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Two-Step Bacterial Artificial Chromosome (BAC) Engineering: Cloning of the A and B Homology Arms into the Shuttle Vector.

TitleTwo-Step Bacterial Artificial Chromosome (BAC) Engineering: Cloning of the A and B Homology Arms into the Shuttle Vector.
Publication TypeJournal Article
Year of Publication2020
AuthorsHeintz N, Gong S
JournalCold Spring Harb Protoc
Volume2020
Issue4
Pagination098053
Date Published2020 04 01
ISSN1559-6095
Abstract

This protocol describes the preparation of the shuttle vector before its introduction into bacterial artificial chromosome (BAC) host cells for BAC two-step engineering. The homology arm sequences, prepared previously, are introduced by ligation into the digested shuttle vector DNA to provide sites for recombination within the BAC clone. Crude lysates of individual bacterial transformants serve as templates in polymerase chain reaction (PCR) analysis to confirm the presence of the homology arms in the recombinant shuttle vector.

DOI10.1101/pdb.prot098053
Alternate JournalCold Spring Harb Protoc
PubMed ID32238591