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题目:: The Transcriptionally Active Regions in the Genome of Bacillus subtilis
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状态:: 发布时间May 18, 2010 , 更新时间 March 27, 2012 , 提交时间 May 12, 2009,
物种:: Bacillus subtilis subsp. subtilis str. 168
摘要:: The majority of all genes have so far been identified and annotated systematically through in silico gene finding. Here we report the finding of 3,217 strand-specific Transcriptionally Active Regions (TARs) in the genome of Bacillus subtilis by the use of tiling arrays. We have measured the genome-wide expression during mid-exponential growth on rich (LB) and minimal (M9) medium. The identified TARs account for 81.5% of the genes as they are currently annotated and additionally we find 44 novel protein-coding genes, 85 putative non-coding RNAs (ncRNAs) and 184 antisense transcripts. Hybridization of labeled genomic DNA (gDNA) has revealed a few thousand regions giving rise to very low signals. These are mostly caused by sequence errors, as is observed in the highly degenerate trp operon and the folding of stable structures such as the regions containing Rho-independent terminators. Through this work we have discovered a group of membrane-associated genes, among these having a long conserved 3’ UnTranslated Region (UTR) predicted to fold into a large and highly stable structure. Among these are YwbN, a target of the twin-arginine translocase (Tat) protein translocation system. The transcriptionally active regions were determined at two different conditions, rich medium (Luria Broth) and minimal medium (M9). Cells were harvested at mid-exponential phase (OD600 = 0.5) and each condition were replicated three times. From each replicate RNA was extracted using FastRNA pro blue kit (Qbiogene) and labelled with Cy3 using a strand-specific protocol. The cDNA was hybridized to 385K NimbleGen tiling arrays 'BaSysBio Bsub T1' covering the entire genome with 45-65 nt isothermal probes, spaced by 22 nt on each strand. Additionally genomic DNA was extracted from 4 replicates at OD600 = 0.5, labelled and hybridized to the tiling arrays. These serve as a reference for the mRNA hybridizations. The normalized RNA hybridization signals were used to create segments using a structural change model (Huber et al, 2006) and transcriptionally active regions were identified by a segment joining-scheme.
实验种类:: transcription profiling by tiling array
样本量:: 10
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数据号:: E-GEOD-16086, GSE16086
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