Project description:Proper control of Auxin localization and accumulation are essential for leaf primordia initiation. Additionally, previous work has shown that the ERECTA family of genes also control leaf initiation. In this study we seek to understand the interaction between Auxin and the er erl2 double mutant in 3 days after germination (DAG) old seedlings. WT and mutant seedlings were either treated with IAA or mock treatments for 30 minutes. RNA was isolated and poly-A enriched strand specific libraries were constructed and paired end sequencing was carried out. The mutant showed mild, yet consistent enrichment of chloroplast and carbohydrate metabolism gene ontology terms in our mutant compared to WT. However, at 30 minutes post IAA treatment, the mutant and WT did not have significantly different responses to auxin. This indicates that the ERECTA family genes do not play a significant role in the immediate and direct response to auxin.
Project description:7-days-old Arabidopsis seedlings of wildtype (Col-0) were treated with 1 μM IAA for 15 minutes or 3 hours and gene expression of whole plant was analyzed using Affymetrix Gene 1.1 ST Array strips.
Project description:7-days-old Arabidopsis seedlings of wildtype (Col-0) were treated with 1 μM IAA for 15 minutes or 3 hours and gene expression of whole plant was analyzed using Affymetrix Gene 1.1 ST Array strips. Arabidopsis seedling (Col-0) 25 plants were grown in each plate (1/2 MS medium) under continuous light at 22 °C for 6 or 7 days. Ten seedlings were cultured in 50 ml plastic tube containing liquid medium (1/2 MS medium), and incubated with shaking. After 24 hours, samples were treated with 0.1% DMSO for 15 minutes or 3 hours (Mock treatments) and with 1 μM indole-3-acetic acid dissolved in DMSO for 15 minutes or 3 hours (IAA tretments). The plants were wiped on papertowels and sampled. Immediately after sampling of the plants, these plants were soaked in liquid nitrogen and stored at -80 °C. The experiment was repeated 3 times in 3 different days. Total RNA was extracted from the frozen samples using RNeasy Plant Mini Kit (Qiagen) following the protocol supplied by manufacturer. The quality of RNA was checked by 2100 Bioanalyzer system (Agilent) and 100 ng RNA of each experiment was applied to microarray analysis. The microarray analysis was carried out using Affymetrix gene 1.1 ST array strips and GeneAtlas (Affymetrix) following the protocol supplied manufacturer.
Project description:To gain more information about the Kyn inhibition of TAA1/TARs, we conducted a RNA-seq analysis to compare the genome-wide gene expression profiles of WT seedlings grown on MS medium or MS plus Kyn, and wei8-/- tar2+/- progeny grown on MS. We report that Kyn treatment largely mimics the loss of TAA1/TAR2 functions.
Project description:Arabidopsis seedlings (Col-0) were grown in suspension in half-strength MS medium with agitation at ~100 rpm at ~22 C under ~50 microeinsteins m-2s-1 cool white fluorescent continuous illumination as described by (Xiao et al., Plant Physiol. 2002 Dec;130(4):2118-28). Seedlings were treated at 10-12 days by addition of freshly made IAA (0.1 or 1.0uM) to each flask, and harvested after a 1 or 3 hour incubation. Controls were not treated and harvested at 0hr. All tissue harvested. Total RNA was extracted using TRIzol (Invitrogen) as described by the manufacturer and then filtered using QIAGEN RNeasy columns. cDNA was synthesized from total RNA using a Superscript double-stranded cDNA synthesis kit (Invitrogen) and a T7-dT24 primer. cRNA was synthesized using the Enzo BioArray HighYield RNA Transcript Labeling kit (Affymetrix p/n 900182) and fragmented by Mg2+ hydrolysis. 15ug per ATH1 array was hybridized overnight at 45 C. Arrays were then washed and scanned using the GeneChip FS400 fluidics station and Agilent GeneArray scanner. Images were analyzed using Affymetrix Microarray Suite 5.0, scaling to a target average intensity of 500. Spiking controls were added to the total RNA before cDNA synthesis and additional spiking samples were added to the resulting cDNA prior to cRNA synthesis.,In comparison table below:,SIGNAL_LOG_RATIO = Mean of log to base two of the experimental divided by control signal ratios across all probe pairs in a set.,CHANGE = Qualitative measurement indicating whether the probe set signal is increased (I), marginally increased (MI), not changed (NC), marginally decreased (MD), or decreased (D) as compared to a control hybridization across all probe pairs, based on a p-value calculation.,change_p-value = Measures the probability that all probe pairs in the set indicate a change, with 0 indicating strong likelyhood for increase, 0.5 indicating little probability for difference, and 1 indicating strong probability for decrease.
Project description:To gain more information about the Kyn inhibition of TAA1/TARs, we conducted a RNA-seq analysis to compare the genome-wide gene expression profiles of WT seedlings grown on MS medium or MS plus Kyn, and wei8-/- tar2+/- progeny grown on MS. We report that Kyn treatment largely mimics the loss of TAA1/TAR2 functions. RNA seq of 3 samples: Col-0 on MS medium, Col-0 on MS+Kyn medium, wei8 tar2(+/-) on MS medium.
Project description:Indica rice seedlings of IR64 variety were grown hydroponically for 7-days in a culture room with a daily photoperiodic cycle of 14h light and 10h dark. Seedlings were incubated in 0.1% dimethyl sulfoxide (control) or 50 micromolar solutions of indole-3-acetic acid (IAA treatment) and benzyl aminopurine (BAP treatment) for 1h and 3h. Equal amounts of 1h and 3h samoles were pooled for each treatment before RNA isolation. The 5 micrograms of each total RNA sample was processed for microarray analysis according to Affymetrix protocol. Experiment Overall Design: Two biological replicates for each sample (control, IAA treated, BAP treated) were used for microarray analysis. The experiment involves the identification of differentially expressed genes in IAA and BAP treated samples as compared to control.
Project description:We tracked the gene expression events following treatment of maize seedlings with the endoplasmic reticulum (ER) stress agent tunicamycin. ER stress elicits the unfolded protein response (UPR) and when plants are faced with persistent stress, the UPR transitions from an adaptive or cell survival phase to programmed cell death.