Illumina NovaSeq 6000 paired end sequencing; RNA-seq of knock-out mutants of Candida parapsilosis CLIB 214 (CBS 604)
RNA-seq of knock-out mutants of Candida parapsilosis CLIB 214 (CBS 604)
The strain CPL2H1 and two homozygous mutants otf1/otf1 and gtf1/gtf1 derived from this strain have the background of Candida parapsilosis CLIB214 (CBS604). The strains CPL2H1 (LH), CPL2H1∆gtf1/∆gtf1 and CPL2H1∆otf1/∆otf1 were pre-cultivated in an SD medium (0.67% [wt/vol] yeast nitrogen base w/o amino acids (Difco), 2% [wt/vol] glucose) for 24 h at 28 °C, then washed in water, inoculated (OD600 ~ 0.2) into an SMix medium (0.67% [wt/vol] yeast nitrogen base w/o amino acids (Difco), 5 mM 3-hydroxybenzoate, 5 mM 4-hydroxybenzoate and 5 mM hydroquinone) and grown for additional 24 h at 28 °C. The pre-cultures were inoculated (OD600 ~ 0.3-0.4) into an SMix medium and cultivated at 28 °C till OD600 ~ 1. All media were supplemented with leucine (20 μg/ml) and histidine (20 μg/ml). Total RNA was extracted using the hot acid phenol protocol essentially as described in Collart and Oliviero (2001) and purified using an Uneasy mini kit (Qiagen); (RIN 8.6 - 9.1). Reference: Collart, M.A. and Oliviero, S. (2001) Preparation of yeast RNA. Curr. Protoc. Mol. Biol. Chapter 13: Unit 13.12.  The sequencing library was prepared using a TruSeq Stranded mRNA LT Sample Prep Kit (2x 151).