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Acta Agronomica Sinica ›› 2022, Vol. 48 ›› Issue (8): 1938-1947.doi: 10.3724/SP.J.1006.2022.14155

• CROP GENETICS & BREEDING · GERMPLASM RESOURCES · MOLECULAR GENETICS • Previous Articles     Next Articles

Bna.C02SWEET15 positively regulates the flowering time of rapeseed through photoperiodic pathway

LI Sheng-Ting1,**(), XU Yuan-Fang1,**(), CHANG Wei1, LIU Ya-Jun2, GU Yuan2, ZHU Hong1, LI Jia-Na1,3,4, LU Kun1,3,4,*()   

  1. 1College of Agronomy and Biotechnology, Southwest University, Chongqing 400715, China
    2Agricultural Technology Extension Station in Lincang City, Lincang 677099, Yunnan, China
    3Engineering Research Center of South Upland Agriculture, Ministry of Education, Chongqing 400715, China
    4Academy of Agricultural Sciences, Southwest University, Chongqing 400715, China
  • Received:2021-08-26 Accepted:2021-11-29 Online:2022-08-12 Published:2021-12-16
  • Contact: LU Kun E-mail:lishengting0123@163.com;1095245283@qq.com;1095245283@qq.com;drlukun@swu.edu.cn
  • About author:First author contact:

    ** Contributed equally to this work

  • Supported by:
    National Natural Science Foundation of China(31871653);National Key Research and Development Program of China(2018YFD0100500);Project of Intellectual Base for Discipline Innovation in Colleges and Universities(B12006);Key Project of Chongqing Natural Science Foundation(cstc2021ycjh-bgzxm0033);Chongqing Postgraduate Research Innovation Project(CYB21115);Germplasm Creation Special Program of Southwest University

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Abstract:

Flowering time, an important agronomic index of crops, is related to the growth cycle and the yield of crops. Sugar transporters are important carbohydrate transporters in plants, whereas there are few reports on the effect of sugar transporters on flowering time in rapeseed (Brassica napus). In this study, sucrose transporter Bna.C02SWEET15 related to flowering in rapeseed were screened and identified from Fox-Hunting library, whose biological function and regulation mechanism were further characterized through expression pattern and subcellular localization analysis, phenotype analysis in transgenic plants and mutant, detection of marker genes expression related to flowering. Bna.C02SWEET15 could be identified in all tissues of Brassica napus, and the relative expression level was the most significant in seeds at 30 days after flowering. Bna.C02SWEET15 was located in cell membrane, and the promoter activity was mainly in anther and seed at 30 days after flowering. Overexpression of Bna.C02SWEET15 in Arabidopsis resulted in flowering in advance and increased the expression of photoperiod key genes CO, FT, and LFY, while decreased the expression of negative regulation gene FLC. In addition, the flowering time of atsweet15a and transgenic RNAi-Bna.C02SWEET15 were later. We proposed that Bna.C02SWEET15 could positively regulate the flowering time through photoperiod pathway, thus affecting the growth cycle and final yield in rapeseed. These results provide important basis for understanding the regulatory role of sugar transporters in crops and lay a theoretical foundation for understanding the regulatory role of sugar transporters in crops.

Key words: Brassica napus, Bna.C02SWEET15, photoperiod, flowering

Table S1

Primers used in this study"

引物名称
Prime name		 引物序列
Primer sequences (5'-3')
26S-F rRNA CACAATGATAGGAAGAGCCGAC
26S-R rRNA CAAGGGAACGGG CTTGGCAGAATC
BnaActin7-F TGGGTTTGCTGGTGACGAT
BnaActin7-R TGCCTAGGACGACCAACAATACT
qRT F-BnaSWEET15 TCCTTGTATTCCTCGCTCCC
qRT R-BnaSWEET15 CAGCCGAAAGAGTTGATGGT
OV-SWEET15 F CACCATGAGTTTGGCACACCAAAT
OV-SWEET15 R ATGCCTGAGAGCTTCTTGCAC
Bna.C02SWEET15-PRO-F acctgcaggcatgcaACAGACCTCGTTTCTATAGCTATCT
Bna.C02SWEET15-PRO-R taccctcagatctaccatggAAACTAATGAGTCTCAGGTGGTGT
F Hyg CTATTTCTTTGCCCTCGGACGA
R Hyg ATGAAAAAGCCTGAACTCACCGCG
F35S3ND GGAAGTTCATTTCATTTGGAGAG
OCS5ND CGATCATAGGCGTCTCGCATATCTC
F Bar CGACATCCGCCGTGCCACCGA
R Bar GTACCGGCAGGCTGAAGTCCAGC
Actin real F GGTAACATTGTGCTCAGTGGTGG
Actin real R AACGACCTTAATCTTCATGCTGC
qRT F-AtSWEET15 CATAGCCATGTTCTTCGCTTAC
qRT R-AtSWEET15 CTTTGTCTTTATCACACGAGCC
M13pF ACACAGGAAACAGCTATGACCAT
M13pR GGGTCCTAACCAAGAAAATGAA
RNAi-SWEET15-F GGATCCGACGTCTCTCCCATACCAACATTCATAACA
RNAi-SWEET15-R TCTAGACCATGGATAGTATAAGCTGTACTACT
BnaPAP2-F GGATCCACCTAAGCATGCATTTGAAAA
BnaPAP2-R ATTTAAATGACGTCAGGTTTACATTCAAGACACA
引物名称
Prime name		 引物序列
Primer sequences (5'-3')
F35S3N GGAAGTTCATTTCATTTGGAGAG
ROCST5N GCTCAGGTTTTTTACAACGTGCAC
CO real F GCCATCAGCGAGTTCCAATTCTAC
CO real R CCTTCCTCTTGATCCACCACCAG
FLC real F CGTCGCTCTTCTCGTCGTCTC
FLC real R TTCGGTCTTCTTGGCTCTAGTCAC
FT real R GCCTGCCAAGCTGTCGAAACAATA
FT real F TCCCTGCTACAACTGGAACAACCT
LFY real F TCCACTGCCTAGACGAAGAAGC
LFY real R TCCCAGCCATGACGACAAGC
N674321-LP CAGCCGAGTACAGAGACTTGG
N674321-RP TTTCCTCGCTTTTATCTTCGG
LBb1.3 ATTTTGCCGATTTCGGAAC

Fig. 1

Relative expression pattern of Bna.C02SWEET15 in Brassica napus A: the expression pattern of Bna.C02SWEET15 in Brassica napus using qRT-PCR. R: root; St: stem; L: leaf; F: flowers; MB: main bud; 5D-S, 10D-S: silique at 5 days and 10 days after flowering; 15D-SE, 21D-SE, 30D-SE: seeds at 15, 21, and 30 days after flowering; 15D-SW, 21D-SW, 30D-SW: seed pericarp at 15, 21, and 30 days after flowering; B: histochemical staining of proBna.C02WEET15::GUS. a: seed pericarp after germination; b: young seedling; c: rosette; d: stem leaf; e: anther; f: silique; g: root; h: seed at 30 days after flowering."

Fig. 2

Subcellular localization of Bna.C02SWEET15-YFP protein Bar: 20 μm."

Fig. 3

Phenotypic observation and marker gene expression detection of Bna.C02SWEET15 overexpressed transgenic plants A: phenotypic observation in overexpressed transgenic plants; bar: 1 cm. B: the expression level of Bna.C02SWEET15 in transgenic seedlings using qRT-PCR; C: flowering time of Bna.C02SWEET15 transgenic plants; D: marker genes expression in flowering pathway. * and ** represent significant difference at the 0.05 and 0.01 probability levels, respectively."

Fig. 4

Phenotypic observation of Arabidopsis thaliana mutant atsweet15 A: sequence similarity between AtSWEET15 and Bna.C02SWEET15; B: map of mutant insertion sites; C: phenotypic observation of atsweet15a; D: the expression level of AtSWEET15 in atsweet15a. Bar: 1 cm. ** represents significant difference at the 0.01 probability level. "

Fig. 5

Phenotypic observation of transgenic RNAi-Bna.C02SWEET15 A: phenotypic observation of RNAi-Bna.C02SWEET15; B: gene expression level in transgenic seedlings; C: flowering time of RNAi-Bna.C02SWEET15 transgenic plants; bar: 10 cm. ** represents significant difference at the 0.01 probability level."

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