Illumina HiSeq 2500 paired end sequencing; IRE1-dependent transcriptional change under a canonical endoplasmic-reticulum stressing stimulus and upon diauxic shift in Saccharomyces cerevisiae cells
IRE1-dependent transcriptional change under a canonical endoplasmic-reticulum stressing stimulus and upon diauxic shift in Saccharomyces cerevisiae cells
Cells were collected from liquid culture by centrifugation (1,500 xg; 1min) Yeast cells were shaken in standard synthetic dextrose (SD) medium at 30 C (starting OD600 0.3). Tunicamycin (2ug/ml final concentration) was added to exponentially growing cultures, which were further incubated for 1 hour. Total RNA was extracted for yeast cells using standard hot phenol method. Residual DNA was digested using DNase I (Takara). Library construction and sequencing for the Illumina HiSeq 2500 was provided as a custom service of Eurofins Genomics K. K. (Tokyo, Japan). The polyA fraction (mRNA) was isolated from total RNA, followed by its fragmentation. Then double-stranded (ds) cDNA was reverse transcribed from fragmented mRNA. The ds cDNA fragments were processed for adaptor ligation, size selection (for 200 bp inserts) and amplification to generate strand-specific cDNA libraries.
Experimental Factor: genotype
wild type genotype
Experimental Factor: environmental stress
none
Experimental Factor: compound
tunicamycin
Experimental Factor: dose
2