Chinese Bulletin of Botany ›› 2022, Vol. 57 ›› Issue (1): 42-55.DOI: 10.11983/CBB21158
• RESEARCH PAPER • Previous Articles Next Articles
Xia Wang1, Wei Yan1, Zhiqin Zhou2, Zhenyi Chang1, Minting Zheng1, Xiaoyan Tang1,2, Jianxin Wu1,*()
Received:
2021-09-14
Accepted:
2021-11-17
Online:
2022-01-01
Published:
2022-01-17
Contact:
Jianxin Wu
Xia Wang, Wei Yan, Zhiqin Zhou, Zhenyi Chang, Minting Zheng, Xiaoyan Tang, Jianxin Wu. Identification and Mapping of a Rice Male Sterility Mutant ms102[J]. Chinese Bulletin of Botany, 2022, 57(1): 42-55.
Primers | Sequences (5′-3′) |
---|---|
OsACTIN-qPCR-F | GCTATGTACGTCGCCATCCA |
OsACTIN-qPCR-R | GGACAGTGTGGCTGACACCAT |
RTS-qPCR-F | ACATGTGGACTCGCTTGACT |
RTS-qPCR-R | CATGGCTGCATGCAGATTCAT |
TDR-qPCR-F | TGCTCTGGGAGCACAAGCC |
TDR-qPCR-R | CTCGCTGTCCCTCACCATG |
UDT1-qPCR-F | GAAGCACTCTGCAGCTAC |
UDT1-qPCR-R | CTGCGTAGCCGTAGAAGG |
DTM1-qPCR-F | AATTTAAGCCCTATCCCCTAAG |
DTM1-qPCR-R | GAAGGTGGATTCCACTAGCTA |
PTC1-qPCR-F | CACCAGATCATGGACCTCTG |
PTC1-qPCR-R | AGCAGCCTCAGCTCCATGTG |
DPW-qPCR-F | ACCACAGGAGACACGGTGATG |
DPW-qPCR-R | CTTGGCCTGATGGTGACAA |
PAIR1-qPCR-F | GGATGGACCCAGATTAACC |
PAIR1-qPCR-R | CTGTTTAGGTGCCACCCTGT |
PAIR2-qPCR-F | TGCCAGAGGAGAGGACCATTC |
PAIR2-qPCR-R | CACGAGATGCTTGCTATTGAC |
DTD-qPCR-F | TCATGAGTCTTTGAGCCGCA |
DTD-qPCR-R | CACCTTCGCTCCAAATCTGT |
OsCP1-qPCR-F | TGCAAGGATCACTCCACCTG |
OsCP1-qPCR-R | GGCTGTTCTTGCTCTTGGAG |
OsMSP1-qPCR-F | AGCATCGCGAGTAAGATGC |
OsMSP1-qPCR-R | GTAGCCTTGTAACTTCAAGTAGGA |
CYP703A3-qPCR-F | TGTACTGCTTTCTTTGCCCG |
CYP703A3-qPCR-R | GCAGCAATCATGTCCTGCAT |
CYP704B2-qPCR-F | GCTGGTTGATGACTTCACCT |
CYP704B2-qPCR-R | CGACAGTATGTCGTGCTTGAT |
OsGAMYB-qPCR-F | GCGACGGTATCATGTTCAAT |
OsGAMYB-qPCR-R | GTCGCATAAGAGAACATCTG |
Table 1 Sequences of primers for qRT-PCR
Primers | Sequences (5′-3′) |
---|---|
OsACTIN-qPCR-F | GCTATGTACGTCGCCATCCA |
OsACTIN-qPCR-R | GGACAGTGTGGCTGACACCAT |
RTS-qPCR-F | ACATGTGGACTCGCTTGACT |
RTS-qPCR-R | CATGGCTGCATGCAGATTCAT |
TDR-qPCR-F | TGCTCTGGGAGCACAAGCC |
TDR-qPCR-R | CTCGCTGTCCCTCACCATG |
UDT1-qPCR-F | GAAGCACTCTGCAGCTAC |
UDT1-qPCR-R | CTGCGTAGCCGTAGAAGG |
DTM1-qPCR-F | AATTTAAGCCCTATCCCCTAAG |
DTM1-qPCR-R | GAAGGTGGATTCCACTAGCTA |
PTC1-qPCR-F | CACCAGATCATGGACCTCTG |
PTC1-qPCR-R | AGCAGCCTCAGCTCCATGTG |
DPW-qPCR-F | ACCACAGGAGACACGGTGATG |
DPW-qPCR-R | CTTGGCCTGATGGTGACAA |
PAIR1-qPCR-F | GGATGGACCCAGATTAACC |
PAIR1-qPCR-R | CTGTTTAGGTGCCACCCTGT |
PAIR2-qPCR-F | TGCCAGAGGAGAGGACCATTC |
PAIR2-qPCR-R | CACGAGATGCTTGCTATTGAC |
DTD-qPCR-F | TCATGAGTCTTTGAGCCGCA |
DTD-qPCR-R | CACCTTCGCTCCAAATCTGT |
OsCP1-qPCR-F | TGCAAGGATCACTCCACCTG |
OsCP1-qPCR-R | GGCTGTTCTTGCTCTTGGAG |
OsMSP1-qPCR-F | AGCATCGCGAGTAAGATGC |
OsMSP1-qPCR-R | GTAGCCTTGTAACTTCAAGTAGGA |
CYP703A3-qPCR-F | TGTACTGCTTTCTTTGCCCG |
CYP703A3-qPCR-R | GCAGCAATCATGTCCTGCAT |
CYP704B2-qPCR-F | GCTGGTTGATGACTTCACCT |
CYP704B2-qPCR-R | CGACAGTATGTCGTGCTTGAT |
OsGAMYB-qPCR-F | GCGACGGTATCATGTTCAAT |
OsGAMYB-qPCR-R | GTCGCATAAGAGAACATCTG |
Primers | Sequences (5'-3') |
---|---|
S1-HRM-F | GACAGGCAAGCAGAAAACCTG |
S1-HRM-R | GTGCAGGTAGCACATCATCCT |
S2-HRM-F | CTACAGATATATACAGGACGGTG |
S2-HRM-R | GCTAAGGTAGCAACGTACACAG |
S3-HRM-F | GATGGAGACTCACGTCCAGA |
S3-HRM-R | CCACAGCCCTCATTTTCGT |
Table 2 Sequences of primers for high resolution melting (HRM)
Primers | Sequences (5'-3') |
---|---|
S1-HRM-F | GACAGGCAAGCAGAAAACCTG |
S1-HRM-R | GTGCAGGTAGCACATCATCCT |
S2-HRM-F | CTACAGATATATACAGGACGGTG |
S2-HRM-R | GCTAAGGTAGCAACGTACACAG |
S3-HRM-F | GATGGAGACTCACGTCCAGA |
S3-HRM-R | CCACAGCCCTCATTTTCGT |
Figure 1 Phenotype analysis of the rice ms102 (A) Phenotype of mature plants (bar=9 cm); (B) Tiller number of mature wild type (WT) and ms102 plant (n=23); (C) Leaf color of WT and ms102 plant at flowering (bar=1 cm); (D) Mature spikelet without palea and lemma (bar=1 mm); (E) Flowering spikelet, the arrow indicates a dehisced anther (bar=1 cm); (F), (G) I2-KI-stained pollen of WT and ms102 mutant (bars=50 μm); (H) Mature panicles (bar=2.4 cm)
Fertile plants | Male sterility plants | χ2(3:1) | χ20.05 |
---|---|---|---|
359 | 102 | 2.034 | 3.841 |
Table 3 Segregation of the F2 progeny of rice WT × ms102
Fertile plants | Male sterility plants | χ2(3:1) | χ20.05 |
---|---|---|---|
359 | 102 | 2.034 | 3.841 |
Figure 2 Transverse sections images of WT and ms102 anthers at the developmental stages 7-12 BMs: Binuclear microspores; DMs: Degenerated microspores; E: Epidermis; En: Endothecium; M: Middle layer; Mp: Mature pollen; Ms: Microspores; PMC: Pollen mother cell; ST: Swollen tapetum; T: Tapetum; Tds: Tetrads. Bars=10 μm
Figure 3 Transmission electron microscope images of WT and ms102 anthers at the developmental stages 9-12 (A), (D) Stage 9; (B), (E) Stage 10 (the arrows indicate the microspore wall); (C), (F) Stage 12; (G), (H) The magnified region of dashed box in B and E (the arrows indicate the Ubisch bodies); T: Tapetum; Ms: Microspores; C: Cuticle. (A), (B), (D), (E) Bars=2 μm; (C), (F), (G), (H) Bars=1 μm
Figure 4 Scanning electron microscope images of WT and ms102 anthers at developmental stage 12 (A), (C), (E), (G) The external surfaces of anther wall; (B), (F) Pollen grains; (D), (H) Ubisch bodies. (A), (E) Bars =100 μm; (B), (C), (F), (G) Bars=10 μm; (D), (H) Bars=2 μm
Figure 5 Expression of genes related to pollen development in WT and ms102 anthers Anthers at stages 7-12 were used for qRT-PCR. OsACTIN1 served as an internal control. Data are shown as means±SD (n=3). * P<0.05, ** P<0.01, *** P<0.001
Chr. locus | Physical location | Genotype | SNP index | Gene |
---|---|---|---|---|
Chr. 1 | 39,147,186 (S1) | C→T | 0.8966 | Intergenic |
Chr. 1 | 39,167,971 (S2) | G→A | 0.8966 | Intergenic |
Chr. 1 | 40,599,831 (S3) | C→T | 0.9474 | LOC_Os01g7-0140-LOC_O-s01g70150 |
Table 4 Candidate genes and annotation information
Chr. locus | Physical location | Genotype | SNP index | Gene |
---|---|---|---|---|
Chr. 1 | 39,147,186 (S1) | C→T | 0.8966 | Intergenic |
Chr. 1 | 39,167,971 (S2) | G→A | 0.8966 | Intergenic |
Chr. 1 | 40,599,831 (S3) | C→T | 0.9474 | LOC_Os01g7-0140-LOC_O-s01g70150 |
The number of male sterile plants | The number of fertile plants | |||||
---|---|---|---|---|---|---|
Physical location | S1 | S2 | S3 | S1 | S2 | S3 |
Wild type | 1 | 1 | 0 | 25 | 25 | 23 |
Heterozygote | 8 | 8 | 0 | 19 | 19 | 21 |
Mutant | 44 | 44 | 53 | 0 | 0 | 0 |
Table 5 Genotype-phenotype correlation analyses of the three candidate SNP sites
The number of male sterile plants | The number of fertile plants | |||||
---|---|---|---|---|---|---|
Physical location | S1 | S2 | S3 | S1 | S2 | S3 |
Wild type | 1 | 1 | 0 | 25 | 25 | 23 |
Heterozygote | 8 | 8 | 0 | 19 | 19 | 21 |
Mutant | 44 | 44 | 53 | 0 | 0 | 0 |
Figure 6 Gene structures of LOC_Os01g70140 and LOC_Os01g70150 and mutant phenotypes (A) The gene structures and mutation sites of LOC_Os01g70140 and LOC_Os01g70150; (B)-(D) Morphology of mature anther and pistil (bars=1 mm); (E)-(G) Mature pollen grains stained with I2-KI (bars=100 μm).
Figure 8 The phenotype of the F1 progeny of dpw2/+ × ms102 The phenotype of the F1 progeny of dpw2/+ × ms102 (A)-(D) Morphology of mature anther and pistil (bars=1 mm); (E)-(H) Mature pollen grains stained with I2-KI (bars=100 μm).
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