TABLE 1.

Primers used for the amplification and assembly of the infectious clone and protein expression constructs^a

Construct and gene/region	Sense	Sequence (5′ to 3′)	Binding site (KC517488)
CTV GFP-p27 fusionb
Outer primers	+	GCAATCTCGAGACTAGTTAGTGCTGTCTCTCCCGTATATC	11661–11688
	−	CGTGTCTAAGTCACGCTAAACAAAGTGAC	17272–17300
P27 to GFP read-through	+	TACGCGATTTGGGTAAGTACCTATAGCAATTACAGATGGCTAGCAAAGGAGAAGAACT	16025–16044
	−	AGTTCTTCTCCTTTGCTAGCCATCTGTAATTGCTATAGGTACTTACCCAAATCGCGTA	16025–16044
GFP CP controller element	+	TACAAATAACTCGAGGGGTAGTTGGTTTGGGGACGGTAACATTATAC	16051–16076
	−	GTATAATGTTACCGTCCCCAAACCAACTACCCCTCGAGTTATTTGTA	16051–16076
E. coli/pRSETc
L1	+	ATGCGGATCCATGAGTGGGCGGCGAGTTTGTTAC	1122–1142
	−	CGATCTCGAGCTAACCAACCAAATGGTGGTTAGG	1542–1562
L2	+	ATGCGGATCCATGCGGTTTGTGGTGTGTGTTGAAG	2580–2601
	−	CGATCTCGAGGCCCATCTTATGATACTTATTGAG	2985–3008
p6	+	TCGATGGGGATCCATGGACTGTGTGATTCAAGG	11874–11893
	−	CGATAAGCTTTTAGATAGTGGCGTGAGTGC	12010–12029
p27	+	TCGATGGGGATCCATGGCGGGTTACACGATG	15325–15342
	−	CGATAAGCTTCTACAAATACTTTCCCAAATC	16027–16047
p25	+	TCGATGGGGATCCATGGACGACGAGACAAAGAA	16141–16160
	−	CGATAAGCTTTCAACGTGTGTTAAATTTCCCAAG	16789–16812
p18	+	TCGATGGGGATCCATGTCAGGCAGCTTGG	16778–16793
	−	CGATAAGCTTCTAAGTCGCGCTAAACAAAGC	17224–17244
p13	+	TCGATGGGGATCCATGGGTATTCGACGCGTG	17315–17332
	−	CGATAAGCTTCTAGTTATCGCAAGGTAAGA	17655–17674
p20	+	TCGATGGGGATCCATGCGAGCTTACTTTAGTG	17750–17768
	−	CGATAAGCTTCTACACGCATGAAGGAGAAAC	18278–18298
p23	+	TCGATGGGGATCCATGAACGATACTAGCGGAC	18380–18398
	−	CGATAAGCTTTCAGATGAAGTGGTGTTCACG	18989–19009
A. tumefaciens/pCAMBIA 1380d
35S promoter start	+	ATCGGGCGCGCCGATCTCCTTTGCCCCAGAGA	NAe
35S–6×His-p61 overlap	+	CATTTCATTTGGAGAGGATGCATCATCATCATCATCATTCGTCTCATCACGTATG	13746–13762
	−	CATACGTGATGAGACGAATGATGATGATGATGATGCATCCTCTCCAAATGAAATG	13746–13762
p61-Nos overlap	+	GGTTACACGATGCTTCCTAGCGTTCAAACATTTGGCAAT	15331–15346
	−	ATTGCCAAATGTTTGAACGCTAGGAAGCATCGTGTAACC	15331–15346
35S–6×His-p65 overlap	+	CATTTCATTTGGAGAGGATGCATCATCATCATCATCATGTGCTTCTGGGTCTAGAC	12038–12055
	−	GTCTAGACCCAGAAGCACATGATGATGATGATGATGCATCCTCTCCAAATGAAATG	12038–12055
p65-Nos overlap	+	TGGAAAGAGTACCTCTCTAGCGTTCAAACATTTGGCAAT	15300–15316
	−	ATTGCCAAATGTTTGAACGCTAGAGAGGTACTCTTTCCA	15300–15316
Nos terminator end	−	CGATCTGCAGCCCGATCTAGTAACATAGATG	NA

^aPrimer binding sites are given according to the sequence of isolate FS577 (NCBI GenBank accession no. KC517488) and restriction enzyme recognition sites are underlined, where appropriate.

^bAssembly and integration of a GFP read-through domain at the C terminus of the minor coat protein (CP) into the full-length T36 infectious clone.

^cCloning of CTV proteins into the pRSET binary vector for expression in E. coli BL21(DE3).

^dCloning of CTV proteins into the pCAMBIA 1380 vector for agrobacterium-mediated transient expression in N. benthamiana.

^eNA, not applicable.