Caffeine biosynthesis involves sequential methylation of the purine ring by the transfer of methyl group from the methyl donor S-adenosyl methionine. Genes that code for the individual methyl transferase have a high degree of sequence homology. To achieve decaffeination by genetic engineering of coffee, the effect and efficiency of different post-transcriptional gene silencing (PTGS) constructs were evaluated. Sense, antisense and invert repeat constructs were developed using a 339 bp fragment from the conserved region of the coffee N-methyltransferase genes involved in caffeine biosynthesis. The silencing constructs were mobilized into Agrobacterium tumefaciens (EHA101). Sonication assisted Agrobacterium-mediated transformation of the somatic embryos of C. canephora was performed, followed by regeneration of the transformants. Molecular analysis and identification of flanking regions confirmed the stable integration of the transgenes in the transformants and their integration in non coding regions of the genome, respectively. Effectiveness of the silencing constructs was evaluated by transcript analysis of the targeted genes by RT-PCR and northern blotting. The silencing efficiency was further evaluated by estimation of the purine alkaloid content in the transgenic lines using HPLC. The three constructs differed in their silencing efficiencies and specificity. Though the constructs were not specific to a single N-methyltransferase, transformants obtained were mainly affected in one of the N-methyltransferases. Results show that the use of homologous coding sequence in the PTGS constructs results in a much higher efficiency in silencing the caffeine biosynthetic pathway. Further, the lowering of caffeine content should be the preferred over complete decaffeination due to extremely low survival rate of transgenic plants with extremely low levels of caffeine.