title: GSM6077774: Candida albicans, CRISPRa non-targeting control, rep1; Candida albicans; RNA-Seq
library_construction_protocol: RNA extraction and sample QC was conducted at GENEWIZ, LLC./Azenta US, Inc (South Plainfield, NJ, USA) as follows:  Total RNA was extracted using Qiagen RNeasy Plus Universal kit following manufacturer's instructions (Qiagen, Hilden, Germany). RNA samples were quantified using Qubit 2.0 Fluorometer (ThermoFisher Scientific, Waltham, MA, USA) and RNA integrity was checked with 4200 TapeStation (Agilent Technologies, Palo Alto, CA, USA). Library preparations and sequencing reactions were conducted at GENEWIZ, LLC./Azenta US, Inc (South Plainfield, NJ, USA) as follows: ERCC RNA Spike-In Mix kit (cat. 4456740) from ThermoFisher Scientific, was added to normalized total RNA prior to library preparation following manufacturer's protocol. The RNA sequencing library was prepared using the NEBNext Ultra II RNA Library Prep Kit for Illumina using manufacturer's instructions (New England Biolabs, Ipswich, MA, USA). Briefly, mRNAs were initially enriched with Oligod(T) beads. Enriched mRNAs were fragmented for 15 minutes at 94 °C. First strand and second strand cDNA were subsequently synthesized. cDNA fragments were end repaired and adenylated at 3'ends, and universal adapters were ligated to cDNA fragments, followed by index addition and library enrichment by PCR with limited cycles. The sequencing library was validated on the Agilent TapeStation (Agilent Technologies, Palo Alto, CA, USA), and quantified by using Qubit 2.0 Fluorometer (ThermoFisher Scientific, Waltham, MA, USA) as well as by quantitative PCR (KAPA Biosystems, Wilmington, MA, USA). The sequencing libraries were multiplexed and clustered onto a flowcell. After clustering, the flowcell was loaded onto the Illumina HiSeq 4000 or equivalent instrument according to manufacturer's instructions. The samples were sequenced using a 2x150bp Paired End (PE) configuration. Image analysis and base calling were conducted by the Illumina Control Software. Raw sequence data (.bcl files) generated from the Illumina instrument was converted into fastq files and de-multiplexed using Illumina bcl2fastq 2.20 software. One mis-match was allowed for index sequence identification.