Table 2. Effect of differences in the spacing between the two palindromes of the as-1 element on binding of ASF-1, TGA2.2, TGA2.1 and TGA1a and on promoter activity.

	ASF-1	TGA2.2	TGA2.1	TGA1a	GUS
as-1	1.00 (± 0.00)	1.00 (± 0.00)	1.00 (± 0.00)	1.00 (± 0.00)	100.0 (± 0.0)
as-1/(–2)GA	2.97 (± 0.12)	1.09 (± 0.14)	0.96 (± 0.32)	1.77 (± 0.14)	16.9 (± 1.2)
as-1/(–2)GG	0.61 (± 0.03)	0.43 (± 0.08)	0.23 (± 0.03)	n.d.	9.5 (± 0.5)
as-1/(+2)	0.16 (± 0.02)	0.17 (± 0.01)	0.08 (± 0.04)	0.29 (± 0.03)	18.4 (± 4.4)
as-1/(+4)	0.27 (± 0.01)	0.23 (± 0.02)	0.09 (± 0.03)	0.46 (± 0.02)	13.5 (± 2.8)
as-1/(+6)	0.23 (± 0.01)	0.16 (± 0.06)	0.09 (± 0.03)	0.44 (± 0.05)	8.7 (± 2.8)
as-1/(+8)	0.37 (± 0.02)	0.33 (± 0.08)	0.21 (± 0.04)	0.62 (± 0.13)	13.7 (± 1.2)
as-1/(+10)	0.32 (± 0.06)	0.21 (± 0.02)	0.12 (± 0.04)	0.61 (± 0.10)	11.8 (± 1.2)

To calculate the relative affinities of the different protein factors to the different as-1 elements, the percentage of radioactivity left in the presence of a 200-fold molar excess of the wild-type as-1 element was set 1 (see Materials and Methods). Relative GUS activities mediated by these elements in transiently transformed protoplasts of tobacco cell line BY-2 are given as percent of the activity of the wild-type as-1 element. Values are taken form three independent experiments.