概要: Stomata play a fundamental role modulating the exchange of gases between plants and the atmosphere. These microscopic structures form in high numbers on the leaf epidermis and are also present on flowers. Although leaf stomata are well-studied, little attention has been paid to the development or function of floral stomata. To characterise gene expression during the stomatal development in IR64 florets, as well as to identify potential gene expression differences in florets of transgenic plants with a reduction in stomatal density, a RNA-sequencing was carried out using floret samples from different developmental stages of IR64 (control) and transgenic plants overexpressing the gene OsEPF1. Differential expression analyses conducted on the RNA-seq dataset indicate that the cellular transitions during the development of floral stomata are regulated by the same genetic network used in rice leaves, and reveal alterations in global gene expression in florets of plants overexpressing OsEPF1.
项目整体设计: This dataset includes samples of florets in the developing stages 1 to 5 and mature from IR64 (control) and OsEPF1-oe-S (transgenic) rice plants. Three biological replicates per sample type were sequenced.
Plants were grown in Conviron growth cabinets under 12 h photoperiod (850 - 1000 µmol m-2 s-1 PAR at canopy level), 30°C day and 24°C night temperature, and 60 % relative humidity, with a constant supply of water to the pot base. From the 4th growth week, plants were fertilized in intervals of two weeks with Chempak High Nitrogen Feed No. 2 (Thompson & Morgan, Ipswich, UK).
处理方案:
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提取方案:
IR64 and OsEPF1-oeS developing florets were collected ~75-85 days after sowing, and mature florets were collected after panicle heading, but prior to anthesis (~ 95 days after sowing). RNA was extracted from floret samples using the Spectrum Plant Total RNA Kit (Sigma-Aldrich), with the On-Column DNase I Digest step, according to the manufacturer’s instructions. RNA samples extracted from developing florets of three individual plants were then mixed, in proportional RNA quantities. Therefore, each biological replicate for developing florets comprised RNA from three different rice plants. Three biological replicates per stage were sequenced.
建库方案:
Enrichment of mRNA was carried out using the NEBNext Poly(A) mRNA Magnetic Isolation Module (Biolabs), and library preparation was performed using the NEBNext Ultra RNA Library Prep Kit (Biolabs).
