8580a3f6-7eff-4466-b29a-f4d538346820 - samples
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PROVIDER: EGAD00001007586 | EGA |
REPOSITORIES: EGA
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Project description:Purpose: Development of specific and precise RNA base editing toolkit. Methods: The transcriptome-wide off-target sites identification, annotation, and transcriptome-wide genes regulation of each editor were analyzed by RNA-seq data. The raw reads (.fastq) were first processed using Trim Glore (v0.6.7) to remove adapter sequences with default parameters. Then the trimmed data were used to remove rRNA data by Bowtie2 (v2.5.0) with key parameters –no-unal -I 1 -X 1000. The cleaned reads data were then aligned to the indexed human GRCh38 reference genome by STAR (v2.7.1a) with key parameters --outSAMstrandField intronMotif --outFilterIntronMotifs RemoveNoncanonical. The aligned BAMs were added RG tag of sample information, sorted, indexed, stated by Samtools (v1.15.1) with default parameters. Then the processed BAMs were removed duplications by the Picard Markduplicates tool (v2.23.9) with key parameters ASO=coordinate REMOVE_DUPLICATES=true. Then remove duplicate BAMs were used to filter reads containing clips by Samtools with default parameters. All the cleaned BAMs were downsampled to an equivalent level for subsequent analysis to ensure a fair comparison of transcriptome-wide off-targets. The uniquely mapped reads in final BAMs were used to calculate gene expression level by Cuffdiff (v2.2.1) with default parameters. Finally, we filtered the differential expression genes using cutoff |Log2FoldChange| > 1 and P-value < 0.05, based on the Cuffdiff quantification results. Results: Our new programmable RNA base editing toolkits have high editing efficiency, flexible editing scope and high transcriptome-wide specificity. Conclusions: We deleveped verstaile RNA editing toolkits enabling presice, specific and versatile RNA base editing.
2024-10-31 | GSE230140 | GEO
Project description:Methods: RNA-Seq was used to identify differentially expressed genes between wild type strains, ∆rrpA and ∆rrpB mutants at mid-log (~6 hrs) of growth. 11168H wild type, 11168H∆rrpA, 11168H∆rrpB and 11168H∆rrpAB were plated out on BA plates and incubated at 37°C under microaerobic conditions for 6 hrs. RNA was extracted from the pelleted bacteria cells using PureLink™ RNA Mini Kit (Invitrogen), following manufactures protocol. RNA was isolated from five independent experiments performed on different days Ribosomal RNA was depleted using Ribominus (Invitrogen).Libraries were prepared using TruSeq® Stranded mRNA (Illumina).The paired-end reads were trimmed and filtered using Sickle v1.200 by using a sliding window approach and trimming the reads where the average base quality drops below 20. Only the reads that were above 10 bp length were kept after trimming. Bowtie2 was used to map the reads against the reference sequences; C. jejuni strains 11168H assembly GCA_900117385.1. This generated the mapping in SAM format which were converted to BAM format using Samtools and were index sorted. We used gffread from cufflinks suite to convert annotations from GFF to GTF format. These were used in Bedtools (with multicov-bams option)with the mapped reads to generate transcript counts per samples. A shell utility by the authorswas then used to collate all these transcripts into a n=20 samples x p=1,628 transcripts for 11168H strain. Statistical analysis was performed in R using the combined data generated from the bioinformatics as well as meta data associated with the study (multifactorial design). We used DESeq DataSet from Matrix()function fromDESeq2 package with the adjusted p-value significance cut-off of 0.05 and log fold change cut-off of 1.5. This function uses negative binomial GLM to obtain maximum likelihood estimates for the transcripts log fold change between the two conditions.
2021-09-30 | GSE174333 | GEO
Project description:Purpose:To understand the change in transcriptome for the overxepression of ΔCSP in HepG2 cells. Methods:Total RNAs of ΔCSP stable expressing or control HepG2 cells were extracted using TRIzol (Invitrogen), following the manufacturer's instructions. RNA-seq and bioinformatic data analysis were performed by Shanghai Sangon Bio Ltd. Briefly, sequencing libraries were generated using VAHTSTM mRNA-seq V2 Library Prep Kit for Illumina® (Vazyme Biotech, Nanjing, China) following manufacturer's recommendations and were sequenced on HiSeq XTen sequencers (Illumina, San Diego, CA). Raw reads in FASTQ format were subjected to quality control using FastQC. RNA-seq reads were aligned to the reference genome using Bowtie. Uniquely mapped reads were used for further analysis. Gene expression levels are expressed as TPM (Transcripts per million reads) and differences in gene expression were calculated with rSeq. Results:There were 492 genes differentially expressed in ΔCSP-overexpressing HepG2 cells compare to the control group respectively (fold change >2 or < 0.5).
2020-12-31 | GSE129323 | GEO
Project description:Rhizophora mucronata Lam., a prevalent mangrove variety of Indo-Pacific region is reported to defy saline stress up to 40 ppt, but the genome or transcriptome behind this tolerance is yet to be investigated. As an initiative to create a reference sequence database, we have forged a set of 46,366,348 paired end RNA-Seq raw reads of Rhizophora mucronata Lam. leaf tissues from Illumina HiSeq 2500 platform (SRA study accession SRP093200 ; Bioproject accession PRJNA345155). All possible gene transcripts were then reconstructed from the RNA raw seq data and 93960 Trinity assembled, annotated transcripts that are being actively expressed at a given time is proposed (TSA accession GGEC00000000). To estimate gene transcript expression, we used Bowtie 2 programme and successfully aligned back up to 95.14% of the filtered reads to the assembled transcriptome. We allowed up to 1-mismatches in the seed region (length =31bp) and all multiple mapped position were reported. Of all filtered reads about 95.14% of reads from each sample were properly aligned back to the assembled transcriptome. Overall we found 52,153 unique transcripts which have expression >=1 FPKM.
2018-03-22 | GSE112162 | GEO
Project description:Purpose:To understand the change in cellular metabolism and function for the overxepression of SCL27A5 in hepatoma cells. Methods:Total RNAs of AdSLC27A5- or AdGFP-infected PLC/PRF/5 cells were extracted using TRIzol (Invitrogen), following the manufacturer’s instructions. RNA-seq and bioinformatic data analysis were performed by Shanghai Novel Bio Ltd. Briefly, strand-specific RNA-seq libraries were prepared using the Total RNA-seq (H/M/R) Library Prep Kit (Vazyme Biotech, Nanjing, China) and were sequenced on a HiSeq X Ten sequencing platform. Raw reads in FASTQ format were subjected to quality control using FastQC. RNA-seq reads were aligned to the reference genome using Bowtie. Uniquely mapped reads were used for further analysis. Gene expression levels are expressed as RPKM (reads per kilobase per million reads) and differences in gene expression were calculated with rSeq. Results:There were 17 genes differentially expressed in AdSLC27A5-infected PLC/PRF/5 cells compare to the GFP control group (fold change >1.5 or < 0.667; FDR < 0.05).
2023-07-01 | GSE117823 | GEO
Project description:Purpose:To understand the change in cellular metabolism and function for the overxepression of GSTZ1 or PCK1 in hepatoma cells. Methods:Total RNAs of AdGFP- , AdGSTZ1-, or AdPCK1-infected Huh7 cells were extracted using TRIzol (Invitrogen), following the manufacturer’s instructions. RNA-seq and bioinformatic data analysis were performed by Shanghai Novel Bio Ltd. Briefly, strand-specific RNA-seq libraries were prepared using the Total RNA-seq (H/M/R) Library Prep Kit (Vazyme Biotech, Nanjing, China) and were sequenced on Ion Torrent Proton sequencing platform. Raw reads in FASTQ format were subjected to quality control using FastQC. RNA-seq reads were aligned to the reference genome using Bowtie. Uniquely mapped reads were used for further analysis. Gene expression levels are expressed as RPKM (reads per kilobase per million reads) and differences in gene expression were calculated with rSeq. Results:There were 498 genes differentially expressed in AdPCK1-infected Huh 7 cells, 513 genes differentially expressed in AdGSTZ1-infected Huh 7 cells compare to the GFP control group respectively (fold change >1.5 or < 0.667; FDR < 0.05).
2019-01-02 | GSE117822 | GEO
Project description:Purpose: Perform RNA-seq study on infectious bovine endometrial tissues to reveal important genes and biological pathways regulating uterine physiology following uterine infections Methods:RNA sequencings were done using Illumina platform. Single-end reads in the FASTQ format were explored using FastQC, low-quality reads were trimmed from both 3’ and 5’ ends until a base pair of Phred quality score of 30 (99.9% accurate) or greater was found, reads having a mean quality score less than 30 and length below 30 nucleotides were filtered out. Cleaned reads were aligned against the bovine reference genome (Bos_taurus.ARS-UCD1.2) using HiSAT2. The resulting SAM files were sorted, converted to BAM files using SAMtools. Read counts mapped to bovine gene models were generated using htseq-count script from HTSeq package. Bioconductor DESeq2 was used to get the differentially expressed genes among infectious vs normal uterine tract groups Conclusions: The study demonstrated that uterine infections altered several genes and pathways related to inflammatory response, immune response, uterine physiology, uterine enviroment and fertility in the intercaruncular region of bovine endometrium.
2022-07-28 | GSE209700 | GEO
Project description:Purpose: RNase Y is a major enzyme responsible for mRNA degradation in Streptococcus pyogenes. The goals of this study are to understand whether RNase Y plays a role in operon transcription of S. pyogenes NZ131 by using RNA-seq analysis. Methods: S. pyogenes mRNA profiles of wild type (WT) and RNase Y mutant (∆rny) were generated by deep sequencing, in duplicate, using Illumina Hiseq 2000. The sequence reads were aligned to the S. pyogenes genome using Bowtie2. The aligned files were sorted to BAM format and indexed using Samtools. The read depth of each base was derived from BAM files using BEDtools. Operon organization of S. pyogenes WT and ∆rny strains were predicted based on base reads. Results: A total of 11 to 12 billion reads were obtained from each sample. More than 99% of these reads were mapped to the S. pyogenes genome. Predictions of operon organization using WT and ∆rny samples showed little difference between the two strains. Conclusions: Our result shows that the mutation of RNase Y does not affect the operon organization of S. pyogenes NZ131.
2017-06-02 | GSE99533 | GEO
Project description:We sequenced 14 mouse tissues' small RNA samples from C57BL/6J mouse and aligned the sequenced reads to miRBase_v16 to profile miRNA expression levels in these 14 tissues. We collect several tissue samples from 10 C57BL/6J mouse. Use high-throughout sequence technology to profile miRNA expression level in each tissue.
2016-04-06 | E-GEOD-67885 | biostudies-arrayexpress
Project description:When GPC transcription factor in hexaploid wheat is down-regulated by stable RNA interference (GPC RNAi), senescence is significantly delayed and grain protein content together with the overall nutrient partitioning to the grain is greatly reduced. mRNA-seq was used to catalogue the genes that are regulated by the GPC transcription factor during monocarpic senescence. cDNA was prepared from wild type bread wheat plants and GPC RNAi plants 12 days after anthesis and sequenced by Illumina. Four biological replications per genotypes were sequenced. To determine gene expression levels, reads were aligned to a reference transcriptome generated by assemblying 454-reads obtained from the same biological material (454 assembled sequences: TSA project 59945 - accession numbers: HP608076-HP639668).
2011-09-23 | E-GEOD-25759 | biostudies-arrayexpress