Supplementary
Functional Characterization of GPC-1Genes in Hexaploid Wheat
Raz Avni1*, Rongrong Zhao2,3*, Stephen Pearce3, Yan Jun4, Cristobal Uauy3, Facundo Tabbita3, Tzion Fahima4, Ann Slade5, Jorge Dubcovsky3, Assaf Distelfeld1
1 Faculty of Life Sciences, Dept. of Molecular Biology and Ecology of Plants, Tel Aviv University, 69978, Israel.
2 Department of Plant Nutrition, College of Resources and Environmental Science, China Agricultural University, Beijing, 100193, P.R. China
3 Dept. of Plant Sciences, University of California, Davis, CA, 95616, USA.
4 Dept. of Evolutionary and Environmental Biology, University of Haifa, Mt. Carmel, Haifa, 31905, Israel.
5 Arcadia Biosciences Inc., Seattle, WA 98119, USA
*These authors contributed equally to this work
Fig. S1 Flag leaf photosynthesis rate (Amax, CO2 m-2s-1) of GPC-1 mutants and control at 3 days before anthesis and at 22, 45 and 59 DAA from the TAU 2011 experiment. Data is the mean value (n=5±SEM). Asterisks indicate significance compared to control using Tukey-HSD test for multiple comparisons. ***P<0.001
Fig. S2 Differences in spike water content (g) between GPC-1 mutants and control (measured as the difference between spike weight at the time of harvest and after drying) at 42, 49, 57 and 63 DAA. Spikes were harvested from the TAU 2011 experiment. Data is mean value (n=5±SE). Asterisks indicate significance compared to control using Tukey-HSD test for multiple comparisons. *P<0.05, **P<0.01, ***P<0.001
Fig. S3 Grain Fe and Zn levels (ppm) in GPC-1 mutants and control plants. Mature grain samples were harvested from the TAU 2012 and UCD 2012 experiments and measured using ICP-MS. Data is mean value (n=5 ±SE). Asterisks indicate significance compared to control using Tukey-HSD test for multiple comparisons. *P<0.05, **P<0.01, ***P<0.001
Fig. S4 Thousand kernel weight (TKW) values for grains harvested from experiments in 2011 (left) and 2012 (right). Measurements revealed no significant difference between mutants and control in any environment, data is mean value (n=5±SE)
Fig. S5 Dry spike weight of GPC-1 mutants and controls. Spikes were harvested from the NY 2012 experiment. The large spike weight is a result of high fertility of these lines resulting in ~4 grains per spikelet. Figure shows similar weight gain of the spikes for all the genotypes. Data is mean value (n=5±SE)
Table S1 Relative leaf chlorophyll content by year, field, DAA and genotype. Table shows the mean value and letters indicate significance group by Tukey-HSD test
Year / Field / DAA / ∆GPC-1 / ∆GPC-A1 / ∆GPC-D1 / Control2011 / TAU / 17 / 39.9AB / 39.3B / 38.0B / 42.8A
32 / 33.7A / 33.3A / 31.3A / 29.6A
38 / 36.8A / 25.4B / 28.9B / 13.8C
43 / 33.4A / 22.3B / 19.9B / 7.6C
53 / 30.2A / 11.0B / 6.5C / 1.7D
NY / 15 / 34.9A / 33.4A / 34.7A / 34.5A
34 / 35.6A / 27.5B / 30.0B / 12.0C
40 / 26.4A / 16.7B / 15.5B / 5.4C
2012 / TAU / 0 / 44.4A / 44.6A / 44.6A / 45.1A
8 / 44.3AB / 43.5B / 44.1AB / 45.0A
17 / 42.8AB / 43.2AB / 42.3B / 44.2A
22 / 41.3A / 42.2A / 40.1A / 42.4A
31 / 37.6A / 38.2A / 34.6AB / 32.7B
38 / 33.9A / 30.4A / 19.7B / 10.5C
41 / 27.3A / 18.7B / 8.5C / 2.4D
46 / 21.6A / 5.5B / 2.4BC / 0.1C
52 / 11.8A / 0.2B / 0.1B / 0.1B
59 / 3.4A / 0.1B / 0.1B / 0.1B
63 / 0.7A / 0.1A / 0.1A / 0.1A
NY / 10 / 46.0A / 44.2B / 45.5AB / 45.1AB
25 / 39.3A / 34.3B / 36.4AB / 35.8B
37 / 31.0A / 15.6B / 14.8B / 1.3C
UCD / H / 44.0B / 44.8AB / 46.5A / 45.0A
0 / 44.8A / 44.4A / 46.1A / 45.4A
7 / 44.8B / 45.4AB / 47.0A / 46.1A
12 / 45.1A / 45.3A / 46.7A / 45.7A
22 / 46.8A / 46.0A / 46.7A / 46.0A
29 / 45.4A / 43.7A / 44.4A / 44.2A
32 / 45.9A / 43.5A / 44.1A / 36.3B
35 / 43.0A / 38.1A / 37.4A / 9.9B
38 / 38.2A / 30.3A / 32.5A / 8.1B
41 / 36.5A / 25.6B / 22.4B / 7.5C
44 / 34.3A / 6.7B / 7.7B / 7.7B
47 / 34.1A / 7.8B / 7.0B / 6.8B
50 / 33.8A / 6.0B / 6.0B / 6.4B
53 / 29.9A / 5.8B / 6.9B / 6.2B
56 / 26.0A / 5.9B / 6.2B / 5.6B
59 / 16.6A / 4.9B / 5.7B / 5.7B
62 / 11.0A / 5.9B / 5.6B / 5.9B
65 / 8.0A / 5.7B / 5.4B / 6.1AB
68 / 7.0A / 4.6B / 5.3B / 5.1B
71 / 6.0A / 4.5BC / 4.4C / 5.3AB
Table S2 Grain protein content (GPC) by year and location
Year / Location / Control / gpc-a1 / ∆(%) / gpc-d1 / ∆(%) / gpc-1 / ∆(%)2011 / TAU / 142.1 ± 1.1 / 137.5 ± 1.3 / -3.5 / 127.7 ± 1.0 / -9.9*** / 115.1 ± 1.3 / -19.0***
KM / 145.8 ± 1.1 / 139.6 ± 1.1 / -4.1 / 135.2 ± 1.2 / -7.5*** / 121.0 ± 1.9 / -17.1***
NY / 105.5 ± 1.3 / 104.2 ± 1.2 / -1.0 / 101.2 ± 3.1 / -3.8 / 94.1 ± 2.0 / -10.5*
2012 / TAU / 142.5 ± 1.4 / 132.5 ± 0.8 / -7.0*** / 128.7 ± 0.8 / -9.2*** / 111.8 ± 2.0 / -21.1***
NY / 128.6 ± 2.4 / 122.8 ± 3.2 / -4.6 / 122.4 ± 2.0 / -5.4 / 111.6 ± 1.5 / -13.2*
UCD / 122.2 ± 1.0 / 118.0 ± 1.0 / -3.3* / 115.6 ± 1.0 / -4.9*** / 97.1 ± 0.9 / -20.5***
Values are given as Mean ± SE for GPC (g Kg-1) and ∆, mean change between each mutant and the average of the two control lines (as a percentage of the control). Asterisks indicate significance compared to control using Tukey-HSD test for multiple comparisons. *P<0.05 and ***P<0.001
Table S3 Yield parameters in TAU and NY 2012
Genotype / Location / Plant biomass (g) / Spike number per plant / Grain weight per plant (g) / Harvest index / Grain weightperspike (g)gpc-a1 / TAU / 69.8± 4.7 / 10.6± 0.7 / 28.5± 2.0 / 40.7± 0.9 / 2.7± 0.1
gpc-d1 / TAU / 70.4± 5.9 / 11.0±0.9 / 30.7± 2.7 / 43.2± 0.5 / 2.8± 0.1
gpc-1 / TAU / 62.1± 5.0 / 10.2± 0.7 / 22.3± 1.9* / 35.7± 1.0*** / 2.2±0.1**
Control / TAU / 75.9± 4.9 / 11.9± 0.6 / 32.3± 2.2 / 42.2± 0.7 / 2.6± 0.1
gpc-a1 / NY / 60.6± 4.7 / 8.0 ± 0.5 / 25.8± 2.1 / 42.6± 1.7* / 3.2± 0.2
gpc-d1 / NY / 67.5± 5.4 / 9.3± 0.8 / 27.6± 2.4 / 40.6± 0.6 / 3.0± 0.1
gpc-1 / NY / 70.0± 4.1 / 9.4± 0.6 / 27.0± 2.1 / 38.1± 1.4 / 2.8± 0.1
Control / NY / 68.1± 3.4 / 9.0± 0.4 / 26.5± 1.6 / 37.7± 1.0 / 2.9± 0.1
Values are mean value ± SE (n=5). Asterisks indicate significance compared to control using Tukey-HSD test for multiple comparisons. *P<0.05, ** P<0.01 and ***P<0.001