The Effect of Gap Length on Double-Strand Break Repair in Drosophila

Johnson-Schlitz, Dena M.; Engels, William R.

doi:10.1534/genetics.106.059436

Cited by 7 publications

(4 citation statements)

References 29 publications

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“…It appears that in Drosophila, repair DNA synthesis tracts are limited to ,50 kb. This finding is also reported by an independent study that shows efficient filling of an 11-kb gap, inefficient filling of a 43-kb gap, and no complete filling of a 210-kb gap ( Johnson-Schlitz and Engels 2006). Such a limitation appears to be unlike that in yeast, where an entire chromosomal arm can be copied into a double-strand break by break-induced replication (Morrow et al 1997).…”

Section: Rsupporting

confidence: 62%

In Vivo Construction of Transgenes in Drosophila

et al. 2007

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Transgenic flies are generated by transposon-mediated transformation. A drawback of this approach is the size limit of transposable elements. Here, we propose a novel method that allows the extension of transgenes in vivo. This method is based on an incomplete transgene that has been constructed in vitro and integrated into the Drosophila genome by conventional transgenesis. The incomplete transgene contains two short stretches of DNA homologous to the 59-and 39-ends of a larger DNA segment of interest. Between the short stretches of homology an I-SceI recognition site is located. Once activated, I-SceI endonuclease introduces a DNA double-strand break, which triggers ectopic recombination between the stretches of homology and the endogenous locus. Through gap repair, the transgene obtains the complete region of interest in vivo. Our results show that this method is effective for copying up to 28 kb of genomic DNA into the transgene, thereby eliminating the technical difficulties associated with the in vitro construction of large transgenes and extending the size limits of current transgenesis protocols. In general, this method may be a useful technique for genetic engineering of eukaryotic model organisms.

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Section: Rsupporting

confidence: 62%

In Vivo Construction of Transgenes in Drosophila

et al. 2007

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“…Notably, the use of TLS polymerases as “first responders” might be particularly advantageous in instances where extensive synthesis might be unfavorable or energetically costly. As a corollary to this, large gaps that require extensive synthesis may be particularly difficult to repair by HR and may be ultimately repaired by end joining [36] .…”

Section: Discussionmentioning

confidence: 99%

Competition between Replicative and Translesion Polymerases during Homologous Recombination Repair in Drosophila

et al. 2012

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In metazoans, the mechanism by which DNA is synthesized during homologous recombination repair of double-strand breaks is poorly understood. Specifically, the identities of the polymerase(s) that carry out repair synthesis and how they are recruited to repair sites are unclear. Here, we have investigated the roles of several different polymerases during homologous recombination repair in Drosophila melanogaster. Using a gap repair assay, we found that homologous recombination is impaired in Drosophila lacking DNA polymerase zeta and, to a lesser extent, polymerase eta. In addition, the Pol32 protein, part of the polymerase delta complex, is needed for repair requiring extensive synthesis. Loss of Rev1, which interacts with multiple translesion polymerases, results in increased synthesis during gap repair. Together, our findings support a model in which translesion polymerases and the polymerase delta complex compete during homologous recombination repair. In addition, they establish Rev1 as a crucial factor that regulates the extent of repair synthesis.

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“…Although gaps of at least 44 kb can be repaired by SDSA, the efficiency of completed SDSA decreases with increasing gap size (Johnson-Schlitz and Engels 2006). In cases where the gap is not completely filled, EJ (primarily TMEJ) completes repair after variable amounts of synthesis from one or both ends (Adams et al 2003;Chan et al 2010).…”

Section: Relationship Between Hr and Ejmentioning

confidence: 99%

DNA Repair inDrosophila: Mutagens, Models, and Missing Genes

2017

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The numerous processes that damage DNA are counterbalanced by a complex network of repair pathways that, collectively, can mend diverse types of damage. Insights into these pathways have come from studies in many different organisms, including Drosophila melanogaster. Indeed, the first ideas about chromosome and gene repair grew out of Drosophila research on the properties of mutations produced by ionizing radiation and mustard gas. Numerous methods have been developed to take advantage of Drosophila genetic tools to elucidate repair processes in whole animals, organs, tissues, and cells. These studies have led to the discovery of key DNA repair pathways, including synthesis-dependent strand annealing, and DNA polymerase theta-mediated end joining. Drosophila appear to utilize other major repair pathways as well, such as base excision repair, nucleotide excision repair, mismatch repair, and interstrand crosslink repair. In a surprising number of cases, however, DNA repair genes whose products play important roles in these pathways in other organisms are missing from the Drosophila genome, raising interesting questions for continued investigations.

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The Effect of Gap Length on Double-Strand Break Repair in Drosophila

Cited by 7 publications

References 29 publications

In Vivo Construction of Transgenes in Drosophila

In Vivo Construction of Transgenes in Drosophila

Competition between Replicative and Translesion Polymerases during Homologous Recombination Repair in Drosophila

DNA Repair inDrosophila: Mutagens, Models, and Missing Genes

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