Skandalis et al. 



JESO Volume 139, 2008 



FIGURE 1. DL2 cell cultures 72h post infection (400x magnification), a) Mock-infected 

 DL2 b) FHV-infected DL2. 



were evident in the infected cells 72 h post inoculation (Fig. lb), at which point the 

 infected cells aggregated into a mass and became individually indistinct. By contrast the 

 mock-infected control DL2 cell culture showed no differences in morphology or growth 

 pattern compared to cells grown under normal conditions (Fig. la). The morphological 

 changes of the infected cells were consistent with cytopathic cell death and no 

 morphological changes associated with apoptosis were observed. 



Cloning of Oggl 



The Drosophila Oggl gene consists of 4 exons and 3 introns totaling 1476 

 nucleotides. The spliced Oggl transcript is 1298 nucleotides in length. In our analysis of 

 alternative Oggl transcripts, the small size of the introns represented a potential problem 

 since it was possible to amplify genomic Oggl sequences directly from contaminating 

 genomic DNA rather than from reverse transcribed mRNA. To avoid this problem, all 

 RNA preparations were treated with DNAse to degrade any contaminating genomic DNA 

 and then each RNA preparation was tested by PCR to confirm that it could not support 

 Oggl amplification without reverse transcribing mRNA (Fig. 2b). 



Splice Variant Analysis 



The effect of FHV infection on DL2 splicing was assessed at 8, 12, 20, and 

 44 hrs post FHV inoculation. Based on previous observations, approximately 95% 

 of the cells are infected 8 h following inoculation with maximal viral RNA synthesis 

 occurring approximately 20 h post inoculation. Amplified Oggl transcript sequences were 

 successfully obtained from each time point (Fig. 2a) and cloned into E.coli DH5a cells. 



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