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Dual ifgMosaic: A Versatile Method for Multispectral and Combinatorial Mosaic Gene-Function Analysis.

Cell | 2017

Improved methods for manipulating and analyzing gene function have provided a better understanding of how genes work during organ development and disease. Inducible functional genetic mosaics can be extraordinarily useful in the study of biological systems; however, this experimental approach is still rarely used in vertebrates. This is mainly due to technical difficulties in the assembly of large DNA constructs carrying multiple genes and regulatory elements and their targeting to the genome. In addition, mosaic phenotypic analysis, unlike classical single gene-function analysis, requires clear labeling and detection of multiple cell clones in the same tissue. Here, we describe several methods for the rapid generation of transgenic or gene-targeted mice and embryonic stem (ES) cell lines containing all the necessary elements for inducible, fluorescent, and functional genetic mosaic (ifgMosaic) analysis. This technology enables the interrogation of multiple and combinatorial gene function with high temporal and cellular resolution.

Pubmed ID: 28802047 RIS Download

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This is a list of tools and resources that we have found mentioned in this publication.


Addgene (tool)

RRID:SCR_002037

Non-profit plasmid repository dedicated to helping scientists around the world share high-quality plasmids. Facilitates archiving and distributing DNA-based research reagents and associated data to scientists worldwide. Repository contains over 65,000 plasmids, including special collections on CRISPR, fluorescent proteins, and ready-to-use viral preparations. There is no cost for scientists to deposit plasmids, which saves time and money associated with shipping plasmids themselves. All plasmids are fully sequenced for validation and sequencing data is openly available. We handle the appropriate Material Transfer Agreements (MTA) with institutions, facilitating open exchange and offering intellectual property and liability protection for depositing scientists. Furthermore, we curate free educational resources for the scientific community including a blog, eBooks, video protocols, and detailed molecular biology resources.

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BacPac Resources Center (tool)

RRID:SCR_007215

It is the distribution arm of their academic laboratory. They operate on a cost-recovery mechanism in order to make the resources generated in their laboratory available to the academic scientific community. While clones and screening services are widely available, library arrays are primarily available to researchers with a scientific need to analyze most clones in the library. This site contains information on currently available BAC and PAC genomic DNA libraries, BAC Clones, PAC Clones, Fosmid Clones, cDNA collections, high-density colony hybridization filters, and BAC and PAC cloning vectors. Protocols used in our laboratory for the hybridization-based screening of colony filters, purification of BAC and PAC DNA, and end-sequencing methodologies, are also provided. BPRC does not list clones, for two reasons: 1)most clones have not been characterized and lack specific data. 2)all clones are part of libraries and all clones from a particular library share common characteristics. Hence, to find out if BPRC has a particular clone, one needs either use Automatic Clone Validation or else find out if the clone is compatible with the range of clone names for a corresponding clone library. Typically (although not always), clone names are derived from the library name. BPRC uses the NCBI-recommended clone nomenclature & library nomenclature. Most arrayed libraries are available in frozen microtiter dish format to academic and non-academic users provided that there is a scientific need for complete-library access. (for instance to annotate, modify or analyze all BAC clones as part of a genome project).

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BacPac Resources Center (tool)

RRID:SCR_001520

It is the distribution arm of their academic laboratory. They operate on a cost-recovery mechanism in order to make the resources generated in their laboratory available to the academic scientific community. While clones and screening services are widely available, library arrays are primarily available to researchers with a scientific need to analyze most clones in the library. This site contains information on currently available BAC and PAC genomic DNA libraries, BAC Clones, PAC Clones, Fosmid Clones, cDNA collections, high-density colony hybridization filters, and BAC and PAC cloning vectors. Protocols used in our laboratory for the hybridization-based screening of colony filters, purification of BAC and PAC DNA, and end-sequencing methodologies, are also provided. BPRC does not list clones, for two reasons: 1)most clones have not been characterized and lack specific data. 2)all clones are part of libraries and all clones from a particular library share common characteristics. Hence, to find out if BPRC has a particular clone, one needs either use Automatic Clone Validation or else find out if the clone is compatible with the range of clone names for a corresponding clone library. Typically (although not always), clone names are derived from the library name. BPRC uses the NCBI-recommended clone nomenclature & library nomenclature. Most arrayed libraries are available in frozen microtiter dish format to academic and non-academic users provided that there is a scientific need for complete-library access. (for instance to annotate, modify or analyze all BAC clones as part of a genome project).

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C57BL/6J (tool)

RRID:IMSR_JAX:000664

Mus musculus with name C57BL/6J from IMSR.

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Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb (antibody)

RRID:AB_2315112

This monoclonal targets Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204)

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MS CD144 PURE MAB 0.5MG 11D4.1 (antibody)

RRID:AB_395707

This monoclonal targets CD144 (VE-Cadherin)

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Anti-Sox2 (antibody)

RRID:AB_2286686

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ERG antibody [EPR3863] (antibody)

RRID:AB_10864794

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anti-tRFP (antibody)

RRID:AB_2571743

This polyclonal targets Full-length recombinant denatured and non-denatured TagRFP comprising 6XHis tag

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MOUSE ANTI V5-TAG:Alexa Fluor 488 (antibody)

RRID:AB_770155

This monoclonal targets Viral V5-TAG

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Living Colors Full-length GFP Polyclonal Antibody (antibody)

RRID:AB_2336883

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Fiji (software resource)

RRID:SCR_002285

Software package as distribution of ImageJ and ImageJ2 together with Java, Java3D and plugins organized into coherent menu structure. Used to assist research in life sciences.

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OP9 (cell line)

RRID:CVCL_4398

Cell line OP9 is a Stromal cell line with a species of origin Mus musculus

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Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP® Rabbit mAb (antibody)

RRID:AB_2315112

This monoclonal targets Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204)

View all literature mentions

MS CD144 PURE MAB 0.5MG 11D4.1 (antibody)

RRID:AB_395707

This monoclonal targets CD144 (VE-Cadherin)

View all literature mentions

Anti-Sox2 (antibody)

RRID:AB_2286686

This polyclonal targets SOX2

View all literature mentions

ERG antibody [EPR3863] (antibody)

RRID:AB_10864794

This monoclonal targets ERG antibody [EPR3863]

View all literature mentions

anti-tRFP (antibody)

RRID:AB_2571743

This polyclonal targets Full-length recombinant denatured and non-denatured TagRFP comprising 6XHis tag

View all literature mentions

MOUSE ANTI V5-TAG:Alexa Fluor 488 (antibody)

RRID:AB_770155

This monoclonal targets Viral V5-TAG

View all literature mentions

Living Colors Full-length GFP Polyclonal Antibody (antibody)

RRID:AB_2336883

This polyclonal targets Full length recombinant GFP

View all literature mentions