Alternative mRNA splice variants in Drosophila 



JESO Volume 139, 2008 



LDC8122044N 



a) b) 



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FIGURE 2. Oggl PCR products, a) From left 200 Q- 

 to right: Molecular size marker (MM), mock- 

 infected control (C), 8 h post-infection, 12 h 1031 

 post-infection, 20 h post-infection , 44 h post- 

 infection, and no-template PCR control, b) 

 PCR using RNA samples as template to assess 

 contamination by genomic DNA. 



Individual cloned transcripts from each stage of infection were analyzed by restriction 

 digest analysis and sequenced to determine the type and numbers of splice variants 

 generated by Oggl . 



Overall, our analysis of Oggl splice variants in Drosophila revealed five different 

 transcripts (labeled SV1-5) in addition to the wild type (WT), all of which maintained 

 some or all introns (Fig. 3). However, different combinations of splice variants were 

 observed under different conditions. Characterization of 95 transcripts recovered from 

 mock-infected DL2 revealed, in addition to WT, two types of splice variants: SV4, 

 which retained introns 3 (frequency 1.1%) and SV2, which retained introns 2 and 3 

 (frequency 1 .1%) (Fig. 4). Eight hours following FHV inoculation the frequency of splice 

 variants increased to 15% (60 transcripts characterized). Of the three splice variants 

 characterized, two were not detected in the control cells: SV1, which was completely un- 

 spliced, retaining all introns (frequency 6.7%) and SV3 that retained introns one and two 

 (frequency 3.3%). The third splice variant observed, SV4, was also present in the mock- 

 infected control and at 8 hrs increased to 5%. Twelve hours post-infection the overall 

 frequency of splice variants was less than at eight hours and similar to the frequency 

 observed in uninfected cells (2.2 % of 95 transcripts characterized). The transcripts 

 detected were SV4 and a novel variant, SV5, which retained intron 2. Twenty hours 

 post-infection only one variant was detectable, SV5, with a frequency of 2.7% (N=74 

 transcripts). Finally, 44 hours post-infection the frequency of splice variants was elevated 

 to 5.3% (N=95 transcripts) which was approximately double the frequency observed in 

 the mock-infected cells but still 3-fold lower than the frequency observed 8 hours post- 

 infection. The splice variants detected were SV1 (1.1%), SV2 (1.1%), SV3 (1.1%), and 

 SV4 (2.1%) (Fig. 4). 



Then frequencies of splice variants at 12 and 20 hrs were similar to the ones 

 observed in mock-infected controls, but the types of variants differ (Fig. 4). Splicing 

 of intron 2 exhibited a unique dependence on splicing of intron 1 . In the hundreds of 



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