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Synthetic promoter AAVs for cell-type specific expression in retinal cells

Posted by Beth Kenkel on Mar 12, 2020 9:15:00 AM

Adeno-associated virus (AAV) is a popular tool for gene delivery, but it has a drawback: how do you ensure your gene goes where you want it to? Knowing that a gene is expressed in a particular cell type is important not only for translational research, such as gene therapy, but also basic research. To improve cell-type specificity of AAV, work has focused on modifying the outside protein shell, or capsid, of the virus so that it only enters and delivers it cargo to certain cell types. 

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Topics: Viral Vectors, Cell Tracing, AAV

Single-cell tracking of lineage and identity with CellTag

Posted by Beth Kenkel on May 9, 2019 9:15:18 AM

Direct reprogramming describes the process where differentiated cells are turned into a cell type of choice while bypassing the intermediate pluripotent state. Though a valuable tool for regenerative medicine, direct reprogramming is an inefficient process, with the majority of cells failing to develop the desired identity.

The development of single cell technologies, such as single cell RNA-seq (scRNA-seq), now allows scientists to identify the gene expression patterns unique to the few cells that successfully reprogram. Unfortunately scRNA-seq only captures the gene expression of cells at a single time point, making it difficult to investigate how early expression patterns influence reprogramming outcomes.
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Topics: Viral Vectors, Cell Tracing

Multicolor Animals: Using Fluorescent Proteins to Understand Single Cell Behavior

Posted by Aliyah Weinstein on Mar 5, 2019 8:08:52 AM

Stochastic multicolor labeling is a popular technique in neuroscience and developmental biology. This type of cell labeling technique involves the introduction of a transgene construct containing fluorescent proteins (XFP) of different colors to label an organ or entire organism. Because each cell can have multiple copies of the transgene that will recombine independently, cells may acquire one of a variety of colors when a combination of XFP are expressed. Each cell remains the same color for its entire lifetime and daughter cells retain the same color, allowing for the fate mapping of cell populations over time. The ability to track single cell dynamics at the organism level has been made possible by tools that allow cells to become persistently fluorescent during development. Stochastic multicolor labeling systems, many based on Brainbow, now exist for a variety of species, cell types, and research applications.

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Topics: Fluorescent Proteins, Cell Tracing, Neuroscience

Tetbow: Bright Multicolor Labeling for Neuronal Tracing

Posted by Guest Blogger on Jan 24, 2019 9:24:20 AM

This post was contributed by Richi Sakaguchi from Kyoto University, and Marcus N. Leiwe and Takeshi Imai from Kyushu University.

Stochastic multicolor labeling is a powerful solution for discriminating between neurons for light microscopy-based neuronal reconstruction. To achieve stochastic multicolor labeling, Brainbow used the Cre-loxP system to express one of the three fluorescent protein (XFP) genes in a transgene. When multiple copies of the transgene cassette are introduced, stochasticity will result in a combinatorial expression of these three genes with different copy numbers, producing dozens of color hues (Livet et al., 2007; Cai et al., 2013). However, the brightness of Brainbow was inherently low. This is because the stochastic and combinatorial expression of fluorescent proteins is only possible at low copy number ranges, resulting in low fluorescent protein level.

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Topics: Fluorescent Proteins, Cell Tracing

Neuronal labeling with Spaghetti Monster

Posted by Benoit Giquel on Aug 14, 2018 8:42:01 AM

The central nervous system (CNS) orchestrates complex processes enabling organisms to control their movements and behavior. These functions and others are controlled by collections of neurons that are intricately wired into circuits through synaptic connections (Shepherd, 2004). Understanding the structure and function of these neural circuits is essential for neuroscience research. The use of genetic tools for visualizing and perturbing circuits together with the development of methodologies to deliver genes to the CNS have recently made it much easier to map these neuronal networks.

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Topics: Viral Vectors, Cell Tracing, Neuroscience

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