Yeast Single-cell RNA-seq, Cell by Cell and Step by Step
Single-cell RNA-seq (scRNA-seq) has become an established method for uncovering the intrinsic complexity within populations. Even within seemingly homogenous populations of isogenic yeast cells, there is a high degree of heterogeneity that originates from a compact and pervasively transcribed genome...
| Autores: | , , , , , , , |
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| Tipo de recurso: | artículo |
| Estado: | Versión publicada |
| Fecha de publicación: | 2019 |
| País: | España |
| Institución: | Universidad de Barcelona |
| Repositorio: | Dipòsit Digital de la UB |
| OAI Identifier: | oai:diposit.ub.edu:2445/220565 |
| Acceso en línea: | https://hdl.handle.net/2445/220565 |
| Access Level: | acceso abierto |
| Palabra clave: | Transcripció genètica Llevats Genetic transcription Yeast |
| Sumario: | Single-cell RNA-seq (scRNA-seq) has become an established method for uncovering the intrinsic complexity within populations. Even within seemingly homogenous populations of isogenic yeast cells, there is a high degree of heterogeneity that originates from a compact and pervasively transcribed genome. Research with microorganisms such as yeast represents a major challenge for single-cell transcriptomics, due to their small size, rigid cell wall, and low RNA content per cell. Because of these technical challenges, yeast-specific scRNA-seq methodologies have recently started to appear, each one of them relying on different cell-isolation and library-preparation methods. Consequently, each approach harbors unique strengths and weaknesses that need to be considered. We have recently developed a yeast single-cell RNA-seq protocol (yscRNA-seq), which is inexpensive, high-throughput and easy-to-implement, tailored to the unique needs of yeast. yscRNA-seq provides a unique platform that combines single-cell phenotyping via index sorting with the incorporation of unique molecule identifiers on transcripts that allows to digitally count the number of molecules in a strand- and isoform-specific manner. Here, we provide a detailed, step-by-step description of the experimental and computational steps of yscRNA-seq protocol. This protocol will ease the implementation of yscRNA-seq in other laboratories and provide guidelines for the development of novel technologies. |
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