Supporting data

Supplementary Table 1 Primer sequences used for RT-PCR and qRT-PCR in this study

Primer Name Accession No. Primer Sequence

IbMYB1a-For AB258984 5’-AGCTAAGAATTTCCGACACCCTTCAATA-3’

IbMYB1a-Rev AB258984 5’-GTGAATTTAACGCTTAGCTTAACAGTTCT-3’

RpSTS-NdeI-For JX673939 5’-CATATGGCACCGGAGGAGT-3’

RpSTS-SpeI-Rev JX673939 5’-ACTAGTTCAGGTAATTAGCGGC-3’

VrROMT-NdeI-For JX673941 5’-CATATG GATTTGGCAAACG-3’

VrROMT-SpeI-Rev JX673941 5’-ACTAGTTCAAGGATAAACCTCAA-3’

NtPAL4-For EU883670 5’-acacaaccaagatgtcaactcct-3’

5’-GTTCGCCTACGCTGATGAT-3’ (for qRT-PCR)

NtPAL4-Rev EU883670 5’-agttctgttccaagctcctttct-3’

5’-TCACATTCTTCTCACTTTCACCATT-3’ (for qRT-PCR)

NtC4H-For DQ350352 5’-tatggtcaattgagtggggtatc-3’

5’-CAATTCCTCTATTAGTCCCACACAT-3’ (for qRT-PCR)

NtC4H-Rev DQ350352 5’-tatggagactgaactgtccacct-3’

5’-ACTCTTCGGGTTTCTTCCAATG-3’ (for qRT-PCR)

NtACTIN-For AB158612 5’-gctacttactgaagcacccttga-3’

5’-TCTGGATTGGAGGATCTATCTTAGC-3’ (for qRT-PCR)

NtACTIN-Rev AB158612 5’-gcagtaatttccttgctcatacg-3’

5’-ATTTGCGGTGGACAATGGAA-3’ (for qRT-PCR)

RT-PCR primers were designed using Primer3 software (Rozen and Skaletsky 2000).

qRT-PCR primers were designed using the Beacon designer™ software (Premier Biosoft)

Supplementary Table 2 Quantification of resveratrol derivatives produced in cross-pollinated transgenic tobacco lines (F1) of SM and RSM

Constructs / Transgenic Lines / Western blot / trans-piceid (mg/g) / cis-piceid (mg/g) / Total piceid (mg/g)
Control (T3) / #1-1-1-1 / - / 0.00 / 0.00 / 0.00
IbMYB1a-OX (T1) / SPO-M #1 / - / 0.00 / 0.00 / 0.00
STS-OX (T3) / #4-2-5-1 / +++ / 5.62 / 16.06 / 21.67
STS x IbMYB1a (F1) / SM371-4 / ++ / 37.82 / 36.16 / 73.98
SM425-1 / ++ / 78.71 / 46.19 / 124.90
SM425-2 / +++ / 112.53 / 65.01 / 177.54
SM425-3 / +++ / 73.96 / 50.12 / 124.09
SM425-4 / +++ / 116.11 / 40.85 / 156.96
SM425-5 / +++ / 81.68 / 36.66 / 118.34
ROST x IbMYB1a (F1) / RSM282-2 / ++ / 34.80 / 29.75 / 64.55
RSM282-3 / ++ / 44.82 / 31.11 / 75.93
RSM282-4 / + / 47.78 / 6.07 / 53.85
RSM282-5 / ++ / 31.76 / 25.35 / 57.11
RSM4104-3 / +++ / 42.24 / 36.16 / 78.40
RSM4104-4 / + / 40.59 / 0.00 / 40.59
RSM4104-5 / +++ / 42.79 / 31.96 / 74.74

-, no expression of RpSTS transgene; +, low expression of RpSTS transgene; ++, moderate expression of RpSTS transgene; +++, high expression of RpSTS transgene

Supplementary Table 3 Quantification of resveratrol derivatives produced in cross-pollinated transgenic tobacco lines (F2) of SM and RSM

Constructs / Transgenic Lines / Western blot / trans-piceid (mg/g) / cis-piceid (mg/g) / Total piceid (mg/g)
Control (T3) / #1-1-1-1 / - / - / - / -
IbMYB1a-OX (T1) / SPO-M #1 / - / - / - / -
STS-OX (T3) / #4-2-5-1 / +++ / 5.62 / 16.06 / 21.67
STS x IbMYB1a (F2) / SM3714-2 / + / 55.80 / 67.57 / 123.37
SM4252-2 / ++ / 70.05 / 71.87 / 141.91
SM4252-3 / +++ / 101.76 / 137.80 / 239.56
SM4252-4 / ++ / 47.83 / 56.35 / 104.18
SM4252-5 / +++ / 95.68 / 93.47 / 189.15
ROST x IbMYB1a (F2) / RSM2823-1 / ++ / 30.19 / 29.12 / 59.31
RSM2823-4 / ++ / 26.52 / 36.33 / 62.86
RSM41045-1 / ++ / 25.49 / 37.57 / 63.06
RSM41045-3 / + / 18.24 / 23.51 / 41.75
RSM41045-4 / + / 21.16 / 16.87 / 38.03

-, no expression of RpSTS transgene; +, low expression of RpSTS transgene; ++, moderate expression of RpSTS transgene; +++, high expression of RpSTS transgene

Supplementary Table 4 HPLC-ESI-MS/MS characterization of resveratrol derivatives produced in cross-pollinated SM plants

Compound / RT / Structure / ESIMS(-) / Formular / MS/MS(+) / Formular / References
1 / 14.456 / trans-Piceid / 389.2 / C20H21O8 / 229.1 / C14H13O3 / Parage et al. (2012)
2 / 18.214 / cis-Piceid / 389.2 / C20H21O8 / 229.1 / C14H13O3 / Parage et al. (2012)
3 / 21.144 / Piceid methyl ether / 403.2 / C21H23O8 / 243.1 / C15H15O3 / Parage et al. (2012)
4 / 23.903 / Resveratrol methyl ether-O-hexoside / 445.2 / C23H25O9 / 243.1 / C15H15O3 / Parage et al. (2012)
5 / 25.346 / Resveratrol methyl ether-O-hexoside / 445.2 / C23H25O9 / 243.1 / C15H15O3 / Parage et al. (2012)
6 / 18.441 / 5-methylresveratrol-3, 4′-O-β-D-diglucopyranoside / 565.2 / C27H34O13 / 403.1 / C21H23O8 / Dai et al. (2009)

RT, retention time in min; ESIMS(-), compound mass in negative ion mode electrospray ionization mass spectrometry ([M-H]-); MS/MS(+), fragment mass in positive ion mode.