A Molecularly Defined Duplication Set for the X Chromosome of Drosophila melanogaster

Abstract

We describe a molecularly defined duplication kit for the X chromosome of Drosophila melanogaster. A set of 408 overlapping P[acman] BAC clones was used to create small duplications (average length 88 kb) covering the 22-Mb sequenced portion of the chromosome. The BAC clones were inserted into an attP docking site on chromosome 3L using FC31 integrase, allowing direct comparison of different transgenes. The insertions complement 92% of the essential and viable mutations and deletions tested, demonstrating that almost all Drosophila genes are compact and that the current annotations of the genome are reasonably accurate. Moreover, almost all genes are tolerated at twice the normal dosage. Finally, we more precisely mapped two regions at which duplications cause diplo-lethality in males. This collection comprises the first molecularly defined duplication set to cover a whole chromosome in a multicellular organism. The work presented removes a long-standing barrier to genetic analysis of the Drosophila X chromosome, will greatly facilitate functional assays of X-linked genes in vivo, and provides a model for functional analyses of entire chromosomes in other species.

Methods to study Drosophila immunity

“…Transposon mutagenesis, albeit suffering from insertion bias [72], allows for easy retrieval of positional information, and forms the basis for a downstream toolkit of genetic applications including imprecise excision knock-out, Gal4-UAS overexpression of flanking genes, or element replacement by targeting vectors, to name but a few [73][74][75][76][77]. Extensive libraries of P-element-based transposon insertions are available through stock centers, along with deletion and duplication lines [78][79][80][81]. Finally, targeted gene knock-out using optimized targeting plasmids in combination with CRISPR will greatly accelerate full KO coverage of the fly genome [82].…”

The sex locus is tightly linked to factors conferring sex-specific lethal effects in the mosquito Aedes aegypti

“…In males inheriting the recombinant m-chromosome, the gene would be present in a double dose (one copy on the M-chromosome and one copy on the m-chromosome; Figure 3c) that would lead to its overexpression and, in effect, embryonic lethality. A study on Drosophila, in which duplication of two short X chromosome regions led to male embryonic lethality (Venken et al, 2010), lends support to this scenario. (2) If a recessive embryonic-lethal gene is involved, males inheriting the maternal recombinant chromosome would be homozygous and inviable (Figure 3c).…”

Fruit Flies in Biomedical Research

“…For example, one approach is to start with many mutations identified in a forward genetic screen (Xiong et al 2012;Yamamoto et al 2012;Zhang et al 2013;Charng et al 2014), to map these mutants with new tools publically available in fly (Zhai et al 2003;Cook et al 2010b). The causative mutations can now be efficiently identified with WGS (Haelterman et al 2014) and rescued with genomic BACs (Venken et al 2009(Venken et al , 2010 to verify the specificity. Then the human homologs can be identified by their sequence homology and studied in thousands of human exomes (Bamshad et al 2012;Yamamoto et al 2014a).…”