Supporting Figure Legends

Supporting figure legends

Fig. S1Examples of phenotypic divergence between the mapping parents and variation in RILs. (a) plant architecture. (b) panicle. (c) grains. Photos of whole plants and panicles were taken 102 days after transplanting, and photos of mature grains were taken after harvesting.

Fig. S2 95% confidence intervals (CI) and standard deviations of top five major QTL of simulated populations with fixed marker position and QTL position.

Fig. S3 Mutations in five candidate genes between Nipponbare and 93-11. Wild-type genes cloned in the previous studies are illustrated, with bars and arrow heads representing exons and UTRs and lines between them representing introns. Mutations are indicated in red color.

A. By comparing the sequence of TAC1between our mapping parents, Nipponbare and 93-11, we found that Nipponbare shared the same mutation with all japonica cultivars which causing small tiller anglesthan indica rice (Yu et al. 2007). The mutation causing small tiller angles, or more upright tillers, is an A to G substitution at the 3’-splicing site of the fourth intron of the gene. This strongly supports the candidacy of TAC1 for qTA-9.

B. When the sequence of the gene was compared between Nipponbare and 93-11, a C to G substitution in the third exon was found to introduce a premature stop codon in 93-11. Meanwhile, a loss-of-function mutation of SD1 through a large deletion across the first and second exons was considered to be responsible for the short stature of rice variety IR8 (Sasaki et al. 2002).This is consistent with the result of the QTL analysis that the 93-11 allele of qPH-1 makes tiller shorter, which supports the candidacy of SD1 for qPH-1.

C. Three nonsynonymous substitutions were found by comparing Nal1 sequences between Nipponbare and 93-11. These different codons are CAT (H) and CGT(R), GTG (V) and GCG (A), and ATC (I) and GTC (V) for Nipponbare and 93-11, respectively (Fig. S2c).At the same time, the mutant with a 30 bp deletion in the fourth exon in the gene has a narrow leaf (Qi et al. 2008). The results suggest that the natural variation of the gene may have contributed to leaf-width variation among rice cultivars.

D. Comparing the coding region of GS3 between Nipponbare and 93-11, we found that the gene of 93-11 had several deletions in the first exon resulting in frame-shift mutations and truncation of the protein.A nucleotide substitution from C to A in the second exon of the gene led to a premature stop codon in Minghui 63. The truncation of protein was responsible for the substantial increased in grain length(Fan et al. 2006). The consistency between the loss-of-function mutation of the gene and longer grains of 93-11 supports the candidacy of GS3 for qGL-3.

E. The loss-of-function mutation of qSW5 resulting from a 1.2 kb deletion in the coding region led to the growth of wider grains in the japonica cultivar Nipponbare (Shomura et al. 2008). The same mutation was found between our mapping parents, Nipponbare and 93-11, which supports the candidacy of qSW5 for qGW5.

Fig. S4 Relation between genetic distance and physical length of adjacent bins on the linkage map.Red dots represent bins overlapped with centromeres.

Table S1Phenotypic difference of fourteen traits between the mapping parents, Oryza sativa spp. japonica cv. Nipponbare and spp. indica cv. 93-11

Traitsa / Nipponbare / 93-11
Heading date (days) / 75 / 92 / NA
Culm diameter (mm) / 7.26±0.755 / 8.49±0.404
Plant height (cm) / 84.3±0.82 / 124.4±1.71 / **
Flag leaf length (cm) / 26.17±4.793 / 38.20±2.052 / *
Flag leaf width (cm) / 1.27±0.104 / 1.98±0.076 / **
Tiller angle (scale) / 2.5 / 3.0 / NA
Tiller number / 18.0±3.16 / 13.5±2.52
Panicle length (cm) / 20.13±0.808 / 26.97±1.380 / **
Awn length (cm) / 0 / 1.30±0.80 / *
Grain length (mm) / 6.85±0.216 / 9.46±0.195 / **
Grain width (mm) / 2.94±0.139 / 2.66±0.119 / **
Grain thickness (mm) / 2.05±0.049 / 2.18±0.039 / **
1,000-grain weight (g) / 25.05±0.012 / 30.62±0.509 / *
Spikelet number / 92.0±8.72 / 207.3±24.19 / **

aMean and standard deviation of the traits except for heading date and tiller angle that were measured as one value for each parent.

*, **Difference significant at the P = 0.05 and 0.01, respectively.

TableS3Physical length and genetic distance of recombination bins and comparison with conventional molecular markers used in a previous study

Binsa / Conventional Markersb
Number of Bins / Physical Length of Bins(Mb) / Genetic Distance of Adjacent Bins (cM) / Genetic Length(cM) / Number of markers / Genetic Distance of Adjacent Markers (cM) / Genetic Length(cM)
Mean / Max / Meanc / Max / Meanc / Max
Chr.01 / 285 / 0.16 / 3.4 / 0.82 / 8.3 / 216.9 / 444 / 0.90 / 5.9 / 181.8
Chr.02 / 241 / 0.15 / 4.0 / 0.70 / 3.2 / 161.4 / 327 / 1.01 / 15.6 / 157.9
Chr.03 / 240 / 0.16 / 0.8 / 0.69 / 3.2 / 157.9 / 381 / 0.95 / 7.9 / 166.4
Chr.04 / 203 / 0.18 / 1.6 / 0.66 / 2.8 / 122.3 / 260 / 1.09 / 8.6 / 129.6
Chr.05 / 176 / 0.17 / 1.8 / 0.73 / 2.9 / 122.6 / 275 / 0.89 / 5.0 / 122.3
Chr.06 / 186 / 0.17 / 3.8 / 0.73 / 4.8 / 122.7 / 322 / 0.96 / 13.6 / 126.3
Chr.07 / 167 / 0.18 / 1.7 / 0.68 / 3.2 / 105.7 / 241 / 1.11 / 13.2 / 118.6
Chr.08 / 192 / 0.15 / 0.8 / 0.73 / 2.5 / 120.5 / 214 / 1.08 / 9.2 / 121.2
Chr.09 / 133 / 0.18 / 3.6 / 0.78 / 3.4 / 95.5 / 179 / 1.26 / 13.9 / 93.5
Chr.10 / 145 / 0.16 / 1.8 / 0.67 / 2.4 / 85.6 / 162 / 1.00 / 9.3 / 83.8
Chr.11 / 182 / 0.17 / 5.8 / 0.66 / 2.1 / 105.5 / 233 / 1.09 / 9.7 / 119.5
Chr.12 / 184 / 0.15 / 0.9 / 0.75 / 3.2 / 122.9 / 197 / 1.30 / 9.9 / 109.5
Genome / 2334 / 0.16 / 5.8 / 0.72 / 8.3 / 1539.5 / 3235 / 1.03 / 15.6 / 1530.4

aThe number of recombination bins, the average and maximal physical length of bins, the average and maximal genetic distance between adjacent bins, and the total genetic length are given for each chromosome and the entire genome.

bThe information of conventional molecular markers was taken from website:

cGenetic distance larger than zero was included for mean calculation.Average value of entire genome was 0.66 cM including distance of zero.

Table S4 Comparison of frequency distribution of genetic distance amonglinkage maps constructed by recombination bins,simulated markersandconventional markers

Genetic Distance (cM) / Percent of Adjacent Bins / Percent of Adjacent 2330Simulated Markers / Percent of Adjacent Conventional Markersa
0 / 7.88 / 32.57 / 53.68
0.1-1 / 76.61 / 46.68 / 31.55
1.1-2 / 12.19 / 14.11 / 9.77
2.1-3 / 2.67 / 4.53 / 2.95
3.1-4 / 0.43 / 1.47 / 0.74
4.1-5 / 0.09 / 0.30 / 0.59
5.1-6 / 0.09 / 0.26 / 0.12
6.1-7 / 0 / 0.04 / 0.12
7.1-8 / 0 / 0 / 0.06
8.1-9 / 0.04 / 0 / 0.06
9.1-10 / 0 / 0 / 0.19
10.1-11 / 0 / 0.04 / 0.03
11.1-12 / 0 / 0 / 0
12.1-13 / 0 / 0 / 0
13.1-14 / 0 / 0 / 0.09
14.1-15 / 0 / 0 / 0
15.1-16 / 0 / 0 / 0.03

aThe information of conventional molecular markers was taken from website:

Table S5. 95% confidence intervals of QTL indentified using recombination bins and simulated markers.

Trait / QTL / 95% Confidence Interval (cM) a
recombination bins / simulation of 2330 markers / simulation of 238 markers
Heading date / qHD-3 / 1.6 / 1.6 / 8.5
Culm diameter / qCD-2 / 1.9 / 2.0 / 6.8
qCD-4 / 3.0 / 6.2 / 12.1
qCD-12 / 2.0 / 2.0 / 20.2
Plant height / qPH-1 / 3.2 / 1.4 / 4.0
qPH-2 / 1.6 / 1.5 / 11.1
qPH-3 / 2.1 / 4.4
qPH-12 / 5.0 / 6.1 / 7.9
Flag leaf length / qFLL-2 / 4.6 / 4.7 / 8.6
qFLL-5 / 6.1 / 7.4
qFLL-12 / 1.8 / 3.3 / 10.7
Flag leaf width / qFLW-1 / 4.2 / 5.3
qFLW-4 / 2.2 / 1.6 / 12.1
qFLW-7 / 7.8
qFLW-8 / 2.0 / 3.8
Tiller angle / qTA-2 / 7.2 / 8.0 / 12.4
qTA-6 / 2.2 / 10.2
qTA-9 / 1.7 / 2.2 / 3.6
Tiller number / qTN-5 / 2.3
Panicle length / qPL-2 / 4.3 / 4.2 / 7.5
qPL-6a / 3.3 / 4.4
qPL-6b / 1.8 / 1.7
qPL-12a / 2.5 / 4.7
qPL-12b / 3.2 / 6.7
Awn length / qAL-1 / 4.2 / 8.0 / 3.5
qAL-3 / 2.6 / 4.4 / 7.5
qAL-4a / 1.3 / 1.9 / 5.5
qAL-4b / 7.0 / 6.7
Grain length / qGL-1 / 2.8 / 5.9 / 7.7
qGL-3 / 2.8 / 2.1 / 5.8
qGL-6 / 3.7 / 4.3
qGL-10 / 10.9 / 11.6
Grain width / qGW-2a / 3.2 / 3.3
qGW-2b / 1.9
qGW-3 / 5.2 / 6.8 / 6.5
qGW-5 / 1.3 / 3.0 / 3.2
qGW-6 / 2.6 / 1.7 / 3.1
Grain thickness / qGT-5 / 2.9 / 3.4
qGT-6 / 2.4 / 2.4 / 2.1
qGT-10 / 5.0 / 5.0
qGT-11 / 1.7 / 2.1
1,000-grain weight / qTGW-5a / 1.8
qTGW-5b / 3.7 / 2.8
qTGW-8 / 3.2 / 5.2 / 20.8
qTGW-10 / 1.9 / 4.2 / 14.4
Spikelet number / qSN-1a / 5.9 / 7.6 / 11.6
qSN-1b / 2.3 / 2.3
qSN-6 / 3.6 / 3.4 / 7.0
qSN-8 / 3.3 / 4.3 / 16.0

a Blank spaces represent no overlapped QTL based on 95% Confidence Interval.

Table S6. 95% confidence intervals and standard deviations of top five major QTL of sub-populations sampled from 150 RILs.

QTL / 95% Confidence Interval (cM)
50a / 100 a
qPH-1,
(candidate gene: SD1) / 1 / NA / 1.70
2 / NA / 2.80
3 / 2.90 / NA
4 / 1.70 / 2.10
5 / NA / 2.30
average / 2.30 / 2.23
SD / 0.85 / 0.46
qTA-9,
(candidate gene: TAC1) / 1 / 1.40 / 1.70
2 / 4.10 / 1.80
3 / 3.00 / 1.40
4 / 2.10 / 1.50
5 / 3.60 / 1.60
average / 2.84 / 1.60
SD / 1.10 / 0.16
qGL-3,
(candidate gene: GS3) / 1 / NA / 2.90
2 / NA / 1.30
3 / 3.10 / NA
4 / NA / 2.00
5 / NA / 1.30
average / 3.10 / 1.88
SD / - / 0.76
qGW-5,
(candidate gene: qSW5) / 1 / 2.40 / 2.60
2 / NA / 3.20
3 / NA / 1.50
4 / 2.50 / 2.40
5 / 3.10 / NA
average / 2.67 / 2.43
SD / 0.38 / 0.70
qFLW-4,
(candidate gene: Nal1) / 1 / NA / 2.20
2 / NA / 2.30
3 / NA / NA
4 / NA / NA
5 / NA / NA
average / - / 2.25
SD / - / 0.07

aPopulation size; NA, no mapped QTL was overlapped with corresponding candidate genes; SD, standard deviation.

Table S7.95% confidence intervals and standard deviations of top five major QTL of each simulated population with fixed marker position and QTL position.

QTL / 95% Confidence Interval (cM)
50a / 100a / 150a / 200a / 300a / 400a / 500a
qPH-1,
(candidate gene: SD1) / 1 / 2.00 / 1.20 / 1.40 / NA / 1.30 / 0.40 / 0.70
2 / 3.40 / 4.20 / 1.80 / NA / 1.20 / 1.60 / 0.90
3 / NA / NA / 2.90 / 1.50 / 1.50 / 0.70 / 0.70
4 / NA / NA / 2.50 / 1.90 / 1.50 / 0.50 / 0.90
5 / 4.00 / NA / NA / 2.00 / 1.10 / 0.60 / 0.70
average / 3.13 / 2.70 / 2.15 / 1.80 / 1.32 / 0.76 / 0.78
SD / 1.03 / 2.12 / 0.68 / 0.26 / 0.18 / 0.48 / 0.11
qTA-9,
(candidate gene: TAC1) / 1 / 15.7 / NA / NA / 1.20 / 0.30 / 0.20 / NA
2 / NA / 4.10 / 3.60 / 2.40 / 2.60 / 3.20 / 2.20
3 / 6.2 / NA / NA / 0.30 / 0.30 / 0.40 / NA
4 / NA / NA / 2.10 / 2.10 / 2.80 / 1.50 / 2.90
5 / NA / 3.00 / 0.50 / 1.10 / 0.60 / 1.20 / 1.30
average / 10.95 / 3.55 / 2.07 / 1.42 / 1.32 / 1.30 / 2.13
SD / 6.72 / 0.78 / 1.55 / 0.84 / 1.27 / 1.19 / 0.80
qGL-3,
(candidate gene: GS3) / 1 / NA / 2.90 / 2.70 / 2.70 / 2.90 / 2.80 / 0.9
2 / 8.70 / 2.90 / 3.10 / 1.00 / 1.00 / 1.20 / 1.00
3 / NA / NA / 2.60 / 4.00 / 2.40 / 3.70 / 3.70
4 / 3.20 / 3.70 / 3.30 / 2.50 / 2.70 / 2.60 / 2.60
5 / NA / 2.60 / 2.70 / 2.70 / 2.90 / 2.80 / 0.50
average / 5.95 / 3.03 / 2.88 / 2.58 / 2.38 / 2.62 / 1.74
SD / 3.89 / 0.47 / 0.30 / 1.07 / 0.80 / 0.90 / 1.36
qGW-5,
(candidate gene: qSW5) / 1 / 4.60 / 3.20 / 0.80 / 0.80 / 0.8 / 0.8 / 2.10
2 / NA / 4.50 / 3.40 / NA / 4.00 / 2.50 / NA
3 / 6.00 / 2.70 / 2.80 / NA / 1.60 / 0.60 / 0.80
4 / 1.60 / 1.70 / 1.70 / 1.10 / 1.00 / 1.20 / 1.10
5 / 7.00 / 4.10 / 1.90 / 1.40 / 1.40 / 2.60 / 2.30
average / 4.80 / 3.24 / 2.12 / 1.10 / 1.76 / 1.54 / 1.58
SD / 2.35 / 1.12 / 1.01 / 0.30 / 1.29 / 0.95 / 0.74
qFLW-4,
(candidate gene: Nal1) / 1 / 7.20 / 1.90 / 6.90 / 0.90 / 0.90 / 0.90 / 0.90
2 / NA / NA / NA / NA / 3.10 / 2.50 / 1.50
3 / 10.70 / NA / NA / 2.00 / 1.90 / 1.90 / 1.70
4 / NA / 4.50 / 3.60 / 2.20 / 1.20 / 2.20 / 0.90
5 / 7.90 / 1.70 / 0.90 / 0.70 / 0.70 / 0.70 / 0.70
average / 8.60 / 2.70 / 3.80 / 1.45 / 1.56 / 1.64 / 1.14
SD / 1.85 / 1.56 / 3.00 / 0.76 / 0.97 / 0.80 / 0.43

aPopulation size; NA, no mapped QTL was overlapped with corresponding candidate genes; SD, standard deviation.