Transcriptomic changes reveal gene networks responding to the overexpression of a blueberry DWARF AND DELAYED FLOWERING 1 gene in transgenic blueberry plants

Abstract

BackgroundConstitutive expression of the CBF/DREB1 for increasing freezing tolerance in woody plants is often associated with other phenotypic changes including dwarf plant and delayed flowering. These phenotypic changes have been observed when Arabidopsis DWARF AND DELAYED FLOWERING 1 (DDF1) was overexpressed in A. thaliana plants. To date, the DDF1 orthologues have not been studied in woody plants. The aim of this study is to investigate transcriptomic responses to the overexpression of blueberry (Vaccinium corymbosum) DDF1 (herein, VcDDF1-OX).ResultsThe VcDDF1-OX resulted in enhanced freezing tolerance in tetraploid blueberry plants and did not result in significant changes in plant size, chilling requirement, and flowering time. Comparative transcriptome analysis of transgenic ‘Legacy-VcDDF1-OX’ plants containing an overexpressed VcDDF1 with non-transgenic highbush blueberry ‘Legacy’ plants revealed the VcDDF1-OX derived differentially expressed (DE) genes and transcripts in the pathways of cold-response, plant flowering, DELLA proteins, and plant phytohormones. The increase in freezing tolerance was associated to the expression of cold-regulated genes (CORs) and the ethylene pathway genes. The unchanged plant size, dormancy and flowering were due to the minimal effect of the VcDDF1-OX on the expression of DELLA proteins, flowering pathway genes, and the other phytohormone genes related to plant growth and development. The DE genes in auxin and cytokinin pathways suggest that the VcDDF1-OX has also altered plant tolerance to drought and high salinity.ConclusionA DDF1 orthologue in blueberry functioned differently from the DDF1 reported in Arabidopsis. The overexpression of VcDDF1 or its orthologues is a new approach to increase freezing tolerance of deciduous woody plant species with no obvious effect on plant size and plant flowering time.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-017-1053-z) contains supplementary material, which is available to authorized users.

An invaluable transgenic blueberry for studying chilling-induced flowering in woody plants

“…These plants were developed from in vitro shoot cultures and were grown in the greenhouse for approximately 14 months after they were rooted. The transgenic plants, three to five plants per transgenic event, were from 49 transgenic events, each containing an overexpression vector for VcDDF1 [ 27 , 28 ]. All three plants of one VcDDF transgenic event, which was designated Mu-Legacy, started flowering in the heated greenhouse in mid-October (Fig.…”

“…6 ). The DE gene networks of Mu-Legacy have much less GO terms than those of VcDDF1 -Legacy [ 27 ], indicating the potential effects of both insertion positions and VcDDF1 expression levels. When flowering pathway genes of A. thaliana were used to identify DE genes (DEGs), only four DEGs, including orthologues of PROTEIN FD (FD), TERMINAL FLOWER 1 (TFL1) , ACTIN-RELATED PROTEIN6 ( ARP6 ), DOF ZINC FINGER PROTEIN5.3 ( DOF5.3 ), were detected and they were all repressed (Fig.…”

“…Unlike Mu-Legacy plants, VcDDF1 -Legacy plants, flowered similar to nontransgenic ‘Legacy’ [ 27 ], suggesting that the overexpressed VcDDF1 might not be the major factor for the altered flowering in the Mu-Legacy. To further explore the potential candidate genes that could be resposible for the mutation in Mu-Legacy, transcriptomic analyses of Mu-Legacy (compared to VcDDF1 -Legacy) were also conducted in leaves and nonchilled flower buds.…”

“…9 ). Since the VcRR2 did not show differential expression in leaves and flower buds of VcDDF1 -Legacy [ 27 ], the upregulated expression of VcRR2 could be responsible for the mutation of Mu-Legacy. It is likely that the CaMV 35S promoter of the gus A gene drove through the NOS terminator and then promoted expression of the adjacent VcRR2 gene.…”

“…Comparative transcriptome analysis provides a powerful tool to study the differentially expressed genes associated with phenotypic changes in transgenic blueberry plants [ 20 , 21 , 26 , 27 ]. In terms of flowering time regulation, overexpression of a blueberry FLOWERING LOCUS T ( VcFT ) (an increase of approximately 2900-fold) was unable to completely eliminate the need for chilling in blueberry for normal flowering [ 20 ].…”