Vernalization treatment induces site-specific DNA hypermethylation at the VERNALIZATION-A1 (VRN-A1) locus in hexaploid winter wheat

Table 1 Characteristics of PCR amplicons used for bisulfite analysis

Fragment number	Primer name	Primer sequence (5′-3′)	Position of fragment within theVRN-A1gene (bp from start of Genbank accession AY747600)	Size of amplified fragment (bp)
0.01 k⁽¹⁾	0.01 k-F	AAATGATTTGGGGAAAGTAAAATT	100–311	212
	0.01 k-R	AAAAAATTTTTAAAAAAATCTAACCC
1.2 k⁽¹⁾	1.2 k-F	ATTAAATTTGTGTTTGTTGTTTGA	1221–1468	248
	1.2 k-R	AACTCTCTACTTTTTAATTTAACTCTTC
2.2 k	2.2 k-F	GGGGATAAGTAATTGTTATGTTTTG	2259–2490	233
	2.2 k-R	ATCCAAAAAATCAACAAACTACAT
6.8 k(a) ^{(2) (3)}	6.8 k(a)-F	TTGTTTGTATGTGAGTAGATTGGA	6847–7147	301
	6.8 k(a)-R	AACTACTCCTCCACCTTATACCAA
7.6 k⁽⁴⁾	7.6 k-F	TGAGGAGGTTGGAAGTATTAAGTA	7679–7393	315
	7.6 k-R	TACACCCCTACAAAACTAAATCTT
9.2 k	9.2 k-F	ATGTAGTATGGATAAAATTTTTGAA	9203–9711	509
	9.2 k-R	TTATTACTAAACCCTTCAAAAACTC
10.1 k	10.1 k-F	GTTGTAGTTTTAATTGAGGGATT	10130–10512	383
	10.1 k-R	ACCCCTATCCATAACTAACTCT
10.5 k	10.5 k-F	AATTTGTTTGGGATTAAAGGTT	10592–10939	340
	10.5 k-R	CAAAATCCTCTCCCATAAAATAC
11.7 k	11.7 k-F	GTGTTYGTTTTGGTTGTGTAGT	11792–12048	257
	11.7 k-R	ACTCTAATTTCTTTTCCTTTCCC

Bold letters in the sequence represent primer positions matching cytosines, which were considered as non methylated and therefore converted into thymines after bisulfite treatment (Cs were changed to Ts in forward primers and Gs to As in reverse primers). ⁽¹⁾Primer pairs that were not genome A specific; ⁽²⁾Fragment for which several primer pairs with different C/T content were designed (see Additional file 1); ⁽³⁾While corresponding PCR amplicon contains a fragment of the Jorge transposon, the reverse primer does not overlap the annotation of this element. The closest Jorge element found shares only 76% nucleotide identity with this amplicon; ⁽⁴⁾While the two primers are included within the Sumaya annotation, the closest Sumaya element found shares only 81% nucleotide identity with this amplicon.

ISSN: 1471-2229