title: Illumina HiSeq 2500 sequencing; CAGE experiment of S. Cerevisiae strain CEN.PK113-7D in four different chemostat conditions
design: CAGE experiment of S. Cerevisiae strain CEN.PK113-7D in four different chemostat conditions
library_name: Glu_Rep2_s
library_construction_protocol: 10OD of cells were collected, pelleted and snap frozen in liquid nitrogen. The S. cerevisiae strain CEN.PK113-7D was pre-cultured in a batch culture in Delft media with 2% glucose at 30°C and 200 rpm in shake flasks and then transferred to the chemostats in duplicates. Four different media compositions with a limitation in a different nutrient were employed in the chemostat runs. The temperature was also set to 30°C. The dilution rate was fixed to 0.1 and the cells were grown for 4 days to achieve stable cell numbers in the culture.  RNA was extracted using the RNeasy® Mini Kit from QIAGEN. Cells were mechanically disrupted using a FastPrep®-24 from MPbio in combination with the lysing matrix tubes type C from MPbio. The FastPrep was run 3 times for 20 seconds with 4.0m/s settings and a 5 minute break in between each run. RNA quality was assessed using NanoDrop and Bioanalyzer. The non-amplification non-tagging CAGE protocol for Illumina sequencing (nAnT-iCAGE) as previously published by Murata et. al. 2014 was used on two biological replicates of each condition, starting with 5 micrograms of extracted RNA.
attributes.tag: Experimental Factor: environmental stress
attributes.value: low glucose