Rapid generation of nested chromosomal deletions on mouse chromosome 2

David F LePage; Deanna M Church; Elise Millie; Terry J Hassold; Ronald A Conlon

doi:10.1073/pnas.97.19.10471

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Rapid generation of nested chromosomal deletions on mouse chromosome 2

Journal article

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Rapid generation of nested chromosomal deletions on mouse chromosome 2

David F LePage, Deanna M Church, Elise Millie, Terry J Hassold and Ronald A Conlon

Proceedings of the National Academy of Sciences - PNAS, Vol.97(19), pp.10471-10476

09/12/2000

DOI: https://doi.org/10.1073/pnas.97.19.10471

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https://hdl.handle.net/2376/118279

PMCID: PMC27048

PMID: 10984539

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Abstract

Biological Sciences

Nested chromosomal deletions are powerful genetic tools. They are particularly suited for identifying essential genes in development either directly or by screening induced mutations against a deletion. To apply this approach to the functional analysis of mouse chromosome 2, a strategy for the rapid generation of nested deletions with Cre recombinase was developed and tested. A loxP site was targeted to the Notch1 gene on chromosome 2. A targeted line was cotransfected with a second loxP site and a plasmid for transient expression of Cre. Independent random integrations of the second loxP site onto the targeted chromosome in direct repeat orientation created multiple nested deletions. By virtue of targeting in an F 1 hybrid embryonic stem cell line, F 1 (129S1×Cast/Ei), the deletions could be verified and rapidly mapped. Ten deletions fell into seven size classes, with the largest extending six or seven centiMorgans. The cytology of the deletion chromosomes were determined by fluorescent in situ hybridization. Eight deletions were cytologically normal, but the two largest deletions had additional rearrangements. Three deletions, including the largest unrearranged deletion, have been transmitted through the germ line. Several endpoints also have been cloned by plasmid rescue. These experiments illustrate the means to rapidly create and map deletions anywhere in the mouse genome. They also demonstrate an improved method for generating nested deletions in embryonic stem cells.

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Title: Rapid generation of nested chromosomal deletions on mouse chromosome 2
Creators: David F LePage - Department of Genetics, Case Western Reserve University School of Medicine and University Hospitals of Cleveland, 10900 Euclid Avenue, Cleveland, OH 44106-4955; and
Deanna M Church - Department of Genetics, Case Western Reserve University School of Medicine and University Hospitals of Cleveland, 10900 Euclid Avenue, Cleveland, OH 44106-4955; and
Elise Millie - Department of Genetics, Case Western Reserve University School of Medicine and University Hospitals of Cleveland, 10900 Euclid Avenue, Cleveland, OH 44106-4955; and
Terry J Hassold - Department of Genetics, Case Western Reserve University School of Medicine and University Hospitals of Cleveland, 10900 Euclid Avenue, Cleveland, OH 44106-4955; and
Ronald A Conlon - Department of Genetics, Case Western Reserve University School of Medicine and University Hospitals of Cleveland, 10900 Euclid Avenue, Cleveland, OH 44106-4955; and
Publication Details: Proceedings of the National Academy of Sciences - PNAS, Vol.97(19), pp.10471-10476
Academic Unit: Molecular Biosciences, School of
Publisher: The National Academy of Sciences
Identifiers: 99900547974001842
Language: English
Resource Type: Journal article