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Title: Novel DNA methylation targets in Arabidopsis thaliana
Author: Gentry, Matthew Steven
Awarding Body: University of Leeds
Current Institution: University of Leeds
Date of Award: 2012
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The initiation of DNA methylation in Arobidopsis is controlled by the RNA-directed DNA methylation (RdDM) pathway that uses 24nt siRNAs to recruit the de novo methyltransferase DRM2 to the target site. The REPETITIVE PETUNIA SEQUENCE (RPS) acts as a hot spot for de novo methylation, in particular, hypermethylation is found at a 62 nt region containing an 11 nt palindromic sequence with an 18 nt spacer that forms a putative stem loop structure. The analysis of deletion/substitution derivatives of RPS showed that de novo methylation did not depend on the DNA sequences in the loop region, but did require the 11nt stem region. In addition, a 10nt region directly downstream of the loop region contributed to the efficiency of de novo methylation. Where hypermethylation occurs, it is associated with histone 3 lysine 9 dimethylation. A mutant construct where the palindromic region had been interrupted, produced two distinct types of either almost fully hypomethylated or fully hypermethylated transgenes. This suggests that the RPS stem loop provides a structural de novo methylation target. Analysis of the RPS derivatives in mutant backgrounds identified three distinct stages of DNA methylation which included an RdDM-independent initiation step, DCL-dependent but RDR2-independent spreading. and RDR2-dependent amplification. The initial step therefore represents a potentially novel mechanism for de novo DNA methylation.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available