From the Cover: Directed, efficient, and versatile modifications of the Drosophila genome by genomic engineering.
With the completion of genome sequences of major model organisms, increasingly sophisticated genetic tools are necessary for investigating the complex and coordinated functions of genes. Here we describe a genetic manipulation system termed "genomic engineering" in Drosophila. Genomic engineering is a 2-step process that combines the ends-out (replacement) gene targeting with phage integrase phiC31-mediated DNA integration. First, through an improved and modified gene targeting method, a founder knock-out line is generated by deleting the target gene and replacing it with an integration site of phiC31. Second, DNA integration by phiC31 is used to reintroduce modified target-gene DNA into the native locus in the founder knock-out line. Genomic engineering permits directed and highly efficient modifications of a chosen genomic locus into virtually any desired mutant allele. We have successfully applied the genomic engineering scheme on 6 different genes and have generated at their loci more than 70 unique alleles.
Department of Cell Biology and Physiology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA.
Juan
J
Huang
Wenke
W
Zhou
Wei
W
Dong
Annie M
AM
Watson
Yang
Y
Hong
D000483:Alleles; D000818:Animals; D001435:Bacteriophages / Q000235:genetics; D001483:Base Sequence; D004330:Drosophila / Q000235:genetics*; D018390:Gene Targeting; D005818:Genetic Engineering / Q000379:methods*; D049750:Genome, Insect / Q000235:genetics*; D019426:Integrases; D008969:Molecular Sequence Data; D009154:Mutation; D016662:Virus Integration
D019426:Integrases