Selective Methylation of One of the Two Promoters Residing in T-DNA Inserted in a Retroelement of Rice

摘要

The upstream sequence of pinb previously isolated from rice and confirmed to be a wound-inducible promoter by detecting GUS in TO transgenic rice transformed via Agrobacterium tumefaciens-mediated procedures. In a transgenic line (pinb-16), the selectable marker hptII driven by CaMV35S promoter was completely silenced in T2 sublines; but the uidA gene driven by pinb promoter was expressed without being affected, though it, together with hptll, exists in the same T-DNA insertion. Analyses of methylation patterns using bisulphite-sequencing in the homozygous T1 and T2 sublines showed that cytosines in CaMV35S were gradually methylated in T1 plants and almost completely methylated in T2 plants. Interestingly, the process of methylation was accompaniedby the occurrence of lesion mimic phenotype in rice leaves. The activity of hygromycin-resistance could be reestablished by treatment with 5-azacytidine. Genomic Southern and isolation of the T-DNA flanking sequences indicated that T-DNA was inserted ina retroelement of rice. These results revealed that methylation shows preference for the heterogeneity promoter fragment in the transgenic rice line and may be induced by the retroelement.