RNA-sequencing from single nuclei
Rashel V. Grindberg,Joyclyn Yee-Greenbaum,Michael J. McConnell,Mark Novotny,Andrew O'Shaughnessy,Georgina M. Lambert,Marcos J. Araúzo-Bravo,Jun Lee,Max Fishman,Gillian E. Robbins,Xiaoying Lin,Pratap Venepally,Jonathan H. Badger,David W. Galbraith,Fred H. Gage,Roger S. Lasken +15 more
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TLDR
In this paper, the authors demonstrate that this method can be done using the extremely low levels of mRNA in a single nucleus, isolated from a mouse neural progenitor cell line and from dissected hippocampal tissue.Abstract:
It has recently been established that synthesis of double-stranded cDNA can be done from a single cell for use in DNA sequencing. Global gene expression can be quantified from the number of reads mapping to each gene, and mutations and mRNA splicing variants determined from the sequence reads. Here we demonstrate that this method of transcriptomic analysis can be done using the extremely low levels of mRNA in a single nucleus, isolated from a mouse neural progenitor cell line and from dissected hippocampal tissue. This method is characterized by excellent coverage and technical reproducibility. On average, more than 16,000 of the 24,057 mouse protein-coding genes were detected from single nuclei, and the amount of gene-expression variation was similar when measured between single nuclei and single cells. Several major advantages of the method exist: first, nuclei, compared with whole cells, have the advantage of being easily isolated from complex tissues and organs, such as those in the CNS. Second, the method can be widely applied to eukaryotic species, including those of different kingdoms. The method also provides insight into regulatory mechanisms specific to the nucleus. Finally, the method enables dissection of regulatory events at the single-cell level; pooling of 10 nuclei or 10 cells obscures some of the variability measured in transcript levels, implying that single nuclei and cells will be extremely useful in revealing the physiological state and interconnectedness of gene regulation in a manner that avoids the masking inherent to conventional transcriptomics using bulk cells or tissues.read more
Citations
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Comprehensive single-cell transcriptional profiling of a multicellular organism
Junyue Cao,Jonathan S. Packer,Vijay Ramani,Darren A. Cusanovich,Chau Huynh,Ray A. M. Daza,Xiaojie Qiu,Choli Lee,Scott N. Furlan,Scott N. Furlan,Scott N. Furlan,Frank J. Steemers,Andrew Adey,Andrew Adey,Robert H. Waterston,Cole Trapnell,Jay Shendure,Jay Shendure +17 more
TL;DR: The authors profiled almost 50,000 single cells from an individual Caenorhabditis elegans larval stage and were able to identify and recover information from different, even rare, cell types and develop combinatorial indexing strategies to profile the transcriptomes of single cells or nuclei.
Journal ArticleDOI
The technology and biology of single-cell RNA sequencing.
Aleksandra A. Kolodziejczyk,Aleksandra A. Kolodziejczyk,Jong Kyoung Kim,Valentine Svensson,John C. Marioni,John C. Marioni,Sarah A. Teichmann,Sarah A. Teichmann +7 more
TL;DR: Recently developed single-cell mRNA-sequencing methods enable unbiased, high-throughput, and high-resolution transcriptomic analysis of individual cells, which provides an additional dimension to transcriptomic information relative to traditional methods that profile bulk populations of cells.
Journal ArticleDOI
Massively-parallel single nucleus RNA-seq with DroNc-seq
Naomi Habib,Naomi Habib,Inbal Avraham-Davidi,Anindita Basu,Tyler Burks,Karthik Shekhar,Matan Hofree,Sourav Choudhury,Sourav Choudhury,François Aguet,Ellen Gelfand,Kristin Ardlie,David A. Weitz,Orit Rozenblatt-Rosen,Feng Zhang,Feng Zhang,Aviv Regev,Aviv Regev +17 more
TL;DR: In this paper, a massively parallel single-nucleus RNA sequencing (sNuc-seq) with droplet technology is proposed. But it does not provide high throughput, and it is not suitable for high-dimensional data.
Journal ArticleDOI
Neuronal subtypes and diversity revealed by single-nucleus RNA sequencing of the human brain.
Blue B. Lake,Rizi Ai,Gwendolyn E. Kaeser,Gwendolyn E. Kaeser,Neeraj Salathia,Yun C. Yung,Rui Liu,Andre Wildberg,Derek Gao,Ho-Lim Fung,Song Chen,Raakhee Vijayaraghavan,Julian Wong,Allison Chen,Xiaoyan Sheng,Fiona Kaper,Richard Shen,Mostafa Ronaghi,Jian-Bing Fan,Wei Wang,Jerold Chun,Kun Zhang +21 more
TL;DR: A scalable approach to sequence and quantify RNA molecules in isolated neuronal nuclei from a postmortem brain, generating 3227 sets of single-neuron data from six distinct regions of the cerebral cortex demonstrates a robust and scalable method for identifying and categorizing single nuclear transcriptomes.
Journal ArticleDOI
In vivo interrogation of gene function in the mammalian brain using CRISPR-Cas9
Lukasz Swiech,Matthias Heidenreich,Abhishek Banerjee,Naomi Habib,Yinqing Li,John J. Trombetta,Mriganka Sur,Feng Zhang +7 more
TL;DR: In this paper, an adeno-associated viral (AAV)-associated endonuclease (Cas)9 was used to edit single or multiple genes in replicating eukaryotic cells, resulting in frame-shifting insertion/deletion (indel) mutations and subsequent protein depletion.
References
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