Identification And Characterization Of Box C/D SnoRNA Genes From Oryza Sativa

2 . - O - ribose methylation is one of the predominant types of nucleotide modification in rRNA . In order to map the specific methylation sites of rice 18S and 28S rRNAs, a series of primers was designed and synthesized to com plementary to both yeast and rice 18S/28S rRNAs. Primer extensions at different dNTP concentrations were carried out to detect the methylation sites of both yeast and rice rRNAs. The data showed that over 80% of yeast rRNA ribose methylation sites were con served with in rice rRNA . It was also observed that much more methylation sites were in plant rRN A than that in yeast rRNA. 16 novel ribose methylation sites were identified for the first time from rice 18S/28S rRNA. These data will provide important inform ation for screening new box C/D snoRNAs from rice genome. Based on the information of rice rRN A methylation sites, 48 different box C/D genes from 27 novel snoRN A clusters were identified by computational screening of Oryza sativa genome sequences, 25 of w hich have never been reported in any other plants. Among them, 7 snoRNAs were implicated to guide 2 . - O - ribose methylation sites, which were previously identified in human or yeast rRNAs but never in plant. The other 16 snoRNAs were implicated to guide comp letely novel 2 . - O - ribose methylation sites, which were identified by our lab. The remaining two novel snoRNAs lack any complementarity to rRNAs or snRNAs, implying that they might target other RN A species. All snoRNAs reported here were experimentally conf irmed by Northern blot and primer extension assa y . Polycistronic precursors from clusters were demonstrated by RT . PCR . It is highly likely each of the rice snoRN A gene clusters identified here is transcribed in polycistronic wa y . This indicates that the mo de of polycistronic expression of snoRN A clusters is widespread in plant.