Table S1 1,8-cineole/-terpineol ratios of flowers of N. bonariensis, N. forgetiana and N. mutabilis during flower development and during the day (n=3-6)

Species / 1d / 2d / 3d / 4d / 5d
N. forgetiana / 2.7:1 / 2.0:1 / 2.7:1 / 2.5:1 / 2.5:1
N. bonariensis / 2.8:1 / 3.0:1 / 2.8:1 / 2.8:1 / 2.1:1
Species / 6:00 AM / 9:00 AM / 12:00 AM / 3:00 PM / 6:00 PM / 9:00 PM / 12:00 PM
N. forgetiana / 2.3:1 / 2.0:1 / 3.0:1 / 2.1:1 / 2.7:1 / 2.5:1 / 2.4:1
N. bonariensis / 2.2:1 / 2.3:1 / 2.2:1 / 2.4:1 / 4.0:1 / 2.5:1 / 2.4:1
N. mutabilis / 3:1 (n=1)

Table S2 Calculation of correlation coefficient (R) and significance value (p)

Florale emission (presented in Fig.2)

species / variable1/variable 2 / R correlation coefficient/ probability p
N. bonariensis / cineole/terpineol / 0.9= significant positive correlation, p=0.003
N. forgetiana / cineole/terpineol / 0.9, strong positive correlation, p= 0.006
N. mutabilis / cineole/terpineol / 0.9, strong positive correlation, p= 0.002

Petal raw extract (presented in Figs. 3, S3, S4)

species / variable1/variable 2 / R correlation coefficient/ probability p
N. forgetiana, GPP
6am to midnight / cineole/terpineol / 0.9, positive correlation ,p= 0.03
N. forgetiana, GPP
1d-5d / cineole/terpineol / 0.8, positive correlation, p=0.024
N. forgetiana, NPP
1d-5d / cineole/terpineol / 0.9, strong positive correlation, p=0.002
N. forgetiana, NPP
6am to midnight / cineole/terpineol / 0.9, strong positive correlation, p= 0.001
N. bonariensis, GPP
6 am to midnight / cineole/terpineol / 0.9, strong positive correlation, p= 0.0083
N. bonariensis, GPP
1d-5d / cineole/terpineol / 0.9, strong positive correlation, p=0.002
N. bonariensis, NPP
1d-5d / cineole/terpineol / 0.8, positive correlation, p=0.04
N. bonariensis, NPP
6 am to midnight / cineole/terpineol / 0.2, positive correlation, p= 0.05

Overexpressed enzymes (presented in Fig 7)

species / variable1/variable 2 / R correlation coefficient/ probability p
N. bonariensis / cineole/terpineol / 0.8= strong positive correlation, p= 0.002
N. forgeteiana / cineole/terpineol / 0.9=strong positive correlation, p= 0.002
N. longiflora / cineole/terpineol / 0.9 = strong correlation, p= 0.002

Table S3 Amino acid sequence comparison of Nicotiana CINs and TERs

N. alata / N. langsdorfii / N. bonariensis / N. forgetiana / N. longiflora / N. mutabilis / N. suaveolens / orf
N. alata / 100 % / 99 %
2 AS / 98 %
11 AS / 98 %
8 AS / 98 %
7 AS / 98 %
10 AS / 90 %
51 AS / 543 aa
N. langsdorfii / 100 % / 97 %
13 AS / 98 %
10 AS / 98 %
9 AS / 97 %
12 AS / 89 %
53 AS / 522 aa
N. bonariensis / 100 % / 98 %
11 AS / 98 %
10 AS / 97 %
13 AS / 90 %
54 AS / 524 aa
N. forgetiana / 100 % / 99 %
1 AS / 98 %
6 AS / 90 %
49 AS / 524 aa
N. longiflora / 100 % / 98 %
5 AS / 90 %
48 AS / 524 aa
N. mutabilis / 100 % / 90 %
50 AS / 524 aa
N. suaveolens / 100 % / 610 aa

Table S4 Amino acid alterations of newly isolated CINs versus TER of N. alata

A

N. alata TER / N. bonariensis CIN
1 / isoleucine 99 / valine
2 / proline 112 / arginine / nonpolar to basic
3 / aa 171 / arginine / basic
4 / aa 172 / asparagine / polar
5 / tyrosine 217 / glycine / polar to polar
6 / glycine 259 / alanine / stays nonpolar
7 / threonine 368 / arginine / polar to basic
8 / isoleucine 376 / arginine / nonpolar to basic
9 / methionine 393 / threonine / nonpolar to polar
10 / valine 496 / isoleucine / nonpolar
11 / serine 508 / threonine / polar

B

N. alata TER / N. forgetiana CIN
1 / aa 171 / arginine / basic
2 / aa 172 / asparagine / polar
3 / valine 191 / glutamic acid / nonpolar to acidic
4 / glutamine 246 / glutamic acid / polar to acidicr
5 / isoleucine 376 / arginine / nonpolar to basic
6 / valine 496 / isoleucine / nonpolar
7 / arginine 536 / serine / basic to polar
8 / leucine 541 / valine / polar

C

N. alata TER / N. longiflora CIN
1 / aa 171 / arginine / basic
2 / aa 172 / asparagine / polar
3 / valine 191 / glutamic acid / nonpolar to acidic
4 / glutamine 246 / glutamic acid / polar to acidic
5 / isoleucine 376 / arginine / nonpolar to basic
6 / valine 496 / isoleucine / nonpolar
7 / arginine 536 / serine / basic to polar

D

N. alata TER / N. mutabilis putative CIN
1 / histidine 129 / glutamine / basic to polar
2 / aa 171 / arginine / basic
3 / aa 172 / asparagines / polar
4 / valine 191 / glutamic acid / nonpolar to acidic
5 / isoleucine 226 / phenylalanin / nonpolar
6 / isoleucine 245 / threonine / nonpolar to polar
7 / glutamine 246 / glutamic acid / polar to acidic
8 / isoleucine 376 / arginine / nonpolar to basic
9 / asparagine 495 / aspartic acid / polar to acidic
10 / valine 496 / isoleucine / nonpolar

aa: amino acid