Chinese Journal OF Rice Science ›› 2018, Vol. 32 ›› Issue (1): 12-22.DOI: 10.16819/j.1001-7216.2018.7055
• Orginal Article • Previous Articles Next Articles
Xiaoyun LIU, Xiao LI, Tengfei LI, Lufang SU
Received:
2017-05-18
Revised:
2017-09-15
Online:
2018-01-10
Published:
2018-01-10
刘小云, 李晓, 李腾飞, 苏鲁方
基金资助:
CLC Number:
Xiaoyun LIU, Xiao LI, Tengfei LI, Lufang SU. Overexpression of OsENO2-2 Affects Heading Date in Rice[J]. Chinese Journal OF Rice Science, 2018, 32(1): 12-22.
刘小云, 李晓, 李腾飞, 苏鲁方. 水稻OsENO2-2基因过表达对水稻抽穗期的影响[J]. 中国水稻科学, 2018, 32(1): 12-22.
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URL: http://www.ricesci.cn/EN/10.16819/j.1001-7216.2018.7055
Fig. 1. Construction of overexpression vector and RNAi vector. A, Structure of overexpression vector; LB, Left border of T-DNA; Hn, Hygromycin; Pubiqutin, Promoter of ubiqutin; GUS, beta-glucuronidase; RB, Right border of T-DNA. B, Structure of RNAi vector. 35S, 35S promoter of CaMV; R-f, RNAi fragment of OsENO2-2.
引物名称 Primer name | 引物序列(5′-3′) Primer sequence(5′-3′) |
---|---|
OsENO2-2-F | GGGGGTACCATGGTTCAACAGCTTGATGGAA |
OsENO2-2-R | GGGGGTACCTTAGTACGGCTCCACAGGGG |
R-OsENO2-2-F | AGAACTAGTGGTACCTGCTTCCACCGGAATATATGAG |
R-OsENO2-2-R | AGAGAGCTCGGATCCCCCTCTTTGTTTTCCTGAATGTTA |
Hn-F | TACACAGCCATCGGTCCAGA |
Hn-R | TAGGAGGGCGTGGATATGTC |
OsENO2-2-QF | GGCCAAGATGCCACAAATGT |
OsENO2-2-QR | TTGCCTGTGTAGCCAGCCTTA |
Actin-QF | TGTATGCCAGTGGTCGTACCA |
Actin-QR | CCAGCAAGGTCGAGACGAA |
RFT1-QF | TGGTGTTCGTGCTGTTCCA |
RFT1-QR | TTGTAGAGCTCGGCGAAGTTC |
RID1-QF | CGACGACAATAGCTCGATCGC |
RID1-QR | GTGCATGGTCACGGAGCCTT |
Hd3a-QF | GCTCACTATCATCATCCAGCATG |
Hd3a-QR | CCTTGCTCAGCTATTTAATTGCATAA |
Hd1-QF | TCAG CAACAGCATATCT TTCTCATCA |
Hd1-QR | TCTGGAATTTGGCTATACTATCACC |
Ehd1-QF | GGATGCAAGGAAATCATGGA |
Ehd1-QR | AATCCCATCGGAAATCTTGG |
OsGI-QF | TGGAGAAAGGTTGTGGATGC |
OsGI-QR | GATAGACGGCACTTCAGCAGAT |
Ghd7-QF | AAATCCGGTACGCGTCCAG |
Ghd7-QR | GACATAGGTGGATGGCGGTG |
Table 1 Primers used in the study.
引物名称 Primer name | 引物序列(5′-3′) Primer sequence(5′-3′) |
---|---|
OsENO2-2-F | GGGGGTACCATGGTTCAACAGCTTGATGGAA |
OsENO2-2-R | GGGGGTACCTTAGTACGGCTCCACAGGGG |
R-OsENO2-2-F | AGAACTAGTGGTACCTGCTTCCACCGGAATATATGAG |
R-OsENO2-2-R | AGAGAGCTCGGATCCCCCTCTTTGTTTTCCTGAATGTTA |
Hn-F | TACACAGCCATCGGTCCAGA |
Hn-R | TAGGAGGGCGTGGATATGTC |
OsENO2-2-QF | GGCCAAGATGCCACAAATGT |
OsENO2-2-QR | TTGCCTGTGTAGCCAGCCTTA |
Actin-QF | TGTATGCCAGTGGTCGTACCA |
Actin-QR | CCAGCAAGGTCGAGACGAA |
RFT1-QF | TGGTGTTCGTGCTGTTCCA |
RFT1-QR | TTGTAGAGCTCGGCGAAGTTC |
RID1-QF | CGACGACAATAGCTCGATCGC |
RID1-QR | GTGCATGGTCACGGAGCCTT |
Hd3a-QF | GCTCACTATCATCATCCAGCATG |
Hd3a-QR | CCTTGCTCAGCTATTTAATTGCATAA |
Hd1-QF | TCAG CAACAGCATATCT TTCTCATCA |
Hd1-QR | TCTGGAATTTGGCTATACTATCACC |
Ehd1-QF | GGATGCAAGGAAATCATGGA |
Ehd1-QR | AATCCCATCGGAAATCTTGG |
OsGI-QF | TGGAGAAAGGTTGTGGATGC |
OsGI-QR | GATAGACGGCACTTCAGCAGAT |
Ghd7-QF | AAATCCGGTACGCGTCCAG |
Ghd7-QR | GACATAGGTGGATGGCGGTG |
Fig. 2. PCR amplification of OsENO2-2 and identification of recombinant plasmid. A, PCR amplification of OsENO2-2 full open reading frame. B, Digesting identification of recombinant plasmid. Lanes 1-4, Four independent clones of recombinant plasmid. Lane 1, 3, 4, Positive clones; Lane 2, Negative control.
Fig. 3. Tissue-specific expression of OsENO2-2. S, Seedling; FL, Flag leaf; L, The second leaf from the top at the heading stage; LS, Leaf sheath; ST, Stem; R, Root; PA, Panicle; EN, Endosperm.
Fig. 4. PCR analysis of Hn gene in T0 transgenic positive plants and expression analysis of OsENO2-2 in transgenic lines. A, PCR identification of Hn gene of the transgenic plants. The transgenic plants of the overexpressed lines, the transgenic plants of the control group lines, the transgenic plants of the RNAi lines, from up to down, respectively. B, The relative expression level of OsENO2-2 in transgenic plants(Mean±SD). WT, Wild type; OX-1 to OX-8, Eight independent overexpression transgenic plants. R-1 to R-7, Seven independent RNAi transgenic plants.
Fig. 5. Heading date of the transgenic plants(Mean±SD). A, Heading phenotypes of wild-type and overexpression plants. WT, Wild type; OX, Overexpression lines. B, Heading date of wild-type and overexpressing plants(n=30). WT, Wild type; OX-1, OX-2, OX-3, Overexpressed transgenetic positive plants; OX-7, The transgenic plants of the control group. C, Statistical analysis of heading date of wild-type, overexpressed plants and RNAi plants under long-day condition(14h light/10 h dark) (n=10). D, Statistical analysis of heading date of wild-type, overexpressed plants and RNAi plants under short-day condition(10h light/14h dark) (n=10). **The differences between wild type and transgenic plants are significant at 0.01 level(Student's t-test).
Fig. 6. Circadian rhythm expression of OsENO2-2 under different photoperiods(Mean±SD). LD, Long-day condition(light 14 h/dark 10 h); SD, Short-day condition(light 10 h/dark 14 h); WT, Wild type; OX, OsENO2-2 overexpressed plants.
Fig. 7. Expression levels of key heading regulatory genes in leaves of OsENO2-2 overexpression plants(OX) compared to wild type(WT) under long-day condition(Mean±SD).
Fig. 8. Expression levels of key heading regulatory genes in leaves of OsENO2-2 overexpression plants(OX) compared to wild type(WT) under short-day condition(Mean±SD).
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