
94°C for 30 s with constant annealing temperature (53°C)
for 45 s and 72°C for 90 s followed by a final extension for
10 min at 72°C.
11
Table S5 SSR markers with polymorphisms at more than one locus in F2 progeny of T.
subterraneum populations 92S05 and 92S80
Number of
Number of markers
Marker name
44
RCS0894, RCS2088, RCS1325, RCS3976, RCS6010,
amplified loci
2
RCS2202, RCS3862, RCS6069, RCS6448, RCS4886,
RCS6272, RCS1632, RCS5324, RCS5058, RCS3226,
RCS6563, RCS5303, RCS0783, RCS4747, RCS4813,
RCS1868, RCS0796, RCS1746, RCS4736, RCS0824,
RCS0831, RCS2481, RCS2850, RCS1234, RCS4695,
RCS6014, RCS6625, RCS2164, RCS4626, RCS2149,
RCS2883, RCS3192, WCS0328, WCS0670,
WCS1212, WCS1376, WCS1322, WCS1249,
WCS1824
3
19
RCS1499, RCS4479, RCS2052, RCS5569, RCS2882,
RCS5288, RCS5549, RCS3159, RCS0889, RCS5603,
RCS7210, RCS1668, RCS2866, RCS2829, RCS4642,
RCS4606, WCS1662, WCS0677, WCS0029
4
12
RCS3171, RCS1630, RCS0861, RCS0967, RCS1411,
RCS3279, RCS2325, RCS3013, RCS3232, RCS1787,
RCS6394, prs279
5
3
RCS1518, RCS5211, WCS0062
6
2
RCS6249, ats003
12