Supplementary Tables

Primer name / Primer sequence
SMG-7L-VIGS-for-EcoRI / CATGAATTCGGCTGCTCTAGACTCCTACCAGTT
SMG-7L-VIGS-rev-EcoRI / CATGAATTCCTCCTGACTTGTCCTTGTCCGCTA
Vv-SMG-7-1-start-for-SmaI / CATCCCGGGTGGCTGATGATTGTACAAATGGAT
Vv-SMG-7-1-stop-rev-XbaI / CATTCTAGACACGAAATAACGACCTGGCCA
Vv-SMG-7-2-start-for-SmaI / CATCCCGGGATGCGAGAGAACTATGAAGCGAT
Vv-SMG-7-2-stop-rev-SpeI / CATACTAGTCACAAAGAATTGGCCTCCCCAT
Vv-SMG-7L-start-for-SmaI / CATCCCGGGATGGGTACTGATTCACTCATCCCCCT
Vv-SMG-7L-stop-rev-XbaI / CATTCTAGATCAATTCCCCATGTAAGTAGGACCTCT
Vv-SMG-7-1-C-468-for-SpeI / CATACTAGTGTGGGAGGATTTTGAGCTGAGAGGAT
Vv-SMG-7-1-C-stop-rev-SalI / CATGTCGACTCACACGAAATAACGACCTGGCCA
Vv-SMG-7L-C-517-start-for-SpeI / CATACTAGTGTGGGAAGACTATGAGCTGCGTGGCTT
Vv-SMG-7L-C-stop-rev-SalI / CATGTCGACTCAATTCCCCATGTAAGTAGGACCTCT
SMG-7-1-T-for-BamHI / CATGGATTCGATCATTTTGAGAGCACAGATGGTA
SMG-7-1-T-rev-EcoRI / CATGAATTCCCCACTATGAATGCATTAGTGTGTA
SMG-7-2-T-for-BamHI / CATGGATTCGGTCATTACATGTGTAGATGGTA
SMG-7-2-T-rev-SmaI / CATCCCGGGGCGTCTTAAGCCGAGCTTTAA
SMG-7L-T-for-XbaI / CATTCTAGATATGAGAAAGGTTCGGTTATCAA
SMG-7L-T-rev-EcoRI / CATGAATTCGGGTTTGATTGCCCAATCCAT
Vv-SMG-7L-N-rev / CAGCTCAAAATCCTCCCACAGAGCA
Vv-SMG-7L-C-for / TGTGGGAGGATTTTGAGCTGAGA
Nb-SMG7L-qPCR-for / ATCTGCCGCTTTCCCTGCCG
Nb-SMG7L-qPCR-rev / AAATCCCCTGTCCCCCTCTCGTT
Tomato-ubi3-qPCR-for / GCCGACTACAACATCCAGAAGG
Tomato-ubi3-qPCR-rev / TGCAACACAGCGAGCTTAACC

Table S1. List and sequences of the primers used.

SMG7L - ubiquitin ∆Ct / SMG7L level is decreased to
PDS-silenced / PDS-SMG7-silenced
leaf 1 / 2.3 / 4.0 / 34%
leaf 2 / -0.5 / 4.0 / 4,9%

Table S2. Quantitative PCR results show that the SMG7L mRNA is downregulated in PDS-SMG7L-silenced leaves compared to PDS-silenced leaves. The SMG7L qPCR Ct values were normalized to ubiquitin qPCR Ct values of the same samples. ∆Ct values were the same for the two different SMG7L-PDS VIGS samples, whilst they were different for the two PDS VIGS samples. Comparison of the SMG7L-PDS to the PDS ∆Ct values shows that SMG7L was significantly downregulated in SMG7L-PDS-silenced leaves.


A.

sequences compared / identity (%)
1 / At S7 and S7L N / 27
2 / Vv S71 and S72 N / 76
3 / Vv S71 and S7L N / 29
4 / Vv S72 and S7L N / 29
5 / Pt S71 and S72 N / 64
6 / Pt S7L1 and S7L2 N / 83
7 / Pt S71 and S7L1 N / 30
8 / Pt S71 and S7L2 N / 30
9 / Pt S72 and S7L1 N / 29
10 / Pt S72 and S7L2 N / 29
11 / Vv S71 and Pt S71 N / 80
12 / Vv S72 and Pt S72 N / 69

B.

sequences compared / identity (%)
1 / At S7 and S7L C / 15
2 / Vv S71 and S72 C / 50
3 / Vv S71 and S7L C / 16
4 / Vv S72 and S7L C / 14
5 / Pt S71 and S72 C / 38
6 / Pt S7L1 and S7L2 C / 83
7 / Pt S71 and S7L1 C / 15
8 / Pt S71 and S7L2 C / 15
9 / Pt S72 and S7L1 C / 14
10 / Pt S72 and S7L2 C / 14
11 / Vv S71 and Pt S71 C / 60
12 / Vv S72 and Pt S72 C / 46

Table S3. Comparison of N- and C-terminal domains of plant SMG homologs. The full-length protein sequences of the A. thaliana (At), V. vinifera (Vv) and P. trichocarpa (Pt) SMG homologs were aligned, then the alignment was separated into the N- and C-terminal domains and the percent of identical amino acids was determined between the different sequence pairs using the MEGA program. a Conservedness of the N-terminal domain of SMG7 proteins. Comparisons between two SMG7 (S7) homologs are highlighted in bold, whilst comparisons between two SMG7L (S7L) homologs are highlighted in bold italic. Identity of the protein sequence is between 64 and 80% in S7-S7 comparisons and it is 83% in the Pt S7L-S7L comparison. Identity is much lower in S7-S7L comparisons, were it is between 27 and 30%, which is around the border of the twilight zone. b Conservedness of the C-terminal domain of SMG proteins. Note that conservedness is considerably lower in the C-terminal region of S7 proteins (38-60% compared to 64-80%) and in S7-S7L comparisons (14-16% compared to 27-30%) than in the N-terminal region. The conservedness of S7 and S7L pairs is far below the twilight zone which makes the reliable alignment of this region impossible, particularly with traditional alignment methods. Note that the identity of both N- and C-terminal domains between Vv S71 and Pt S71 (A,B line 11) is higher than between Vv S71 and Vv S72 (A,B line 2) or between Pt S71 and Pt S72 (A,B line 5).

SMG7-1 / SMG7-2 / SMG7L
CB970809 / CB969879* / CD800872*
CB970887* / CF404865* / DT030409
DV940959* / CF607511 / DT031238
DY474202* / CF609286 / DT033336
EC940073 / EC986861 / EV242749*
EC958861 / EE081427 / FC057581
EE076456
EE093348

Table S4. EST sequences used to in silico assemble the 3’ UTRs of the three grapevine SMG7 homologs (SMG7-1, SMG7-2 and SMG7L). ESTs having polyA sequence at their 3’ ends are indicated by asterisk. Note that although full-length cDNAs were not available, it was possible to predict full length cDNA sequences for all the three genes by the alignment of overlapping ESTs.