Serial Recombineering Cloning to Build Selectable and Tagged Genomic P[acman] Bac Clones for Selection Transgenesis and Functional Gene Analysis Using Drosophila melanogaste

Authors

Koen J T Venken
Nick Matinyan
Yezabel Gonzalez
Herman A Dierick

Publication Date

2-1-2023

Journal

BMC Biology

DOI

10.1186/s12915-020-0740-7

PMID

31987035

PMCID

PMC6986101

PubMedCentral® Posted Date

1-27-2020

PubMedCentral® Full Text Version

Post-print

Published Open-Access

yes

Keywords

Animals, Drosophila melanogaster, Animals, Genetically Modified, Gene Transfer Techniques, DNA, Drosophila, Genomics, Cloning, Molecular, Clone Cells

Abstract

Transgenes with genomic DNA fragments that encompass genes of interest are the gold standard for complementing null alleles in rescue experiments in the fruit fly Drosophila melanogaster. Of particular interest are genomic DNA clones available as bacterial artificial chromosomes (BACs) or fosmids from publicly available genomic DNA libraries. Genes contained within BAC and fosmid clones can be easily modified by recombineering cloning to insert peptide or protein tags to localize, visualize, or manipulate gene products, and to create point mutations or deletions for structure-function analysis of the inserted genes. However, since transgenesis efficiency is inversely correlated with transgene size, obtaining transgenic animals for increasingly larger BAC and fosmid clones requires increasingly laborious screening efforts using the transgenesis marker commonly used for these transgenes, the dominant eye color marker white

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