Systematic generation of high-resolution deletion coverage of the Drosophila melanogaster genome.

In fruit fly research, chromosomal deletions are indispensable tools for mapping mutations, characterizing alleles and identifying interacting loci. Most widely used deletions were generated by irradiation or chemical mutagenesis. These methods are labor-intensive, generate random breakpoints and result in unwanted secondary mutations that can confound phenotypic analyses. Most of the existing deletions are large, have molecularly undefined endpoints and are maintained in genetically complex stocks. Furthermore, the existence of haplolethal or haplosterile loci makes the recovery of deletions of certain regions exceedingly difficult by traditional methods, resulting in gaps in coverage. Here we describe two methods that address these problems by providing for the systematic isolation of targeted deletions in the D. melanogaster genome. The first strategy used a P element-based technique to generate deletions that closely flank haploinsufficient genes and minimize undeleted regions. This deletion set has increased overall genomic coverage by 5-7%. The second strategy used FLP recombinase and the large array of FRT-bearing insertions described in the accompanying paper to generate 519 isogenic deletions with molecularly defined endpoints. This second deletion collection provides 56% genome coverage so far. The latter methodology enables the generation of small custom deletions with predictable endpoints throughout the genome and should make their isolation a simple and routine task.

A complementary transposon tool kit for Drosophila melanogaster using P and piggyBac.

With the availability of complete genome sequence for Drosophila melanogaster, one of the next strategic goals for fly researchers is a complete gene knockout collection. The P-element transposon, the workhorse of D. melanogaster molecular genetics, has a pronounced nonrandom insertion spectrum. It has been estimated that 87% saturation of the approximately 13,500-gene complement of D. melanogaster might require generating and analyzing up to 150,000 insertions. We describe specific improvements to the lepidopteran transposon piggyBac and the P element that enabled us to tag and disrupt genes in D. melanogaster more efficiently. We generated over 29,000 inserts resulting in 53% gene saturation and a more diverse collection of phenotypically stronger insertional alleles. We found that piggyBac has distinct global and local gene-tagging behavior from that of P elements. Notably, piggyBac excisions from the germ line are nearly always precise, piggyBac does not share chromosomal hotspots associated with P and piggyBac is more effective at gene disruption because it lacks the P bias for insertion in 5' regulatory sequences.

Health and safety practices in the nanomaterials workplace: results from an international survey.

This article reports the findings of an international survey of nanomaterials firms and laboratories regarding their environmental health and safety (EHS) programs, engineering controls, personal protective equipment (PPE), exposure monitoring, waste disposal, product stewardship, and risk beliefs. While many participants reported not believing that nanomaterials pose special risks, nanospecific EHS programs were still widely reported. Most nanospecific EHS programs appeared to build from general EHS programs but included nanospecific workplace engineering controls and recommendations for clothing, gloves, eye protection, and respirators. Organizations with nanospecific EHS programs also reported providing product (safe use) guidance to consumers. However, workplace monitoring and nanospecific waste disposal were uneven and were only associated with the subset of organizations believing in special risks. A majority of organizations expressed a need for more toxicological information and EHS guidance. Overall, this study suggests that nanomaterials firms and laboratories are already attentive to nanospecific EHS and product stewardship issues. However, improved risk communication is needed to further the implementation of related programs. Organizations that are wholly inattentive to EHS would likely engage in nanospecific EHS upon implementing a staffed, general EHS program.