Temporal RNA-Seq Transcriptional Profiling in the Facultative Crassulacean Acid Metabolism (CAM) Model Mesembryanthemum crystallinum

The common ice plant (Mesembryanthemum crystallinum) is an important facultative Crassulacean acid metabolism (CAM) model that can switch from C₃ photosynthesis to CAM following high salinity or water-deficit stress treatments. In order to explore the circadian clock-regulated mechanisms that control the expression of CAM, transcriptome sequencing was performed on well-watered and water-deficit stress treated leaf tissue collected in parallel every 4 h over a 72 h time course under both 24 h light/dark (diel entrainment) and 48 h light/light (zeitgeber or free running) conditions in order to capture the full repertoire of circadian clock-controlled transcriptional outputs. cDNA libraries were constructed for 114 ice plant samples and Illumina HiSeq2000 was performed using a total of 19 flow-cell channels, with 6 samples in each. Approximately 2,241,015,571 bp of raw data were generated yielding a total of 2,415,294,564 trimmed sequence reads. All trimmed, cleansed reads were assembled de novo using the Trinity program. The complete read dataset assembled into 109,902 contigs, which ranged in size from 201 bp to 15,797 bp with a mean contig length of 1228 bp. RSEM and Bowtie were then used to assign reads to multiple genes and count relative transcript abundances. Read counts were mapped, normalized and differentially expressed genes were detected using the DESeq package. Circadian clock output differences between the C₃ photosynthesis and CAM states will be discussed in detail. These results will facilitate the identification of the molecular genetic machinery required for CAM using comparative genomics as well as aid in future ice plant genome annotation efforts.