A critical requirement for research using model organisms is an appropriate, well-defined and consistent diet. There is currently no complete chemically defined (holidic) diet available for Drosophila melanogaster. We describe a holidic medium that is equal in performance to an oligidic diet optimized for adult fecundity and lifespan. It is also sufficient to support development over multiple generations, but at a reduced rate. During seven years of experiments, the holidic diet yielded more consistent experimental outcomes than oligidic food for adult fitness traits. Furthermore, nutrients and drugs are more accessible to flies in holidic medium and, similar to dietary restriction on oligidic food, amino acid dilution increases fly lifespan. We also report amino acid specific effects on food choice behavior and that folic acid from the microbiota is sufficient for development. These insights could not be gained using oligidic or meridic diets.

Using two-dimensional agarose gel electrophoresis, we show that mitochondrial DNA (mtDNA) replication of birds and mammals frequently entails ribonucleotide incorporation throughout the lagging strand (RITOLS). Based on a combination of two-dimensional agarose gel electrophoretic analysis and mapping of 5' ends of DNA, initiation of RITOLS replication occurs in the major non-coding region of vertebrate mtDNA and is effectively unidirectional. In some cases, conversion of nascent RNA strands to DNA starts at defined loci, the most prominent of which maps, in mammalian mtDNA, in the vicinity of the site known as the light-strand origin.

Recently, we presented evidence for conventional, strand-coupled replication of mammalian mitochondrial DNA. Partially single-stranded replication intermediates detected in the same DNA preparations were assumed to derive from the previously described, strand-asymmetric mode of mitochondrial DNA replication. Here, we show that bona fide replication intermediates from highly purified mitochondria are essentially duplex throughout their length, but contain widespread regions of RNA:DNA hybrid, as a result of the incorporation of ribonucleotides on the light strand which are subsequently converted to DNA. Ribonucleotide-rich regions can be degraded to generate partially single-stranded molecules by RNase H treatment in vitro or during DNA extraction from crude mitochondria. Mammalian mitochondrial DNA replication thus proceeds mainly, or exclusively, by a strand-coupled mechanism.