Supplemental Figure 1

Transcription

1 Embryo Silk days after pollination hours after pollination Endosperm days after pollination Division expansion Leaf Transition Whole leaf mature Growth activity Growth activity Supplemental Figure 1. CNR1 expression patterns in different tissues. MPSS transcript expression of CNR1 in various tissue series organized left to right by decreasing growth activity or increasing maturation. The tissue samples are the same as those for CNR2 expression of Figure 5 A. Y-axis is expression level in PPM (parts per million). 1

2 Leaf area (cm2) (Ear leaf) Leaf position on the plants TG Supplemental Figure 2. Leaf area of transgenic plants over-expressing CNR1 (Pro ZmUBI :CNR1) and non-transgenic controls. Leaf area was measured in transgenic plants over-expressing CNR1 and non-transgenic sibling controls of event 5 using the leaf meter (LI-COR LI-31, LI-COR). Plants were grown in greenhouse to maturity. Ear leaf (leaf no. ), two leaves below (leaf no. -1 and -2, respectively) and two leaves above the ear leaf (leaf no. +1 and +2, respectively) were removed from the plants and measured. Data were collected from five transgenic and non transgenic control plants, respectively. Bars in the graph show the standard error. 2

3 Event May, 27 July, Average internode length (cm) Average internode length (cm) Event 7 Average internode length (cm) May, 27 October, Average internode length (cm)

4 Supplemental Figure 3. Internode length of transgenic plants over-expressing CNR1 (Pro ZmUBI :CNR1) and non-transgenic controls. Internode length was measured by the distance between nodes, with the brace roots considered the first node, and the base of the tassel the final node. Data represent the average of 5 plants. X-axis, number of internode from base, y-axis, length of the internode. Upper panel, Event 5, a growth-suppressing event; Lower panel, Event 7, a growth-enhancing event. For each event, data was collected from two different times of the year, May and July 27, for Event 5 and May and October 27, respectively. Plants were grown in the greenhouse until maturity. Bars in the graph show the standard error. 4

5 4copies 1 copy 2 copies Supplemental Figure 4. plants with different copy numbers of the transgene for over-expressing CNR1 (Pro ZmUBI :CNR1). Each plant is an independent T generation transgenic event. All plants shown were at the same age. All of the pots were the same size and the photos are in the same scale. 5

6 A B Cell counts per area Event 7 Event 5 Supplemental Figure 5. Leaf epidermal cell number counts. (A). Example image of the leaf epidermis from a control plant. (B). Cell number counts per fixed unit area, a measure of relative cell size, in transgenic plants over-expressing CNR1 and non-transgenic sibling controls. Data were collected from 5 transgenic and non transgenic control plants, respectively, per event. Event 5 is a growth-suppressing event; event 7 is a growth-enhancing event. Bars in the graph show the standard error. 6

7 Supplemental Figure 6 A linear range for ethidium bromide quantification X-axis, DNA amount loaded on the gel in ul of a concentration of 1 ng/ul. Y-axis, arbitrary unit of DNA quantification based upon Quantity One Image Quant software (Bio-Rad). The standard error bar was based upon 3 replicates. The gel image is an example of the 3 gel replicates, showing various DNA amount loaded. The lowest and highest Image Quant reading for CNR2/tubulin quantification in the experiments (1,891 and 3,645, respectively) before normalization (Figure 6), were within the linear range shown here. 7

8 Supplemental Table 1. RT-PCR and Cloning Primers PCR Primer/Probe CNR2_forward CNR2_reverse α tubulin_forward α tubulin_ reverse CNR1_ forward CNR1_ reverse CNR1_probe β-actin_ forward β-actin_ reverse β-actin_ probe CNR1 over-expression vector_forward CNR1 over-expression vector_reverse CNR1 RNAi vector_forward CNR1 RNAi vector_reverse Sequence (5 to 3 ) CGGCGGCGGCGGCTACTACCAG CGGCAGAGCGTCAGCGGGTCATCC AGCCCGATGGCACCATGCCCAGTGATACCT AACACCAAGAATCCCTGCAGCCCAGTGC GACATCCTGGTGCACTGCT CTTGAGCTCGCGGTACTCTT TGGCTCTG CTTCGAATGCCCAGCAATGT GTTCGCCCACTAGCGTACAAC TCGAGGCTGTTCTTT GATCCTAGTTCTACTCCCAAGCAATGT GTTCTGCCACCGCATCAATCAATCATC GGATGAAGAGCTGTGCTGAGAGAATCA GCAGCAGCAGTGCACCAGGATGTCCGG 8