GSE119686
Characterization of the transcriptome of Haloferax volcanii with mixed RNA-Seq
Recently a dRNA-Seq study with Haloferax volcanii has been published that led to the discovery of nearly 2800 novel transcription start sites for non-coding RNAs. However, the dRNA-Seq results are confined to the 5''-end of transcripts and does not contain any length information. Therefore, a major aim of the present RNA-Seq study was the elucidation of the lengths of the novel non-coding RNAs. A second aim was to analyze the operon structure of protein-coding genes. The recent dRNA-Seq study was confined to cultures grown under optimal conditions. It was revealed that less than half of the protein-coding genes were expressed. Therefore, the present RNA-Seq study used cultures grown under four different conditions to identify further transcripts of protein-coding genes that are not needed under optimal conditions. RNA samples from the four conditions were mixed prior to the RNA-Seq analyses (mixed RNA-Seq). The results allowed to characterize the sum of transcriptomes of H. volcanii under four conditions. Northern blot analyses were used to characterize selected examples in detail. Many important results were obtained, including the length determination of all transcripts, a genome-wide operon analysis, verification of a high fraction of antisense RNAs, and the discovery that 30% of all protein-coding transcripts have overlapping 3''-UTRs. Overall design: Three H. volcanii cultures were grown to mid-exponential growth phase under optimal conditions, at low salt concentration, and with glucose as sole carbon source, In addition, a fourth culture was grown to stationary phase. Total RNA was isolated from all four cultures and equal fractions were mixed. RNA-Seq was performed with the mixed RNA sample. 50 million paired end reads of 150 nt length were generated.