Fast and efficient Drosophila melanogaster gene knock-ins using MiMIC transposons.

ABSTRACT: Modern molecular genetics studies necessitate the manipulation of genes in their endogenous locus, but most of the current methodologies require an inefficient donor-dependent homologous recombination step to locally modify the genome. Here we describe a methodology to efficiently generate Drosophila knock-in alleles by capitalizing on the availability of numerous genomic MiMIC transposon insertions carrying recombinogenic attP sites. Our methodology entails the efficient PhiC31-mediated integration of a recombination cassette flanked by unique I-SceI and/or I-CreI restriction enzyme sites into an attP-site. These restriction enzyme sites allow for double-strand break-mediated removal of unwanted flanking transposon sequences, while leaving the desired genomic modifications or recombinatio

SUBMITTER: Vilain S 

PROVIDER: S-EPMC4267933 | biostudies-literature | 2014 Oct

REPOSITORIES: biostudies-literature