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Natural genetic variation in Brassica homologs of FLOWERING LOCUS T and characterization of its expression domains

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Abstract

FLOWERING LOCUS T (FT), a major effect gene, regulates flowering time in Arabidopsis. We analyzed evolutionary changes distinguishing two FT homeologous loci in B. rapa, described genetic variation in homologs isolated and reported expression pattern of FT in B. juncea. Synteny analysis confirmed presence of two FT genomic copies in B. rapa ssp. pekinensis and resolved pre-existing anomalies regarding copy number in “AA” genome. Synteny analysis of B. rapa homeologous regions CR1 (129 kb) and CR2 (232 kb) revealed differential gene fractionation and wide-spread re-arrangements. Seven genomic DNA (gDNA) variants (2.1–2.2 kb) and 10 complementary DNA (cDNA) variants (528 bp) were isolated from 6 Brassica species. The gDNA variants shared 72–99 % similarity within Brassica and 58–60 % between Arabidopsis and Brassica. FT cDNA variants shared 92–100 % similarity within Brassica and 87 % between Arabidopsis and Brassica. Phylogenetic analysis of FT gDNA, cDNA and protein sequences revealed two major clades, differentiating homologs derived from species containing shared “BB” and “CC” genomes. Phylogram based on Brassica FT gDNA differentiated homeologs derived from AA-LF (Least fractioned) and AA-MF1 (Moderately fractioned) sub-genomes. Analysis of FT expression pattern in B. juncea revealed increasing levels correlating with attainment of physiological maturity; highest levels were detected in older leaves implying conservation in spatio-temporal expression pattern vis-à-vis Arabidopsis. In conclusion, our study reveals that polyploidy in Brassicas resulted in expansion of FT gene copies with homologs charting independent evolutionary course through accumulation of mutations. However, expression domains of FT remained conserved across Brassicaceae to preserve the critical function of FT in controlling flowering time.

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Abbreviations

AtFT:

Arabidopsis thaliana FT

BAC:

Bacterial Artificial Chromosome

BRAD:

Brassica database

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Acknowledgments

The authors thank Dr. Arvind Kumar, DDG (Education, ICAR), Dr J.S. Chauhan, ADG (Seed, ICAR) and Dr. Abha Agnihotri (Amity University, Noida) for providing the germplasm. Dr. S.B. Tripathi and Mr M.S.Negi for offering valuable suggestions. S.M. Shivaraj, Priyanka Dhakate and Shikha Tyagi for technical assistance and motivation. Contribution of Mr. Hari Ram Gupta in field-related activities is appreciated. Financial support for this study from Department of Biotechnology, Govt. of India (BT/PR8659/PBD/16/739/2006, BT/PB/01/01/2011) is gratefully acknowledged. Infrastructural support from TERI and TERI University are also acknowledged.

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  1. Pratiksha Mayee

    Present address: Department of Research, Ankur Seeds Pvt Ltd, Nagpur, 440018, Maharashtra, India

Authors and Affiliations

  1. Department of Biotechnology, TERI University, 10 Institutional Area, Vasant Kunj, Delhi, 110070, India

    Pratiksha Mayee & Anandita Singh

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Correspondence to Anandita Singh.

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Funding

Author Anandita Singh has received research grants from Department of Biotechnology, Govt. of India (BT/PR8659/PBD/16/739/2006, BT/PB/01/01/2011).

Conflict of interest

We Pratiksha Mayee and Anandita Singh hereby declare that we have no conflict of interest.

Electronic supplementary material

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Supplementary Fig. A1

Sliding window analysis of FT gDNA sequences from Brassica species and A. thaliana. The graphical output using DNAsp 5.0 (window size = 25 bp, step = 2) shows that the sequences corresponding to exons are more conserved compared to the introns. The regions where the Pi value peaks and dips correspond to predicted introns and exons, respectively (PDF 361 kb)

Supplementary Fig. A2

Phylogram of Brassica FT gDNA sequences. The sequences are clustered in two distinct clades. The homologs isolated from B. napus, B. carinata and B. oleracea (containing shared “CC” genome) form one clade (Group I), while sequences isolated from B. nigra and B. juncea containing shared “BB” genome form a distinct clade (Group II) (PDF 122 kb)

Supplementary Fig. A3

Phylogenetic analysis of in-silico translated FT. The protein variants from Brassica display similarity with FT and TSF compared to the other family members. FT sequences from B. oleracea and B. napus belong to one clade (Group I) while B. juncea, B. rapa, B. nigra and B. carinata form a different clade (Group II) (PDF 350 kb)

Supplementary Fig. A4

Phylogenetic analysis of FT proteins from angiosperms. The sequences from genus Brassica are monophylogenetic and form a distinct clade closer to AtFT (PDF 262 kb)

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Mayee, P., Singh, A. Natural genetic variation in Brassica homologs of FLOWERING LOCUS T and characterization of its expression domains. J. Plant Biochem. Biotechnol. 25, 270–277 (2016). https://doi.org/10.1007/s13562-015-0338-2

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