High-Throughput Mapping of Single-Neuron Projections by Sequencing of Barcoded RNA

Kebschull, Justus M.; Silva, Pedro Garcia da; Reid, Ashlan P.; Peikon, Ian D.; Albeanu, Dinu F.; Zador, Anthony M.

doi:10.1016/j.neuron.2016.07.036

Cited by 307 publications

(306 citation statements)

References 60 publications

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“…We cut out the left auditory cortex from the brain and post-fixed it in 10% formalin at 4 for 8 hrs, and snap-froze the rest of the brain on a razor blade on dry ice. The snap-frozen brain (without the injection site) was then processed for conventional MAPseq as described 5 . The postfixed auditory cortex was cryo-protected, embedded, and cryo-sectioned as described above.…”

Section: Animals and Tissue Processingmentioning

confidence: 99%

“…For validation of BaristaSeq in brain slices, we used a barcode library previously described by Kebschull et al 5 . The library contained 1.5 million known 30-nt random barcode sequences, which represented ~97% of all barcodes in the library.…”

Section: Viruses Constructs and Oligosmentioning

confidence: 99%

“…Sample slice images from the brain XC9 after dissection indicating the locations of the collected projection sites are available at Dryad (see Data and software availability in Methods). The projection sites were sequenced as described for MAPseq 5 .…”

Section: Barseqmentioning

confidence: 99%

“…As a first step toward this goal we began with MAPseq 4,5 (Fig. 1A, left), a sequencing-based method capable of mapping long-range projections of thousands of single neurons in a single brain.…”

mentioning

confidence: 99%

“…To evaluate barcode sequencing in brain slices, we sequenced 25 bases in a sample infected with a diverse (>10 6 ) pool of barcoded Sindbis virus 5,15 injected in the auditory cortex (See Methods; Fig. 2B).…”

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confidence: 99%

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High-throughput mapping of long-range neuronal projection using in situ sequencing

Chen

Sun

Zhan

et al. 2018

Preprint

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SummaryUnderstanding neural circuits requires deciphering the interactions of myriad cell types defined by anatomy, spatial organization, gene expression, and functional properties. Resolving these cell types requires both single neuron resolution and high-throughput, a combination that is challenging to achieve with conventional anatomical methods. Here we introduce BARseq, a method for mapping the projections of thousands of spatially resolved neurons by combining the high throughput of DNA sequencing with the high spatial resolution of microscopy. We used BARseq to determine the projections of 1309 neurons in mouse auditory cortex to 11 targets. We observed 264 distinct projection patterns. Hierarchical clustering confirmed the major classical classes of projection neurons, segregated across cortical laminae. Further analysis revealed 25 subclasses, largely intermingled across laminae. Unlike cell types defined by gene expression, projection subclasses beyond the major classes were rarely enriched in specific laminae, raising the possibility that the organization of projection patterns in mature neurons is orthogonal to that of gene expression. In this way, downstream brain areas could receive information from multiple cell types through parallel pathways. By sequencing in situ, BARseq has the potential to bridge anatomical, transcriptomic, functional, and other approaches at single neuron resolution with high throughput, and thereby offer unprecedented insight of the structure and function of a neural circuit.. CC-BY-NC 4.0 International license It is made available under a was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.The copyright holder for this preprint (which . http://dx.doi.org/10.1101/294637 doi: bioRxiv preprint first posted online Apr. 3, 2018; 2 An important challenge in neuroscience is to relate diverse characteristics of single neurons, in a co-registered fashion, within single brains 1 . Even simultaneous co-registration of two characteristics can be challenging, and has led to insights about the functional organization of neural circuits 2,3 . A high-throughput method capable of such multimodal co-registration would yield a "Rosetta Brain"-an integrative dataset that could constrain theoretical efforts to bridge across levels of structure and function in the nervous system 1 .As a first step toward this goal we began with MAPseq 4,5 (Fig. 1A, left), a sequencing-based method capable of mapping long-range projections of thousands of single neurons in a single brain. MAPseq achieves multiplexing by uniquely labeling individual neurons with random RNA sequences, or "barcodes". Because MAPseq, like most other sequencing methods, relies on tissue homogenization, it cannot resolve the spatial organization of the neuronal somata. This spatial organization, however, potentially allows the registration of distinct neuronal characteristics. We therefore sought to develop a method that would preserve the spatial organization of barc...

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Section: Animals and Tissue Processingmentioning

confidence: 99%

Section: Viruses Constructs and Oligosmentioning

confidence: 99%

Section: Barseqmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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High-throughput mapping of long-range neuronal projection using in situ sequencing

Chen

Sun

Zhan

et al. 2018

Preprint

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Generation of Projection Neuron Diversity in the Neocortex

Uzquiano,

Arlotta

2023

Neocortical Neurogenesis in Development and Evolution

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Vom Design der Moleküle des Lebens zum Design von Leben: Zukünftige Anwendungen von DNA‐Technologien

et al. 2018

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Seit der Aufklärung ihrer Struktur steht die DNA im Zentrum der biologischen Forschung. In den letzten 50 Jahren ist die Entwicklung von DNA‐Technologien rasant vorangeschritten, insbesondere im Bereich der DNA‐Sequenzierung. Parallel dazu wurden auch gewaltige Fortschritte in der DNA‐Synthese und DNA‐Editierung erzielt, die sich von der Oligonukleotid‐ bis zur Genom‐Ebene erstrecken. In diesem Aufsatz erörtern wir vier verschiedene Teilbereiche von DNA‐Technologien und folgen dabei diesem Verlauf von kleinerem zu größerem Maßstab. Wir beginnen mit dem Aufbau von Materialien aus DNA, die wiederum als Transportsysteme in vivo dienen können. Anschließend diskutieren wir, wie die Modifizierung mikrobieller Genome zu neuartigen Methoden für die Produktion industrieller Biologika führen kann. Als nächstes reden wir über die Zukunft der Genomsynthese als eine Methode für die Evolutionsforschung. Schließlich erläutern wir, wie das Barcoding biologischer Systeme deren dreidimensionale Analyse in hoch parallelisierter Weise ermöglicht.

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High-Throughput Mapping of Single-Neuron Projections by Sequencing of Barcoded RNA

Cited by 307 publications

References 60 publications

High-throughput mapping of long-range neuronal projection using in situ sequencing

High-throughput mapping of long-range neuronal projection using in situ sequencing

Generation of Projection Neuron Diversity in the Neocortex

Vom Design der Moleküle des Lebens zum Design von Leben: Zukünftige Anwendungen von DNA‐Technologien

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