Trove directx buffer error fix 2016
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RSEM: accurate transcript quantification from RNA-seq data with or without a reference genome. Quantitative assessment of single-cell RNA-sequencing methods. Isolation of nuclei from mammalian tissues through the use of Triton X-100. A method for isolating intact mitochondria and nuclei from the same homogenate, and the influence of mitochondrial destruction on the properties of cell nuclei. The RIN: an RNA integrity number for assigning integrity values to RNA measurements. Isolation of nuclei from animal cells in culture. Isolation of neuronal chromatin from brain tissue. Jiang, Y., Matevossian, A., Huang, H.S., Straubhaar, J. Microarray analysis of cytoplasmic versus whole cell RNA reveals a considerable number of missed and false positive mRNAs. Genome-wide analyses show that nuclear and cytoplasmic RNA levels are differentially affected by dioxin. Transcriptional maps of 10 human chromosomes at 5-nucleotide resolution. Comparison of the contributions of the nuclear and cytoplasmic compartments to global gene expression in human cells. Trypsin-induced proteome alteration during cell subculture in mammalian cells. Cell types in the mouse cortex and hippocampus revealed by single-cell RNA-seq. A survey of human brain transcriptome diversity at the single cell level. Single-neuron isolation for RNA analysis using pipette capture and laser capture microdissection. Single-neuron RNA-seq: technical feasibility and reproducibility. Comprehensive qPCR profiling of gene expression in single neuronal cells. Transcriptome in vivo analysis (TIVA) of spatially defined single cells in live tissue. RNA-seq analysis to capture the transcriptome landscape of a single cell. Full-length mRNA-seq from single-cell levels of RNA and individual circulating tumor cells. Full-length RNA-seq from single cells using Smart-seq2. Global single-cell cDNA amplification to provide a template for representative high-density oligonucleotide microarray analysis. CEL-Seq: single-cell RNA-seq by multiplexed linear amplification. By following this procedure, it takes about 4 d to construct cDNA libraries that are ready for sequencing. The method also allows investigation of biological features unique to nuclei, such as enrichment of certain transcripts and precursors of some noncoding RNAs.
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Nuclear transcriptomes can be obtained from postmortem human brain tissue stored at −80 ☌, making brain archives accessible for RNA-seq from individual neurons. We isolate nuclei at 4 ☌ from tissue homogenates, which cause minimal damage.
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Previous single-cell approaches for RNA-seq from tissues include cell dissociation using protease treatment at 30 ☌, which is known to alter the transcriptome. Some steps follow published methods (Smart-seq2 for cDNA synthesis and Nextera XT barcoded library preparation) and are not described in detail here. Nuclei are isolated from specimens and sorted by FACS, cDNA libraries are constructed and RNA-seq is performed, followed by data analysis. A protocol is described for sequencing the transcriptome of a cell nucleus.