Project description:These data correspond to one SMRT cell sequencing run (performed on Sequel II, PacBio) of full length cDNAs from 3 pooled glioma stem cell line libraries. No tag was added to distinguish the 3 different samples
Project description:BACKGROUND: In order to contribute to the establishment of a complete map of transcribed regions of the human genome, we constructed a testicular cDNA library for the cynomolgus monkey, and attempted to find novel transcripts for identification of their human homologues. RESULT: The full-insert sequences of 512 cDNA clones were determined. Ultimately we found 302 non-redundant cDNAs carrying open reading frames of 300 bp-length or longer. Among them, 89 cDNAs were found not to be annotated previously in the Ensembl human database. After searching against the Ensembl mouse database, we also found 69 putative coding sequences have no homologous cDNAs in the annotated human and mouse genome sequences in Ensembl.We subsequently designed a DNA microarray including 396 non-redundant cDNAs (with and without open reading frames) to examine the expression of the full-sequenced genes. With the testicular probe and a mixture of probes of 10 other tissues, 316 of 332 effective spots showed intense hybridized signals and 75 cDNAs were shown to be expressed very highly in the cynomolgus monkey testis, but not ubiquitously. CONCLUSIONS: In this report, we determined 302 full-insert sequences of cynomolgus monkey cDNAs with enough length of open reading frames to discover novel transcripts as human homologues. Among 302 cDNA sequences, human homologues of 89 cDNAs have not been predicted in the annotated human genome sequence in the Ensembl. Additionally, we identified 75 dominantly expressed genes in testis among the full-sequenced clones by using a DNA microarray. Our cDNA clones and analytical results will be valuable resources for future functional genomic studies. Keywords: other
Project description:We tried two methods which are the DNase I treated full-length double-strand cDNA sequencing and the poly(A) capture full-length double-strand cDNA sequencing to avoid the non-specific genomic DNA amplification.
Project description:While numerous studies have described the transcriptomes of EVs in different cellular contexts, these efforts have typically relied on sequencing methods requiring RNA fragmentation, which limits interpretations on the integrity and isoform diversity of EV-encapsulated RNA populations. Furthermore, it has been assumed that mRNA signatures in EVs are likely to be fragmentation products of the cellular mRNA material, and little is known about the extent to which full-length mRNAs are present within EVs. Using Oxford nanopore long-read RNA sequencing, we sought to characterize the full-length polyadenylated (poly-A) transcriptome of EVs released by human chronic myelogenous leukemia K562 cells. We detected 441 and 280 RNAs that were respectively enriched or depleted in EVs. EV-enriched poly-A transcripts consist of a variety of biotypes, including mRNAs, long non-coding RNAs, and pseudogenes. Our analysis revealed that 12.72% of all reads present in EVs corresponded to known full-length transcripts, 65.34% of which were mRNAs. We also observed that for many well-represented coding and non-coding genes, diverse full-length transcript isoforms were present in EV specimens, and these isoforms were reflective-of but often in different ratio compared to cellular samples. Here we report a full-length transcriptome from human EVs, as determined by long-read nanopore sequencing.