Characterization of Cre recombinase  mouse lines enabling cell type‐specific targeting of postnatal intervertebral discs

AbstractThe basolateral amygdalar complex (BLA) is implicated in behaviors ranging from fear acquisition to addiction. Optogenetic methods have enabled the association of circuit-specific functions to uniquely connected BLA cell types. Thus, a systematic and detailed connectivity profile of BLA projection neurons to inform granular, cell type-specific interrogations is warranted. Here, we apply machine-learning based computational and informatics analysis techniques to the results of circuit-tracing experiments to create a foundational, comprehensive BLA connectivity map. The analyses identify three distinct domains within the anterior BLA (BLAa) that house target-specific projection neurons with distinguishable morphological features. We identify brain-wide targets of projection neurons in the three BLAa domains, as well as in the posterior BLA, ventral BLA, posterior basomedial, and lateral amygdalar nuclei. Inputs to each nucleus also are identified via retrograde tracing. The data suggests that connectionally unique, domain-specific BLAa neurons are associated with distinct behavior networks.

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Cytometric Profiling of CD133+ Cells in Human Colon Carcinoma Cell Lines Identifies a Common core Phenotype and Cell Type-specific Mosaics

The International Journal of Biological Markers ◽

10.5301/jbm.5000020 ◽

2013 ◽

Vol 28 (3) ◽

pp. 267-273 ◽

Cited By ~ 11

Author(s):

Marica Gemei ◽

Rosa Di Noto ◽

Peppino Mirabelli ◽

Luigi Del Vecchio

Keyword(s):

Colorectal Cancer ◽

Stem Cells ◽

Cancer Stem Cells ◽

Colon Carcinoma ◽

Cell Lines ◽

Carcinoma Cell ◽

Cell Type ◽

Carcinoma Cell Lines ◽

Cell Type Specific

In colorectal cancer, CD133+ cells from fresh biopsies proved to be more tumorigenic than their CD133– counterparts. Nevertheless, the function of CD133 protein in tumorigenic cells seems only marginal. Moreover, CD133 expression alone is insufficient to isolate true cancer stem cells, since only 1 out of 262 CD133+ cells actually displays stem-cell capacity. Thus, new markers for colorectal cancer stem cells are needed. Here, we show the extensive characterization of CD133+ cells in 5 different colon carcinoma continuous cell lines (HT29, HCT116, Caco2, GEO and LS174T), each representing a different maturation level of colorectal cancer cells. Markers associated with stemness, tumorigenesis and metastatic potential were selected. We identified 6 molecules consistently present on CD133+ cells: CD9, CD29, CD49b, CD59, CD151, and CD326. By contrast, CD24, CD26, CD54, CD66c, CD81, CD90, CD99, CD112, CD164, CD166, and CD200 showed a discontinuous behavior, which led us to identify cell type-specific surface antigen mosaics. Finally, some antigens, e.g. CD227, indicated the possibility of classifying the CD133+ cells into 2 subsets likely exhibiting specific features. This study reports, for the first time, an extended characterization of the CD133+ cells in colon carcinoma cell lines and provides a “dictionary” of antigens to be used in colorectal cancer research.

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Characterization of cell type-specific S100B expression in the mouse olfactory bulb

Cell Calcium ◽

10.1016/j.ceca.2020.102334 ◽

2021 ◽

Vol 94 ◽

pp. 102334

Author(s):

Xin Su ◽

Tamara Vasilkovska ◽

Nicole Fröhlich ◽

Olga Garaschuk

Keyword(s):

Olfactory Bulb ◽

Cell Type ◽

Cell Type Specific

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Identification and characterization of cell type-specific and ubiquitous chromatin regulatory structures in the human genome

PLoS Genetics ◽

10.1371/journal.pgen.0030136.eor ◽

2005 ◽

Vol preprint (2007) ◽

pp. e136

Author(s):

Hualin Xi ◽

Hennady P Shulha ◽

Jane M Lin ◽

Teresa R Vales ◽

Yutao Fu ◽

...

Keyword(s):

Human Genome ◽

Cell Type ◽

Cell Type Specific ◽

Identification And Characterization

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Zebrafish Cre/lox regulated UFlip alleles generated by CRISPR/Cas targeted integration provide cell-type specific conditional gene inactivation

10.1101/2021.06.18.448732 ◽

2021 ◽

Author(s):

Maira P. Almeida ◽

Sekhar Kambakam ◽

Fang Liu ◽

Zhitao Ming ◽

Jordan M. Welker ◽

...

Keyword(s):

Gene Function ◽

Cre Recombinase ◽

Gene Trap ◽

Gene Inactivation ◽

Loss Of Function ◽

Cell Type ◽

Active Gene ◽

Indel Mutation ◽

Targeted Integration ◽

Cell Type Specific

The ability to regulate gene activity spatially and temporally is essential to investigate cell type specific gene function during development and in postembryonic processes and disease models. The Cre/lox system has been widely used for performing cell and tissue-specific conditional analysis of gene function in zebrafish, but simple and efficient methods for isolation of stable, Cre/lox regulated alleles are lacking. Here we applied our GeneWeld CRISPR/Cas9 short homology-directed targeted integration strategy to generate floxed conditional alleles that provide robust gene knockdown and strong loss of function phenotypes. A universal targeting vector, UFlip, with sites for cloning short 24-48 bp homology arms flanking a floxed mRFP gene trap plus secondary reporter cassette, was integrated into an intron in hdac1, rbbp4, and rb1. Active, gene off orientation hdac1-UFlip-Off and rb1-UFlip-Off integration alleles result in >99% reduction of gene expression in homozygotes and recapitulate known indel loss of function phenotypes. Passive, gene on orientation rbbp4-UFlip-On and rb1-UFlip-On integration alleles do not cause phenotypes in trans-heterozygous combination with an indel mutation. Cre recombinase injection leads to recombination at alternating pairs of loxP and lox2272 sites, inverting and locking the cassette into the active, gene off orientation, and the expected mutant phenotypes. In combination with our endogenous neural progenitor Cre drivers we demonstrate rbbp4-UFlip-On and rb1-UFlip-On gene inactivation phenotypes can be restricted to specific neural cell populations. Replacement of the UFlip mRFP primary reporter gene trap with a 2A-RFP in rbbp4-UFlip-Off, or 2A-KalTA4 in rb1-UFlip-Off, shows strong RFP expression in wild type or UAS:RFP injected embryos, respectively. Together these results validate a simplified approach for efficient isolation of highly mutagenic Cre/lox responsive conditional gene alleles to advance zebrafish Cre recombinase genetics.

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