Molecular Marker-Directed Development of a Novel Cytoplasmic Male Sterile Line in Rice

Molecular marker-directed development of a novel cytoplasmic male sterile line in rice

Fengfeng Fan1,2, Nengwu Li1, Jie Wang1, Xingdan Liu1,2, Jianfeng Liu2*, Yingguo Zhu1, Shaoqing Li1*

1 State Key Laboratory of Hybrid Rice; Key Laboratory for Research and Utilization of Heterosis in Indica Rice of Ministry of Agriculture; Engineering Research Center for Plant Biotechnology and Germplasm Utilization of Ministry of Education; College of Life Science, Wuhan University, Wuhan 430072, China.

2 College of Agronomy, Hunan Agricultural University, Changsha 410128, China

Corresponding author:

Jianfeng Liu

Email:

Tel: 86-0731-84638412

Fax: 86-0731-84638412

Shaoqing Li

Email: ;

Tel: 86-27-68752285

Fax: 86-27-68752095

Supplementary materials

Fig. S1 Schematic diagram for breeding of new CMS line LuofeiA

Fig. S2 PCR amplification of screened mitochondrial genome with mitotype-specific molecular markers. Mitochondrial gene atp6 was used as a control.

Fig. S3 Phylogenetic tree of Luofei A, IRRI86258 and the seven CMS lines based on the 27 MSS markers.

Table S1 Rice accessions and varieties used in this research

Rice types / Accessions/ Varieties / Species / Source country
Gametophytic CMS line / Yuetai A / O. sativa / China
Sporophytic CMS lines / Maxie A / O. sativa / China
Ⅱ-32A / O. sativa / China
K17A / O. sativa / China
Yue4 A / O. sativa / China
Xieqingzao A / O. sativa / China
Gang46 A / O. sativa / China
Zhenshan97 A / O. sativa / China
Rongfeng A / O. sativa / China
Zaofeng A / O. sativa / China
Wufeng A / O. sativa / China
Xie A / O. sativa / China
Sporophytic maintainer
lines / Maxie B / O. sativa / China
Ⅱ-32B / O. sativa / China
K17B / O. sativa / China
Xieqingzao B / O. sativa / China
Gang46 B / O. sativa / China
Zhenshan97 B / O. sativa / China
Yue4 B / O. sativa / China
Landrace
accessions / 103940 / O. glaberrima / Liberia
96717 / O. glaberrima / Senegal
103601 / O. glaberrima / Cameroon
103922 / O. glaberrima / Nigeria
103946 / O. glaberrima / Liberia
102928 / O. glaberrima / Burkina Faso
103937 / O. glaberrima / Liberia
100127 / O. glaberrima / Gambia
104507 / O. glaberrima / Liberia
103593 / O. glaberrima / Cameroon
86258 / O. glaberrima / Africa
100956 / O. latifolia / India
105167 / O. latifolia / Indonesia
81967 / O. longistaminata / Botswana
104152 / O. longistaminata / Cameroon
104153 / O. longistaminata / Cameroon
104302 / O. longistaminata / Malawi
105203 / O. longistaminata / Ethiopia
101097 / O. minuta / Philippines
106487 / O. minuta / Philippines
86467 / O. nivara / India
89171 / O. nivara / Kampuchea
89176 / O. nivara / Kampuchea
89179 / O. nivara / Kampuchea
89180 / O. nivara / Kampuchea
89182 / O. nivara / Kampuchea
100951 / O. nivara / India
102179 / O. nivara / India
103419 / O. nivara / Sri Lanka
93272 / O. officinalis / Indonesia
100878 / O. officinalis / Thailand
100973 / O. officinalis / Philippines
105106 / O. officinalis / Malaysia
88827 / O. punctata / Madagascar
88828 / O. punctata / Madagascar
100881 / O. punctata / Sri Lanka
104059 / O. punctata / Nigeria
105153 / O. punctata / Nigeria
105154 / O. punctata / Nigeria
86466 / O. rhizomatis / Sri Lanka
103414 / O. rhizomatis / Sri Lanka
103417 / O. rhizomatis / Sri Lanka
80528 / O. rufipogon / India
80608 / O. rufipogon / India
81890 / O. rufipogon / India
82988 / O. rufipogon / China
82990 / O. rufipogon / China
86486 / O. rufipogon / Thailand
86557 / O. rufipogon / Kampuchea
88788 / O. rufipogon / Bangladesh
88817 / O. rufipogon / Laos
88820 / O. rufipogon / Thailand
89030 / O. rufipogon / Kampuchea
93040 / O. rufipogon / Kampuchea
93043 / O. rufipogon / Kampuchea
93080 / O. rufipogon / Kampuchea
93123 / O. rufipogon / Kampuchea
93125 / O. rufipogon / Kampuchea
93215 / O. rufipogon / Nepal
93288 / O. rufipogon / Nepal
99555 / O. rufipogon / China
Early rice varieties or lines / Ezaoxuan / O. sativa / China
Xiangzaoxian7 / O. sativa / China
Jiangnongzao1 / O. sativa / China
Yingyouzao1 / O. sativa / China
Yingyouzao2 / O. sativa / China
Ganzaoxian24 / O. sativa / China
Ganzaoxian30 / O. sativa / China
Ganzaoxian40 / O. sativa / China
Ganzaoxian58 / O. sativa / China
Jiayu164 / O. sativa / China
Xiangzaoxian3 / O. sativa / China
Xiangzaoxian5 / O. sativa / China
Xiangzaoxian13 / O. sativa / China
Xiangzaoxian15 / O. sativa / China
Xiangzaoxian17 / O. sativa / China
Xiangzaoxian18 / O. sativa / China
Xiangzaoxian20 / O. sativa / China
Xiangzaoxian29 / O. sativa / China
Xiangzaoxian30 / O. sativa / China
Xiangzaoxian31 / O. sativa / China
CE14002 / O. sativa / China
CE14006 / O. sativa / China
CE14008 / O. sativa / China
CE14009 / O. sativa / China
CE14010 / O. sativa / China
CE14012 / O. sativa / China
Yingyouzao3 / O. sativa / China
Youfengzao / O. sativa / China
Zhenfengzao / O. sativa / China
Ganzaoxian24 / O. sativa / China
Xiangyouzao / O. sativa / China
Ganzaoxian37 / O. sativa / China
Xiangzaoxian22 / O. sativa / China
Xiangzaoxian24 / O. sativa / China
Xiangzaoxian27 / O. sativa / China
Xiangzaoxian28 / O. sativa / China
CE14014 / O. sativa / China
CE14019 / O. sativa / China
CE14023 / O. sativa / China
CE14027 / O. sativa / China
CE14028 / O. sativa / China
CE14030 / O. sativa / China
CE14039 / O. sativa / China
CE14047 / O. sativa / China
CE14050 / O. sativa / China
CE14091 / O. sativa / China
CE14092 / O. sativa / China
CE14093 / O. sativa / China
CE14096 / O. sativa / China
CE14103 / O. sativa / China
CE14104 / O. sativa / China
CE14107 / O. sativa / China
CE14124 / O. sativa / China
CE14131 / O. sativa / China
CE14134 / O. sativa / China
CE14145 / O. sativa / China
CE14159 / O. sativa / China
CE14170 / O. sativa / China
CE14176 / O. sativa / China
CE14177 / O. sativa / China
CE14195 / O. sativa / China
CE14201 / O. sativa / China
Sporophytic restorer lines / Zhi2 / O. sativa / China
Zhi29 / O. sativa / China
Zhi31 / O. sativa / China
Zhi32 / O. sativa / China
Zhi5 / O. sativa / China
Zhi6 / O. sativa / China
Zhi7 / O. sativa / China
Minghui63 / O. sativa / China
Hybrid rice / Tianyou-huazan / O. sativa / China

Table S2 Primers related to CMS and restorer genes used in this study

Primers / Primer sequences / Annealing temperature (oC) / Reference
orf(H)79 / F:GAGGAGCCGAAGATTTTAG / 57 / Li et al. 2007
R:TTACTTAGGAAAGACTACAC
orf352 / F:ATGACGAGAGATAGAATGAG / 50 / Luo et al. 2013
R:CTAGGAGGCTGAGTTTTG
RM490 / F:ATCTGCACACTGCAAACACC / 55 / Majid et al. 2008
R:AGCAAGCAGTGCTTTCAGAG
RM6100 / F:TCCTCTACCAGTACCGCACC / 55 / Ahmadikhah and karlov. 2006
R:GCTGGATCACAGATCATTGC

Table S3 Agronomic traits of F1 hybrid combinations derived from Luofei A and other four sporophytic CMS lines

Combinations / Plant height (cm) / Effective panicle / Grain number/panicle / Seed setting rate (%) / 1000-grain weight (g) / Grain yield/ plant (g)
XieA/Zhi2 / 106.6±2.3 / 10.8±1.3 / 171±8 / 85.1±2.6 / 25.5±0.9 / 39.9±4.6
XieA/Zhi29 / 119.2±1.9 / 9.4±1.1 / 202±21 / 90.6±0.9 / 26.0±1.1 / 44.2±1.1
XieA/Zhi31 / 115.6±2.3 / 9.4±1.7 / 200±22 / 91.0±2.0 / 24.7±0.3 / 42.2±8.7
XieA/Zhi32 / 123.6±3.8 / 8.0±0.7 / 240±22 / 82.9±3.2 / 26.7±0.5 / 42.6±5.8
XieA/Zhi5 / 120.2±2.6 / 9.2±1.1 / 232±29 / 75.3±4.2 / 24.0±0.7 / 38.2±3.3
XieA/Zhi6 / 113.2±2.5 / 8.0±0.7 / 228±17 / 78.3±3.3 / 26.1±0.8 / 37.0±3.1
XieA/Zhi7 / 114.2±2.4 / 9.6±1.1 / 202±19 / 79.2±1.5 / 25.6±0.9 / 39.0±3.8
Luofei A/Zhi2 / 116.4±3.9 / 11.2±1.6 / 185±24 / 67.9±4.9 / 27.2±0.9 / 37.9±5.0
Luofei A/Zhi29 / 122.6±2.8 / 8.8±1.3 / 232±33 / 86.6±1.6 / 28.7±0.5 / 50.2±7.1
Luofei A/Zhi31 / 119.0±1.4 / 10.6±1.5 / 201±23 / 83.4±2.0 / 25.1±0.6 / 44.4±5.8
Luofei A/Zhi32 / 133.0±3.4 / 8.0±1.0 / 250±30 / 77.5±2.4 / 27.7±0.7 / 41.4±5.6
Luofei A/Zhi5 / 126.0±1.6 / 9.0±1.9 / 229±24 / 71.8±0.3 / 26.0±0.8 / 38.9±3.1
Luofei A/Zhi6 / 127.6±2.7 / 8.4±1.1 / 234±21 / 79.3±6.6 / 26.2±0.8 / 40.5±5.0
Luofei A/Zhi7 / 122.0±3.0 / 8.6±0.9 / 212±11 / 81.5±4.8 / 26.9±1.3 / 40.0±5.8
RongfengA/Zhi2 / 106.4±2.2 / 12.4±1.8 / 138±16 / 78.4±1.9 / 26.6±0.9 / 35.7±6.9
RongfengA/Zhi29 / 116.4±1.1 / 8.8±0.8 / 189±12 / 91.5±0.8 / 28.7±0.5 / 43.8±7.1
RongfengA/Zhi31 / 112.6±2.9 / 12.8±1.8 / 159±15 / 91.8±0.7 / 25.5±0.3 / 47.2±3.5
RongfengA/Zhi32 / 115.2±3.3 / 8.4±1.3 / 240±23 / 86.0±3.4 / 28.9±0.4 / 49.6±5.3
RongfengA/Zhi5 / 120.2±0.8 / 9.0±0.7 / 213±10 / 83.8±0.7 / 26.7±0.3 / 42.9±4.0
RongfengA/Zhi6 / 111.2±4.0 / 8.6±1.3 / 191±9 / 86.0±1.9 / 27.6±0.7 / 38.9±5.2
RongfengA/Zhi7 / 115.0±3.0 / 8.6±0.9 / 182±27 / 88.1±3.3 / 28.2±0.4 / 38.7±6.6
WufengA/Zhi2 / 104.0±1.4 / 10.2±1.1 / 155±32 / 77.0±4.0 / 24.8±0.5 / 29.9±6.0
WufengA/Zhi29 / 114.2±1.3 / 8.8±1.5 / 186±22 / 94.1±0.7 / 26.2±0.5 / 40.1±5.3
WufengA/Zhi31 / 111.2±1.6 / 10.8±1.8 / 183±19 / 90.3±3.0 / 24.6±0.5 / 43.5±7.6
WufengA/Zhi32 / 117.0±3.4 / 8.0±1.4 / 219±42 / 85.4±2.9 / 25.4±0.9 / 37.4±6.2
WufengA/Zhi5 / 117.6±3.6 / 9.2±1.1 / 196±25 / 78.8±3.4 / 25.2±0.7 / 37.0±6.6
WufengA/Zhi6 / 107.4±2.9 / 9.0±0.7 / 212±22 / 88.6±2.1 / 25.9±0.4 / 43.6±3.6
WufengA/Zhi7 / 112.0±4.4 / 8.8±1.5 / 206±19 / 81.1±3.2 / 25.9±1.2 / 37.7±3.2
ZaofengA/Zhi2 / 102.2±2.4 / 11.0±2.0 / 152±15 / 74.7±4.2 / 25.4±0.5 / 31.8±7.8
ZaofengA/Zhi29 / 108.4±2.1 / 9.4±0.9 / 184±21 / 91.8±1.9 / 27.1±0.6 / 42.8±3.0
ZaofengA/Zhi31 / 107.4±2.3 / 9.6±0.9 / 175±12 / 89.4±4.2 / 24.8±0.6 / 37.1±4.2
ZaofengA/Zhi32 / 110.6±2.2 / 10.0±1.0 / 223±26 / 83.9±4.1 / 24.5±1.9 / 44.4±3.6
ZaofengA/Zhi5 / 115.6±4.6 / 9.2±1.5 / 221±17 / 83.8±4.2 / 26.2±0.9 / 44.0±3.6
ZaofengA/Zhi6 / 103.4±2.4 / 9.4±0.9 / 211±18 / 81.0±5.5 / 26.1±1.2 / 42.1±7.1
ZaofengA/Zhi7 / 110.0±2.4 / 9.6±1.1 / 168±9 / 90.2±2.4 / 27.5±0.8 / 39.9±3.9
Tianyou-huazhan / 113.0±3.4 / 10.2±1.6 / 205±19 / 85.0±3.0 / 24.3±0.6 / 43.0±6.6

Table S4 Variance on agronomic traits of the hybrids derived from five CMS lines for combining ability (F value)

Source of variation / Plant height / Effective
panicle / Grain No.
/panicle / Seed set rate / 1,000-grain weight / Grain yield/ plant
Block / 1.37 / 1.61 / 1.57 / 0.38 / 1.02 / 0.32
Combination / 14.54** / 4.78** / 8.20** / 10.42** / 9.99** / 3.34**
P1 / 31.48** / 1.17 / 11.72** / 4.08* / 6.90** / 2.09
P2 / 12.86** / 9.26** / 25.33** / 8.47** / 4.46** / 5.50**
P1×P2 / 2.26** / 1.96* / 1.35 / 3.65** / 4.80** / 1.73*
Error / 18.76 / 1.68 / 469.70 / 0.002 / 0.79 / 30.06

Note: *and **Significant at 0.05 and 0.01 probability level, respectively. P1 including CMS lines Rongfeng A, Loufei A, Xie A, Zaofeng A and Wufeng A. P2 including restorer lines Zhi2, Zhi29, Zhi31, Zhi32, Zhi5, Zhi6, Zhi7. “Error” are sum of squares of the traits.