Rapid and efficient cloning of proviral flanking fragments by kanamycin resistance gene complementation | Zendy

Boris Fehse | Zendy; Klaus Kühlcke | Zendy; Andreas Langer | Zendy; Wolfram Ostertag | Zendy; Heinz Lother | Zendy

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Rapid and efficient cloning of proviral flanking fragments by kanamycin resistance gene complementation

Author(s) -

Boris Fehse,

Klaus Kühlcke,

Andreas Langer,

Wolfram Ostertag,

Heinz Lother

Publication year - 1999

Publication title -

nucleic acids research

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 9.008

H-Index - 537

eISSN - 1362-4954

pISSN - 0305-1048

DOI - 10.1093/nar/27.2.706

Subject(s) - biology , complementation , genetics , 5' flanking region , cloning (programming) , kanamycin , gene , molecular cloning , virology , promoter , peptide sequence , gene expression , phenotype , computer science , programming language

We have developed a technique for the rapid cloning of unknown flanking regions of transgenic DNA. We complemented a truncated kanamycin resistance gene of a bacterial plasmid with a neomycin resistance gene fragment from a gene transfer vector. Optimized transformation conditions allowed us to directly select for kanamycin-resistant bacteria. We cloned numerous proviral flanking fragments from growth factor-independent cell mutants that were obtained after infection with a replication incompetent retroviral vector and identified integrations into the cyclin D2 and several unknown genomic sequences. We anticipate that our method could be adapted to various vector systems that are used to tag and identify genes and to map genomes.

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