To control for cell synchronization, cells for the+N and — N conditions were harvested at the same time of day. Total RNA was extracted and purified separately from each of the two nitrogen replete and the two nitrogen limited cultures using the RNeasy Lipid Tissue Mini Kit (Qiagen, Valencia, CA,

USA). The quality of purified RNA was determined on an Agilent 2100 bio­analyzer (Agilent Technologies, Santa Clara, CA, USA). Isolation of mRNA from total RNA was carried out using two rounds of hybridization to Dynal oligo(dT) magnetic beads (Invitrogen, Carlsbad, CA, USA). Aliquots from mRNA samples were used for construction of the cDNA libraries using the mRNA-Seq Kit supplied by Illumina (Illumina, Inc., San Diego, CA, USA). Briefly, the mRNA was fragmented in the presence of divalent cations at 94°C, and subsequently converted into double stranded cDNA following the first — and second-strand cDNA synthesis using random hexamer primers. After polishing the ends of the cDNA using T4 DNA polymerase and Kle — now DNA polymerase for 30 min at 20°C, a single adenine base was added to the 3’ ends of cDNA molecules. Illumina mRNA-Seq Kit specific adap­tors were then ligated to cDNA 3’ ends. Next, the cDNA was PCR-amplified for 15 cycles, amplicons were purified (QIAquick PCR purification kit, Qia — gen Inc., Valencia CA, USA), and the size and concentration of the cDNA libraries were determined on an Agilent 2100 bioanalyzer. Each of the four cDNA libraries (two nitrogen deplete and two nitrogen replete) was layered on a separate Illumina flow cell and sequenced at the Yale University Center for Genome Analysis using Illumina HiSeq 100 bp single-end sequencing. An additional lane was dedicated to sequencing PhiX control libraries to provide internal calibration and to optimize base calling. The sequence data produced in this study can be accessed at NCBI’s Sequence Read Achieve with the accession number SRA048723.