Moonvista™ carnation

Moonvista™ is a carnation variety in which the flowers have a violet/mauve colour due to the insertion of Viola sp. flavonoid 3', 5'-hydroxylase gene and Petunia hybrida dihydroflavonol-4-reductase to enable the biosynthesis of delphinidin (anthocyanin) pigment. The flowers also carry a variant form of acetolactate synthase from Nicotiana tabacum for selection sulfonylurea selection during transformation.

NOTE: This LMO was formerly referred to with the UID FLO-4Ø685-1.

pCGP1991

Gene expression
Three gene cassettes are present: Nicotiania tabacum acetolactate synthase (ALS; acetohydroxy acid synthase), Petunia hybrida dihydroflavonol-4-reductase (DFR) and Viola sp. flavonoid3', 5'-hydroxylase (F3'5'H).

Transcription of ALS is under control of a Cauliflower mosaic virus (CaMV) 35S promoter and a N. tabacum ALS terminator. A 5' untranslated leader sequence from P. hybrida chlorophyll a/b-binding protein is also present at the 5' end of ALS, but is not expected to be translated. The leader sequence promotes high levels of transcription of ALS.

Transcription of DFR is under control of its endogenous promoter and terminator. The coding sequence contains 6 exons and 5 introns.

Transcription of F3'5'H is under control of an Antirrhinum majus chalcone synthase promoter and a P. hybrida D8 terminator.

Note:
- The size of the ALS coding sequence includes the size of the terminator (3.76 kb = size of ALS coding sequence + ALS terminator)
- The size of the DFR coding sequence represents the size of the full genomic cone (4.96 kb = DFR promoter + DFR coding sequence + DFR terminator)
- The T-DNA is present at one integration locus and contains one copy of each T-DNA component as determined by Southern blot analysis

There are up to five copies of each integrated component of the T-DNA, and integration of T-DNA has occurred at four loci. Estimated copy numbers of T-DNA components integrated in transgenic line FLO-4Ø685-1 as follows: LB - 2 copies, NtALS (SurB) - 2 copies, VhF3'5'H - 4 copies, PhDFR - 2 copies, RB - 5 copies.

These carnations were developed using recombinant DNA techniques to produce flowers with a unique violet/mauve colour by introducing two genes from petunia (Petunia hybrida) that function together in the biosynthesis of the anthocyanin pigment delphinidin. The transgenic lines were derived from the parent cultivar 'White Unesco', which is a white coloured carnation that was selected for a mutation in the dihydroflavonol reductase (DFR) encoding gene that did not allow for expression of a functional enzyme, and thus did not produce the anthocyanin type pigments that give rise to blue and red coloured flowers. The two genes from Petunia hybrida introduced into the transgenic carnation lines included a functional dihydroflavonol reductase encoding gene (dfr) and a gene (hf1) encoding the enzyme flavonoid 3', 5'-hydroxylase (F3'5'H), a member of the NADPH-Cytochrome P450 reductase family. Expression of the F3'5'H encoding gene allows for the production of blue coloured delphinidin anthocyanin pigments, which are not normally found in carnations.

Tolerance to sulfonyl urea herbicides was produced via the introduction of a chlorsulfuron tolerant version of the acetolactate synthase (ALS) encoding gene from tobacco (SuRB).