Colour break in reverse bicolour daffodils is associated with the presence of Narcissus mosaic virus

Table 3 Sequence identification of the viruses present in tepals of Narcissus flowers displaying colour break

Cultivar tested	Primer name	Genus expected	Fragment size cloned/expected (kb)	Sequence results	Genus obtained
Twelve Gauge	Potex1RC (For) Potex5(Rev)	Potexvirus	0.74/0.74¹	NMV (1, 97%)	Potexvirus
	Sprimer (For) KS(Rev)	Potyvirus	1.65/1.60²	NLSYV (1, 92%) NMV (6, 97%)	Potyvirus Potexvirus
	pCar-1 (For) KS(Rev)	Carlavirus	0.80/1.80²	NMV (2, 98%)	Potexvirus Text
Daydream	Potex1RC (For) Potex5(Rev)	Potexvirus	0.74/0.74¹	NMV (2, 97%)	Potexvirus
	pCar-1 (For) KS(Rev)	Carlavirus	0.80/1.80² 0.30/1.80²	NMV (2, 98%) NMV (2, 98%)	Potexvirus
Twilight Zone	Potex1RC (For) Potex5(Rev)	Potexvirus	0.74/0.74¹	NMV (1, 97%)	Potexvirus
	Sprimer (For) KS(Rev)	Potyvirus	1.65/1.60²	NMV (2, 97%) NLV (4, 98%)	Potexvirus
	pCar-1 (For) KS(Rev)	Carlavirus	0.80/1.80² 0.80/1.80² 0.30/1.80²	NSV (1, 95%) NMV (1, 98%) NMV (2, 98%)	Carlavirus Potexvirus
Flying Cloud	Sprimer (For) KS(Rev)	Potyvirus	1.65/1.60²	NMV (4, 97%)	Potexvirus

NMV = Narcissus mosaic virus (AY225449, D13747); NLSYV = Narcissus late season yellows virus (AJ493579); NSV = Narcissus symptomless virus (AM182569); NLV = Narcissus latent virus (FJ024083). RT-PCR was performed on tepal RNA from four flowers displaying colour break. The predominant amplification products indicated in Figure 4 were gel purified, cloned and sent for sequencing. Numbers in brackets refers to number of clones sequenced and average percentage nucleotide identity to virus accession in GenBank. There are two NMV accessions in the GenBank (New Zealand strain, accession AY225449 and Dutch strain, D13747). Sequences showed equal % similarity to both. Expected size (kb) of PCR product as reported by ¹van der Vlugt & Berendsen [11], and ²Chen et al. [12].

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