Next we analyzed to what extent the competition between the luciferase genome segments is affected by characteristics of the genome segments themselves. An obvious difference between the firefly and Gaussia genome segments is their gene length as the firefly and Gaussia luciferase genes consist of 1653 and 558 nucleotides, respectively. Small genome segments are likely to be replicated faster than long ones. To test this hypothesis, we generated an extended Gaussia luciferase gene construct, in which part of the firefly luciferase gene (39-terminal half) was inserted immediately behind the stop codon of the Gaussia luciferase gene in the GNP plasmid (referred to as GFsNP, Fig. 1A) to produce a genome segment with exactly the same length as the firefly luciferase genome segment. The extended and the normal Gaussia luciferase reporter segments were compared for their ability to compete with the firefly luciferase reporter segment. The absolute Gaussia luciferase expression level from the GFsNP segment was lower than that from the GNP segment (Fig. S4B ), probably resulting from its extended length, for which is corrected by plotting normalized Fluc/Gluc ratios in Figure 4 . These norm-Fluc/Gluc ratios (Fig. 4A) indicate that the extended GFsNP segment is still a strong competitor of the FNP segment, although much less efficient than the smaller GNP segment. We conclude that segment size is an important factor in the competition between vRNA segments. However, our results also suggest that coding regions are important as FNP and GFsNP segments do not differ in size, while they contain identical 59 and 39UTRs.