Molecular Markers and Candidate Genes for Thermo-Sensitive Genic Male Sterile in Rice

doi:10.1016/j.rsci.2018.08.006

Abstract

Abstract:

The discovery of thermo-sensitive genic male sterility (TGMS) has led to development of a simple and highly efficient two-line breeding system. In this study, genetic analysis was conducted using three F₂ populations derived from crosses between IR68301S, an indica TGMS rice line, and IR14632 (tropical japonica), Supanburi 91062 (indica) and IR67966-188-2-2-1 (tropical japonica), respectively. Approximately 1:3 ratio between sterile and normal pollen of F₂ plants from the three populations revealed that TGMS is controlled by a single recessive gene. Bulked segregant analysis using simple sequence repeat (SSR) and insertion-deletion (InDel) markers were used to identify markers linked to the tms gene. The linkage analysis based on the three populations indicated that the tms locus was located on chromosome 2 covering the same area. Using IR68301S × IR14632 F₂ population, the results showed that the tms locus was located between SSR marker RM12676 and InDel marker 2gAP0050058. The genetic distance from the tms gene to these two flanking markers were 1.10 and 0.82 cM, respectively. InDel marker 2gAP004045 located between these two markers showed complete co-segregation with the TGMS phenotype. In addition, InDel marker vf0206114052 showed 2.94 cM linked to the tms gene using F₂ populations of IR68301S × Supanburi 91062. These markers are useful tool for developing new TGMS lines by marker-assisted selection. There were ten genes located between the two flanking markers RM12676 and 2gAP0050058. Using quantitative real-time PCR for expression analysis, 7 of the 10 genes showed expression in panicles, and response to temperatures. These genes could be the candidate gene controlling TGMS in IR68301S.

Key words: hybrid rice, thermo-sensitive genic male sterility, insertion/deletion, simple sequence repeat, marker-assisted selection

Khlaimongkhon Sudthana, Chakhonkaen Sriprapai, Pitngam Keasinee, Ditthab Khanittha, Sangarwut Numphet, Panyawut Natjaree, Wasinanon Thiwawan, Mongkolsiriwatana Chareerat, Chunwongse Julapark, Muangprom Amorntip. Molecular Markers and Candidate Genes for Thermo-Sensitive Genic Male Sterile in Rice[J]. Rice Science, 2019, 26(3): 147-156.

Figures/Tables 8

References 53

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F₂ population	Total No. of lines	No. of fertile individuals	No. of sterile individuals	Segregation rate	χ²
B2 × B30	917	700	217	3.22	0.91
B2 × B11	981	722	259	2.79	0.98
B2 × KM9	1 784	1 334	450	2.96	0.042

F₂ population	Total No. of lines	No. of fertile individuals	No. of sterile individuals	Segregation rate	χ²
B2 × B30	917	700	217	3.22	0.91
B2 × B11	981	722	259	2.79	0.98
B2 × KM9	1 784	1 334	450	2.96	0.042

Marker	Forward primers (5´-3´)	Reverse primers (5´-3´)	Position (Mb)
RM12557	AGCACCACCTCCTCGAACTCC	CAACCCTACCTTGCTTCTTCTTGC	4.19
RM4355	GGGATGAGAGTAGAAGGCACAAGG	GCTTAATGCCTTTGATCGTTGC	4.26
RM12593	ACAATACGCTGTGCCAATCTGC	TATTGGCCGTGATGATGAAGTGC	4.57
RM12601	GCGAACTCGACGACTACTCAACC	CACAACGTCGTCTCCAAGTGC	4.69
RM7638	GGCTGTCCGTCTTGTAGTGAGAAGC	TCCTTGCAACTTCCGGAGACC	4.86
RM12632	GGCTTTATTTGTTCTGGCCTTGTGG	CCAAATCAAATGGGTCCTTTGTGC	5.08
RM12649	TTGAGGTAGGGCACGAGGTCTGG	AATCCGAATGGGACCGAATCACC	5.25
RM12655	AGGACCAATCCAAAGCGTTTAGC	CCTGCAGTAGATTGCATTGAACC	5.34
RM5664	GTTCGGCTCCACCTAAACCAAGC	GGCATTCGTCTCGTCTTTGAGG	5.37
RM6378	CTGATCATCTCATGCCTCCTACG	TCCATCTCCCAATATGACCAACC	5.47
2gAP004869	AGCTTAAACATATTGGCTGCACG	TAGCCAAGCACGTATACACTCCC	5.67
RM12674	TAAATGCCAACCAACTCCAAGC	AACTGCGTTTGGGAATATCTCG	5.68
RM12676	ACTGACGATTGGGCACATTATTCC	CTGCAAATTGGTGGGTGATTGC	5.74
2gAP004045	CCGAAACCATATATCCAACACGA	TCAAATGTTATTTGGGCGTAACTTT	5.76
2gAP0050058	CAAACAACCACTTTTGTTCAGCA	ATATATGCTGTGGGCAGGATCAA	5.81
2gAP003974	CAATTCGGAGATTACATTCGGTC	ACTATAGACGCAGCTTGTGTGCC	6 .0
vf0206114052	CGAGACCAATCGATCCTCCA	TAACCTGCATCTGTGGTGGA	6.11
2gAP004085	TGGCAACTAATAGGCATCGTCG	TCTTGTAGCTGCAACATCCCCC	6.26
2gAP004086	TCAAGCAATCACAACCCCTTCC	GGCTACCCAAACCATGCATCA	6.28
Os02g12300	TCCCATTTAATTTGGTTCATCG	GCTAACGTGCGTGGATTGAG	6.4
Os02g12350	TCCATCTGCAAATCCATAGCA	TCTGGTGCATAGCTGCTGGT	6.42
Os02g12370	ACTTATGCGGTGGTCAAGCA	TTGTTCCTCCACGGCTGATA	6.44
2gAP005394	TGCAAACAGTTGGATTAGCTTGA	TGACAGTGCGCTTATCTTCCATA	7.17
2gAP005756	AATTCATTGGACGCACACACAT	TGCATTGGGAAATTGAAAAGATT	7.26
2gAP004070	ATGGTGCCGGATAACGTATTACA	GGCTTGCGGTAAGAATATAGGAGA	7.44

Marker	Forward primers (5´-3´)	Reverse primers (5´-3´)	Position (Mb)
RM12557	AGCACCACCTCCTCGAACTCC	CAACCCTACCTTGCTTCTTCTTGC	4.19
RM4355	GGGATGAGAGTAGAAGGCACAAGG	GCTTAATGCCTTTGATCGTTGC	4.26
RM12593	ACAATACGCTGTGCCAATCTGC	TATTGGCCGTGATGATGAAGTGC	4.57
RM12601	GCGAACTCGACGACTACTCAACC	CACAACGTCGTCTCCAAGTGC	4.69
RM7638	GGCTGTCCGTCTTGTAGTGAGAAGC	TCCTTGCAACTTCCGGAGACC	4.86
RM12632	GGCTTTATTTGTTCTGGCCTTGTGG	CCAAATCAAATGGGTCCTTTGTGC	5.08
RM12649	TTGAGGTAGGGCACGAGGTCTGG	AATCCGAATGGGACCGAATCACC	5.25
RM12655	AGGACCAATCCAAAGCGTTTAGC	CCTGCAGTAGATTGCATTGAACC	5.34
RM5664	GTTCGGCTCCACCTAAACCAAGC	GGCATTCGTCTCGTCTTTGAGG	5.37
RM6378	CTGATCATCTCATGCCTCCTACG	TCCATCTCCCAATATGACCAACC	5.47
2gAP004869	AGCTTAAACATATTGGCTGCACG	TAGCCAAGCACGTATACACTCCC	5.67
RM12674	TAAATGCCAACCAACTCCAAGC	AACTGCGTTTGGGAATATCTCG	5.68
RM12676	ACTGACGATTGGGCACATTATTCC	CTGCAAATTGGTGGGTGATTGC	5.74
2gAP004045	CCGAAACCATATATCCAACACGA	TCAAATGTTATTTGGGCGTAACTTT	5.76
2gAP0050058	CAAACAACCACTTTTGTTCAGCA	ATATATGCTGTGGGCAGGATCAA	5.81
2gAP003974	CAATTCGGAGATTACATTCGGTC	ACTATAGACGCAGCTTGTGTGCC	6 .0
vf0206114052	CGAGACCAATCGATCCTCCA	TAACCTGCATCTGTGGTGGA	6.11
2gAP004085	TGGCAACTAATAGGCATCGTCG	TCTTGTAGCTGCAACATCCCCC	6.26
2gAP004086	TCAAGCAATCACAACCCCTTCC	GGCTACCCAAACCATGCATCA	6.28
Os02g12300	TCCCATTTAATTTGGTTCATCG	GCTAACGTGCGTGGATTGAG	6.4
Os02g12350	TCCATCTGCAAATCCATAGCA	TCTGGTGCATAGCTGCTGGT	6.42
Os02g12370	ACTTATGCGGTGGTCAAGCA	TTGTTCCTCCACGGCTGATA	6.44
2gAP005394	TGCAAACAGTTGGATTAGCTTGA	TGACAGTGCGCTTATCTTCCATA	7.17
2gAP005756	AATTCATTGGACGCACACACAT	TGCATTGGGAAATTGAAAAGATT	7.26
2gAP004070	ATGGTGCCGGATAACGTATTACA	GGCTTGCGGTAAGAATATAGGAGA	7.44

Gene	Description
Os02g0202500	Conserved hypothetical protein
Os02g0202600	Tetratricopeptide-like helical domain containing protein
Os02g0202800	Zinc ion binding protein
Os02g0202900	OsFBK3-F-box domain and kelch repeat containing protein
Os02g0202950	Concanavalin A-like lectin/glucanases superfamily and Legume lectin domain
Os02g0203000	Carbohydrate binding, sequence-specific DNA binding, sequence-specific DNA binding transcription factor activity
Os02g0203300	UDP-glucuronosyl/UDP-glucosyltransferase
Os02g0203401	hypothetical gene, protein coding
Os02g0203500	Putative uncharacterized protein OJ1135_F06.25; Putative uncharacterized protein P0544H11.7
Os02g0203700	Zinc finger protein; Zinc finger transcription factor ZFP30