Comprehensive single-cell atlas of the mouse retina

Li, Jin; Choi, Jongsu; Cheng, Xuesen; Ma, Justin; Pema, Shahil; Sanes, Joshua R.; Mardon, Graeme; Frankfort, Benjamin J.; Tran, Nicholas M.; Li, Yumei; Chen, Rui

doi:10.1101/2024.01.24.577060

Cited by 3 publications

(2 citation statements)

References 46 publications

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“…Raw sequencing reads were processed using the CellRanger v6.1.2 (10X Genomics) pipeline, against the hg38 reference genome (https://cf.10xgenomics.com/supp/cell-exp/refdata-gex-GRCh38-2020-A.tar.gz). Then, quality control was performed separately for each sample through a quality control pipeline (https://github.com/lijinbio/cellqc) 25,26 . Real cells were filtered by dropkick v1.2.8 27 and ambient RNA was removed using SoupX v1.6.2 28 .…”

Section: Methodsmentioning

confidence: 99%

To be or not to be - Decoding the Trabecular Meshwork Cell Identity

Tian,

Baidouri,

Kim

et al. 2024

Preprint

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The trabecular meshwork within the conventional outflow apparatus is critical in maintaining intraocular pressure homeostasis.In vitrostudies employing primary cell cultures of the human trabecular meshwork (hTM) have conventionally served as surrogates for investigating the pathobiology of TM dysfunction. Despite its abundant use, translation of outcomes fromin vitrostudies toex vivoand/orin vivostudies remains a challenge. Given the cell heterogeneity, performing single-cell RNA sequencing comparing primary hTM cell cultures to hTM tissue may provide important insights on cellular identity and translatability, as such an approach has not been reported before. In this study, we assembled a total of 14 primary hTMin vitrosamples across passages 1-4, including 4 samples from individuals diagnosed with glaucoma. This dataset offers a comprehensive transcriptomic resource of primary hTMin vitroscRNA-seq data to study global changes in gene expression in comparison to cells in tissuein situ. We have performed extensive preprocessing and quality control, allowing the research community to access and utilize this public resource.

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Section: Methodsmentioning

confidence: 99%

To be or not to be - Decoding the Trabecular Meshwork Cell Identity

Tian,

Baidouri,

Kim

et al. 2024

Preprint

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“…18_Novel, along with other 'Novel' types described in Tran et al 2019 12 , were molecularly defined but have not been completely characterized. However, annotation of these four types was independently verified in a large-scale integrated analysis of mouse retinal scRNA-seq datasets 22 . In addition, we analyzed our clustered dataset to validate and revise cell-type specific markers.…”

Section: Rgc Atlas Version 11mentioning

confidence: 99%

Sample multiplexing for retinal single-cell RNA-sequencing

Ma,

Chu,

Polo Prieto

et al. 2024

Preprint

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Single-cell RNA sequencing (scRNA-seq) is a powerful tool for evaluating cell-type specific transcription but can be challenging to apply to rare cell populations. Efficient collection of low abundance cell types can be achieved by pooling biological samples, but this precludes resolution of sample origin together with phenotypic data. This limitation is particularly problematic in experiments in which biological or technical variation is expected to be high (e.g., disease models which vary in phenotypic severity, high throughput genetic perturbation screens, or human patient samples). One solution is sample multiplexing where each sample is tagged with a unique sequence barcode that is resolved bioinformatically. We have established a scRNA-seq sample multiplexing pipeline for mouse retinal ganglion cells (RGCs) using cholesterol-modified-oligos (CMOs). RGCs are a scarce and highly heterogenous cell type (~47 types) that represent less than 1% of all retinal cells. We found that CMOs labeled RGCs efficiently and did not have a significant impact on cell viability, gene expression, or cluster type representation. Through these assignments, we evaluated differential gene expression and identified transcriptional variance between RGCs of individual retinas both as a whole and within specific RGC types. Overall RGC type distributions and transcriptomic correlations between retinas were remarkably similar. Additionally, sample multiplexing enabled the identification of multiplets and resolution of key biological features such as sex-specific gene expression differences. In conclusion, we found that the addition of CMOs is not detrimental to scRNA-seq analysis and enabled the evaluation of differential gene expression among individual biological samples for a rare cell population. As single-cell transcriptomics are becoming a more widely used approach to research development and disease, sample multiplexing represents a useful method to enhance the precision of scRNA-seq analysis.

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Generation of an Armcx1 Conditional Knockout Mouse

Bright,

Bomze,

Bhaumik

et al. 2024

Genesis

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Armadillo repeat‐containing X‐linked protein‐1 (Armcx1) is a poorly characterized transmembrane protein that regulates mitochondrial transport in neurons. Its overexpression has been shown to induce neurite outgrowth in embryonic neurons and to promote retinal ganglion cell (RGC) survival and axonal regrowth in a mouse optic nerve crush model. In order to evaluate the functions of endogenous Armcx1 in vivo, we have created a conditional Armcx1 knockout mouse line in which the entire coding region of the Armcx1 gene is flanked by loxP sites. This Armcx1fl line was crossed with mouse strains in which Cre recombinase expression is driven by the promoters for β‐actin and Six3, in order to achieve deletion of Armcx1 globally and in retinal neurons, respectively. Having confirmed deletion of the gene, we proceeded to characterize the abundance and morphology of RGCs in Armcx1 knockout mice aged to 15 months. Under normal physiological conditions, no evidence of aberrant retinal or optic nerve development or RGC degeneration was observed in these mice. The Armcx1fl mouse should be valuable for future studies investigating mitochondrial morphology and transport in the absence of Armcx1 and in determining the susceptibility of Armcx1‐deficient neurons to degeneration in the setting of additional heritable or environmental stressors.

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Comprehensive single-cell atlas of the mouse retina

Cited by 3 publications

References 46 publications

To be or not to be - Decoding the Trabecular Meshwork Cell Identity

To be or not to be - Decoding the Trabecular Meshwork Cell Identity

Sample multiplexing for retinal single-cell RNA-sequencing

Generation of an Armcx1 Conditional Knockout Mouse

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