Chrysanthemum is a vegetative propagated culture in which viral transmission with planting material is important for its production. Chrysanthemum virus B (CVB) belongs to the viruses that strike this plant culture. Chrysanthemum virus B is found everywhere where chrysanthemum is cultivated. Damage to plants by CVB often leads to a complete loss of floral yield. Chrysanthemum (Chrysanthemum morifolium Ramat cv. White Snowdon) was transformed via Agrobacterium-mediated DNA delivery with the aim of improving resistance to CVB infection. Transformation vectors contain the nucleotide sequence of CVB coat proteins (CP) in sense, antisense, and double sense orientation. The transformative vectors also invert repeats of CVB coat protein gene fragments for the induction of RNA-interference. The transgenic chrysanthemum plants were successfully obtained. The integration of the target sequences in plant genomes was confirmed by polymerase chain reaction (PCR) and Southern blot analyses. Chrysanthemum lines were transformed with antisense, sense, and double sense CVB CP sequences, as well as with hairpin RNA-interference constructs that were assayed for resistance to CVB. Infection of transgenic plants by CVB through the grafting of infected scions shows resistance only among plants with carried double sense (16.7%) and hairpin (12.5%) constructs. The plants transformed by sense and double sense sequences were observed and classified as tolerant.

We report a systematic comparison of 19 plant promoters and 20 promoter-terminator combinations in two expression systems: agroinfiltration in Nicotiana benthamiana leaves, and Nicotiana tabacum BY-2 plant cell packs. The set of promoters tested comprised those not present in previously published work, including several computationally predicted synthetic promoters validated here for the first time. The expression of EGFP driven by different promoters varied by more than two orders of magnitude and was largely consistent between two tested Nicotiana systems. We confirmed previous reports of significant modulation of expression by terminators, as well as synergistic effects of promoters and terminators. Additionally, we observed non-linear effects of gene dosage on expression level. The dataset presented here can inform the design of genetic constructs for plant engineering and transient expression assays.

Chrysanthemum transgenic plants with integrated single and double copies of the gene encoding for the virus B coat protein have been created via Agrobacterium-mediated transformation. Both mRNA and the gene product have been revealed in several transformed lines only. The use of the transgenic chrysanthemum plants in the development of varieties resistant to the B virus is discussed.