Project description:We have developed both WTSS-seq (whole transcriptome start site sequencing) and WTTS-seq (whole transcriptome termini site sequencing) methods to capture either 5’- or 3’-ends of transcripts. HATT-seq (head and tail tag sequencing) is still under development, which can be used to capture both 5’- and 3’ ends of each transcript simultaneously. Iso-seq was used to produce full-length transcripts, which can be used to validate both alternative transcription start sites and alternative polyadenylation sites. CAGE-seq was used to confirm alternative transcription start sites only.
Project description:The mammalian sex-determining gene Sry induces male development. Through analyzing transcriptome of pre-Sertoli cells, we identified a novel sequence transcribed within palindromic sequence surrounding Sry. To examine whether this sequence contains transcription start site, Cap analysis gene expression (CAGE) -seq analysis was performed, in which putative transcriptional start sites can be identified by sequencing the 3’ end of cDNA corresponding to 5’ end of RNA. CAGE seq revealed that Cap site was absent within the novel sequence, whereas Sry transcription start site found to locate in minus strand at chrY:2,663,800-2,663,900. Combining the result from long-read RNA seq, we concluded that this sequence is the second exon of mouse Sry.
Project description:We used SLIC-CAGE to map transcription start sites (TSSs) of mouse primordial germ cells from embryonic days 9.5-16.5, postnatal oocytes (P6, P14 and MII), and early 2-cell and 4-cell mouse embryos. We use this TSS data to show that the mouse germline development starts with the somatic promoter code with a prominent switch to the maternal code (W-box dependent) occurring during the follicular oogenesis. We also find that the promoters of gonadal germ cells are characterised by a previously unknown divergence from the somatic transcription initiation. This divergence is distinct from the promoter code used later by the developing oocytes and reveals genome-wide promoter remodelling during early female and male germline development.
Project description:We performed CAGE-Seq on dissected ovaries and testes as well as female and male carcasses of two species of Drosophila (D. melanogaster and D. pseudoobscura). These data are used to map transcription start sites. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf
Project description:We develop a method Re-Cappable-seq for determining eukaryotic transcription start sites derived from all RNA polymerases at nucleotide resolution. In particular, this method identifies the Pol-I and Pol-III TSSs, which are missing by CAGE. Applied to human A549 cell line, our method results in the identification of 33,468 and 5,269 high confidence Pol-II and non-Pol-II TSS respectively. Re-Cappable-seq identifies Pol-II TSS with higher specificity than CAGE.
Project description:Here we quantified the transcription start site usage in a WT strain (BY4741) and a ∆set2 strain associated with the appearence of cryptic transcription start sites.
Project description:CAGE-seq (Cap Analysis of Gene Expression coupled with deep sequencing) was used to map transcription start site (TSS) usage in human embryonic lung fibroblast (HELF) cells in response to infection with the human cytomegalovirus (HCMV) HAN strain. Total RNA from infected cells at 72 hours post-infection and mock-infected controls was profiled. The analysis detected 224,206 and 292,294 CAGE transcription start sites (CTSSs) in infected and mock samples, respectively, which grouped into 20,532 and 26,407 transcript clusters (TCs). Comparative analysis identified 3,302 consensus clusters with significant shifts in TSS positions, spanning 2,487 protein-coding genes and 134 non-coding RNAs. These findings reveal extensive infection-associated remodeling of promoter choice across the host transcriptome.