Integration of molecular biology tools for identifying promoters and genes abundantly expressed in flowers of Oncidium Gower Ramsey

Abstract

BackgroundOrchids comprise one of the largest families of flowering plants and generate commercially important flowers. However, model plants, such as Arabidopsis thaliana do not contain all plant genes, and agronomic and horticulturally important genera and species must be individually studied.ResultsSeveral molecular biology tools were used to isolate flower-specific gene promoters from Oncidium 'Gower Ramsey' (Onc. GR). A cDNA library of reproductive tissues was used to construct a microarray in order to compare gene expression in flowers and leaves. Five genes were highly expressed in flower tissues, and the subcellular locations of the corresponding proteins were identified using lip transient transformation with fluorescent protein-fusion constructs. BAC clones of the 5 genes, together with 7 previously published flower- and reproductive growth-specific genes in Onc. GR, were identified for cloning of their promoter regions. Interestingly, 3 of the 5 novel flower-abundant genes were putative trypsin inhibitor (TI) genes (OnTI1, OnTI2 and OnTI3), which were tandemly duplicated in the same BAC clone. Their promoters were identified using transient GUS reporter gene transformation and stable A. thaliana transformation analyses.ConclusionsBy combining cDNA microarray, BAC library, and bombardment assay techniques, we successfully identified flower-directed orchid genes and promoters.

BeMADS1 is a key to delivery MADSs into nucleus in reproductive tissues-De novo characterization of Bambusa edulis transcriptome and study of MADS genes in bamboo floral development

“…To identify the promoter region and to clone full length genes, a BAC strategy was used [8,33-35] to identify 7 additional full length BeMADS genes in B. oldhamii (Table 2). …”

The location and translocation of ndh genes of chloroplast origin in the Orchidaceae family

“…The BAC clones containing the ndh regions and cp-like DNA of interest were obtained by PCR screening from the super pool, plate, row and spot31. These primers were also used to screen the O.…”

Identification of Heat Shock Transcription Factor Genes Involved in Thermotolerance of Octoploid Cultivated Strawberry

“…Each construct was subsequently recombined as an N-terminal fusion of GFP into the Gateway destination binary vector pK7WGF2 (Functional Genomics Division of the Department of Plant Systems Biology, Gent, Belgium), yielding 35S::GFP–FaTHSFA2a and 35S::GFP–FaTHSFB1a by an attL× attR recombination reaction (Invitrogen, Carlsbad, CA, USA). These GFP fusion constructs were isolated and transformed into floral lips of orchids by bombardment transformation [71]. A Zeiss LSM 510 META laser-scanning confocal microscope using an LD C-Apochromat 409/1.1 W objective lens was subsequently used to observe the fluorescence emitted in the transformed cells, as previously described [71].…”

“…These GFP fusion constructs were isolated and transformed into floral lips of orchids by bombardment transformation [71]. A Zeiss LSM 510 META laser-scanning confocal microscope using an LD C-Apochromat 409/1.1 W objective lens was subsequently used to observe the fluorescence emitted in the transformed cells, as previously described [71]. …”