RNA-seq analysis of Wild Type and Mll5-/- Retinal Transcriptomes
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ABSTRACT:
PROVIDER: PRJNA807898 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR18057114_1.fastq.gz | Fastqsanger.gz | |||
| SRR18057114_2.fastq.gz | Fastqsanger.gz | |||
| SRR18057115_1.fastq.gz | Fastqsanger.gz | |||
| SRR18057115_2.fastq.gz | Fastqsanger.gz | |||
| SRR18057116_1.fastq.gz | Fastqsanger.gz |
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Project description:Quantitative Analysis of Wild Type and CTGF-/- Retinal Transcriptomes During Postnatal Vascular Development
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Project description:Purpose: The goal of this study is to compare the retinal transcriptomes of wild-type and CTGF-/- mice during postnatal development of the retinal vasculature. Results: The results from principal component analysis (PCA) showed a distinguishable gene expression profiling among the samples. The expression level (FPKM value) of known genes and transcripts were calculated using ballgown through the transcript abundances estimated with StringTie. The number of identified genes and transcripts per group was calculated based on the mean of FPKM in group > 0.5. We have identified 24,818 transcripts for 12946 gene count in WT mouse retinas and 24708 transcripts for 12852 gene count in knockout mouse retinas. The significantly enriched GO terms of the differentially expressed genes include those involved in protein, lipid and DNA metabolism, neurogenesis and signal transduction and gene regulation.
2020-03-13 | GSE146900 | GEO
Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived retinal transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis. Methods: Retinal mRNA profiles of 21-day-old wild-type (WT) and neural retina leucine zipper knockout (Nrl-/-) mice were generated by deep sequencing, in triplicate, using Illumina GAIIx. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays. Results: Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the mouse genome (build mm9) and identified 16,014 transcripts in the retinas of WT and Nrl−/− mice with BWA workflow and 34,115 transcripts with TopHat workflow. RNA-seq data confirmed stable expression of 25 known housekeeping genes, and 12 of these were validated with qRT–PCR. RNA-seq data had a linear relationship with qRT–PCR for more than four orders of magnitude and a goodness of fit (R2) of 0.8798. Approximately 10% of the transcripts showed differential expression between the WT and Nrl−/− retina, with a fold change ≥1.5 and p value <0.05. Altered expression of 25 genes was confirmed with qRT–PCR, demonstrating the high degree of sensitivity of the RNA-seq method. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to retinal function. Data analysis with BWA and TopHat workflows revealed a significant overlap yet provided complementary insights in transcriptome profiling. Conclusions: Our study represents the first detailed analysis of retinal transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions. Retinal mRNA profiles of 21-day old wild type (WT) and Nrl-/- mice were generated by deep sequencing, in triplicate, using Illumina GAIIx.
2011-10-25 | E-GEOD-33141 | biostudies-arrayexpress
Project description:Purpose: Identify Top2b-dependent genes by comparing retinal transcriptome profiling (RNA-seq) Methods: Retinal mRNA profiles of neonatal (postnatal day 0 and 6) wild-type (WT) and Topoisomerase IIbeta (Top2b) conditional knockout (Top2b−/−, or cKO) mice were generated using the SOLiD System (Applied Biosystems). Data analysis was performed according to published protocols (Trapnell et al., 2012) with minor modifications. Briefly, colorspace data of raw 50 bp reads was aligned to the mouse genome (mm9 or GRCm37) using Bowtie (Langmead et al., 2009). Gene expression levels were analyzed with Cufflinks, with differentially expressed genes determined by Cuffdiff (Trapnell et al., 2010). Results: After sequencing, ~120 million 50 bp reads for each time point and each group were obtained and mapped to the mouse genome (MGI, as of Feb 15, 2012). Over 62,000 transcripts were identified; these include ~22,000 annotated genes (including alternative-spliced variants) which cover 76% of the whole mouse genome. Among all the annotated genes, 8.80% (1,935/22,000) for P0 and 1.25% (274/22,000) for P6 showed differential expression between the control and cKO samples (p-value ≤ 0.05, q-value ≤ 0.05). Conclusions: All genes with differential expression levels in Top2b cKO samples are direct or indirect dependent to Top2b.
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Project description:Analysis of Wild Type and MFN2-/- Macrophage Transcriptomes
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Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived retinal transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis. Methods: Retinal mRNA profiles of 21-day-old wild-type (WT) and neural retina leucine zipper knockout (Nrl−/−) mice were generated by deep sequencing, in triplicate, using Illumina GAIIx. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks. qRT–PCR validation was performed using TaqMan and SYBR Green assays. Results: Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the mouse genome (build mm9) and identified 16,014 transcripts in the retinas of WT and Nrl−/− mice with BWA workflow and 34,115 transcripts with TopHat workflow. RNA-seq data confirmed stable expression of 25 known housekeeping genes, and 12 of these were validated with qRT–PCR. RNA-seq data had a linear relationship with qRT–PCR for more than four orders of magnitude and a goodness of fit (R2) of 0.8798. Approximately 10% of the transcripts showed differential expression between the WT and Nrl−/− retina, with a fold change ≥1.5 and p value <0.05. Altered expression of 25 genes was confirmed with qRT–PCR, demonstrating the high degree of sensitivity of the RNA-seq method. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to retinal function. Data analysis with BWA and TopHat workflows revealed a significant overlap yet provided complementary insights in transcriptome profiling. Conclusions: Our study represents the first detailed analysis of retinal transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.
2011-10-25 | GSE33141 | GEO