DOI: 10.5061/dryad.h9w0vt4vf
title: Raw data for: Dual-cycle CO2 fixation enhances growth and lipid synthesis in Arabidopsis thaliana

description: Carbon fixation through the Calvin-Benson-Bassham (CBB) cycle accounts for
 the majority of CO2 uptake from the atmosphere. The CBB cycle generates C3
 carbohydrates but is inefficient at producing acetyl-CoA (C2), which is
 the universal precursor for synthesizing lipids. Here, we introduce in
 Arabidopsis thaliana a new-to-nature CO2 fixing cycle, Malyl-coA-glycerate
 (McG) cycle, which, together with the CBB cycle, forms a dual-cycle CO2
 fixation system. This cycle can fix one additional carbon by
 phosphoenolpyruvate carboxylase and convert the photorespiration product,
 glycolate, to acetyl-CoA. Plants with the McG cycle show enhanced protein
 abundance in their photosystems and enhanced photosystem II efficiency.
 McG plants had doubled CO2 fixation rates under atmospheric CO2,
 increased lipid production, pronounced growth enhancement, and tripled the
 seed yield.
description: # Dual-cycle CO2 fixation enhances growth and lipid synthesis in
 Arabidopsis thaliana Dataset DOI:
 [10.5061/dryad.h9w0vt4vf](10.5061/dryad.h9w0vt4vf) ## Description of the
 data and file structure The adp3528_Raw_data.zip file contains data for
 generating figures and tables in "Dual-cycle CO2 fixation enhances
 growth and lipid synthesis in Arabidopsis thaliana" by Lu et al.,
 2025. The Fig.1 to 5 are used for generating the main figures Fig.1 to
 Fig. 5. The data in Fig.S1 to Fig. S32 are used to generate supplementary
 figures FigS1 to FigS32. The data in TableS1 to TableS7 are used for
 generating supplementary tables TableS1 to TableS7. The plants used in
 this study are Arabidopsis wild-type (WT) plants, Arabidopsis transgenic
 plants expressing complete McG (McG_1, McG_2, McG_3, McG_4), plants
 expressing partial McG for catalyzing PEP and CO2 to acetyl-coA and
 glyoxylate (Mc_1, Mc_2, Mc_3), plants expressing partial McG for
 catalyzing glyoxylate to glycerate (G_1, G_2, G_3), and plants expressing
 McG without GDH (McG-~GDH~_1, McG-~GDH~_2, McG-~GDH~_3). Their growth
 phenotypes were imaged at different days after plant germination, at
 different light periods, on argar plates, under greenhouse without
 temperature controls, under ambient CO~2~ (400 ppm) and high CO~2~ (3000
 ppm), under normal light intensity (100 uE) and higher light intensities
 (150 and 300 uE). Light micrographs of mesophyll cells, scanning electron
 micrographs of shoot apical meristem with leaf primordia, transmission
 electron micrographs of leaf and seed sections, confocal images of lipid
 droplets in leaves are taken by using WT and McG plants. Photosynthetic
 carbon assimilation rate, light-response carbon assimilation rate, and
 quantum efficiency of photosystem II of the WT, McG, Mc, G, and McG-GDH
 plants are measured by LI-COR LI6800. The lipid analyses and proteomic
 analyses data of the WT and the McG plants are generated by using R, which
 R scripts and the raw data excel files are provided in the same folder.
 Statistical analyses are performed by using GraphPad Prism 10.3.0.  ####
 File: adp3528_Raw_data.zip **Description:** The file contains Fig. 2- Fig.
 5 for generating the main figures in the main text, Fig. S1-S26 and Table
 S1-S7 for the figures and tables in supplementary materials. **Fig2
 folder**: (1) The file Fig2_A.csv contains fresh weight (FW) mg of
 2-week-old WT and four McG line plants (McG_1,_2,_3,_4) Number of
 variables: 5 Variable list: * genotype: wild-type (WT), McG_1, McG_2,
 McG_3, McG_4 - bio-rep: 4 biological repeats * FW mg: fresh weight
 (milligram; mg) - AVG: average of fresh weight * SE: standard error of
 mean Statistical analysis: Shapiro-Wilk test for data normality test.
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value (2) The
 file Fig2_B.csv contains rosette leaf area of 4-week-old WT and four McG
 line plants (McG_1,_2,_3,_4) Number of variables: 5 Variable list: *
 genotype: wild-type (WT), McG_1, McG_2, McG_3, McG_4 - bio-rep: 1-5
 biological repeats * rosette leaf area (cm2) - AVG cm2: average of rosette
 leaf area (cm2) * SE: standard error of mean Statistical analysis:
 Shapiro-Wilk test for data normality test. Welch's ANOVA test for
 One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value (3) The
 file Fig2_C.csv contains dry weight (mg) of 4-week-old WT and four McG
 line plants (McG_1,_2,_3,_4) Number of variables: 5 Variable list: *
 genotype: wild-type (WT), McG_1, McG_2, McG_3, McG_4 - bio-rep: 1-4
 biological repeats * DW mg: dry weight (milligram; mg) - AVG: average of
 dry weight * SE: standard error of mean Statistical analysis:
 Shapiro-Wilk test for data normality test. Welch's ANOVA test for
 One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value (4) The
 file Fig2_D.csv contains dry weight (mg) of 6-week-old WT and four McG
 line plants (McG_1,_2,_3,_4) Number of variables: 5 Variable list: *
 genotype: wild-type (WT), McG_1, McG_2, McG_3, McG_4 - bio-rep: 9-13
 biological repeats * DW mg: dry weight (milligram; mg) - AVG: average of
 dry weight * SE: standard error of mean Statistical analysis:
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value (5) The
 file Fig2_E.csv contains the rosette leaf number of the 6-week-old plants
 of 6-week-old WT and four McG line plants (McG_1,_2,_3,_4) Number of
 variables: 5 Variable list: * genotype: wild-type (WT), McG_1, McG_2,
 McG_3, McG_4 - bio-rep: 4 biological repeats * leaf number: rosette leaf
 number of a plant - AVG: average of rosette leaf number of a plant * SE:
 standard error of mean Statistical analysis: Welch's ANOVA test for
 One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value (6)
 Fig.2F_WT, \_McG1, \_McG2, \_McG_3, \_McG4 are images of individual
 rosette leaves from 6-week-old WT and McG plants Fig.2G.csv contains the
 silique number from the primary inflorescence of the WT and McG plants at
 14 days after fertilization (DAF) Number of variables: 5 Variable list: *
 genotype: wild-type (WT), McG_1, McG_2, McG_3, McG_4 - bio-rep: 6
 biological repeats * silique per shoot: silique number from the primary
 inflorescence - AVG: average of silique number from the primary
 inflorescence * SE: standard error of mean Statistical analysis:
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value (7)
 Fig.2H.csv is the representive image of siliques from the primary
 inflorescence of the WT and McG plants (8) Fig.2I.csv contains the seed
 number per silique of WT and McG plants Number of variables: 5 Variable
 list: * genotype: wild-type (WT), McG_1, McG_2, McG_3, McG_4 - bio-rep: 6
 biological repeats * Seeds per silique: seed number per silique - AVG:
 average of seed number per silique * SE: standard error of mean
 Statistical analysis: Shapiro-Wilk test for data normality test.
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value (9)
 Fig.2J.csv contains the seed weight per seed of WT and McG plants. Seed
 weight was calculated as total seed weight divided by the number of seeds
 per plant Number of variables: 5 Variable list: * genotype: wild-type
 (WT), McG_1, McG_2, McG_3, McG_4 - bio-rep: 5 biological repeats * seed
 weight ug/seed: weight  (microgram) per seed - AVG: average of weight 
 (microgram) per seed * SE: standard error of mean Statistical analysis:
 Shapiro-Wilk test for data normality test. Welch's ANOVA test for
 One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value (10)
 Fig.2J.csv contains the number of seeds per plant Number of variables: 5
 Variable list: * genotype: wild-type (WT), McG_1, McG_2, McG_3, McG_4 -
 bio-rep: 5 biological repeats * seed number : seed number per plant - AVG:
 average of seed number per plant * SE: standard error of mean Statistical
 analysis: Shapiro-Wilk test for data normality test.
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value **Fig3
 folder**: (1) "Fig3A.csv" is used for analyzing carbon
 assimilation rates of WT and four McG lines. Number of variables: 5
 Variables: * Four genotypes: WT, McG_1, McG_2, McG_3, McG_4 - AVG Ci:
 Average of intercellular concentration of CO~2~ (µmol mol⁻¹; Ci) from
 three measurements of three plant replicates * SE Ci: standard errors of
 intercellular concentration of CO~2~ (µmol mol⁻¹; Ci) from three
 measurements of three plant replicates - AVG A: Average of net carbon
 assimilation rate (µmol m⁻² s⁻¹; A) from three measurements of three plant
 replicates * SE A: standard errors of net carbon assimilation rate (µmol
 m⁻² s⁻¹; A) from three measurements of three plant replicates (2)
 Fig3B.csv contains A400 (CO2 umol m-2s-1) of WT and four McG lines. Number
 of Variables: 4 Variables: * genotype: WT, McG_1, McG_2, McG_3, McG_4 -
 bio-rep: 3 biological replicate for WT and each McG line * A400: net
 carbon assimilation rate measured under CO2 concentration of 400 umol
 mol-1. - AVG: average of A400 of biological replicates * SE: standard
 error of mean of A400 of biological replicates (3) Fig3C.csv contains Amax
 (CO2 umol m-2s-1) of WT and four McG lines. Number of Variables: 4
 Variables: * genotype: WT, McG_1, McG_2, McG_3, McG_4 - bio-rep: 3
 biological replicate for WT and each McG line * Amax: maximal net carbon
 assimilation rate (Amax) measured under CO2 concentrations higher then
 1200 umol mol-1. - AVG: average of Amax of biological replicates * SE:
 standard error of mean of Amax of biological replicates (4)
 "Fig3D.csv" is used for determining CO~2~ compensation points
 (Ci*) for WT and McG plants. Number of variables: 5 Variables: * Four
 genotypes: WT, McG_1, McG_2, McG_3, McG_4 - repeats: three biological
 repeats from each line * Ci* (umol mol-1): CO~2~ compensation points (Ci*)
 for WT and McG plants - AVG: average values of CO~2~ compensation points
 (Ci*) * SE: standard errors of means of CO~2~ compensation points (Ci*)
 (5) "Fig3E.csv" contains data net CO~2~ assimilation rates at
 various light intensities were measured under ambient CO~2~ (400 ppm) for
 plants grown under ambient CO~2~ condition. Number of variables: 6
 Variables: * Four genotypes: WT, McG_1, McG_2, McG_3, McG_4 - repeats:
 four biological repeats from each line * Qin (µmol m⁻² s⁻¹): various light
 intensities from 1500 µmol m⁻² s⁻¹ to 0 µmol m⁻² s⁻¹ - A (µmol m⁻² s⁻¹):
 net carbon assimilation rate * AVG: average A values of four biological
 repeats - SE: standard errors of means of A values of four biological
 repeats (6) "Fig3F.csv" file contains the net CO~2~ assimilation
 rates at various light intensities were measured under 1500 ppm CO~2~ for
 plants grown under high CO~2~ (3000ppm) condition Number of variables: 6
 Variables: * Four genotypes: WT, McG_1, McG_2, McG_3, McG_4 - repeats:
 four biological repeats from each line * Qin (µmol m⁻² s⁻¹): various light
 intensities from 1500 µmol m⁻² s⁻¹ to 0 µmol m⁻² s⁻¹ - A (µmol m⁻² s⁻¹):
 net carbon assimilation rate * AVG: average A values of four biological
 repeats - SE: standard errors of means of A values of four biological
 repeats **Fig4 folder**: (1) "Fig4A.csv" file contains ^13^CO~2~
 incorporation data into metabolites of McG cycle and carbohydrate
 precursor glucose-1-phosphate (G1P) in WT and McG plants Number of
 variables: 5 Variables: * Four genotypes: WT, McG_1, McG_2, McG_3, McG_4 -
 Six metabolites: G1P: glucose-1-phosphate; PEP: phosphoenolpyruvate;
 malate, Tartonic semialdehyde; 2PG: 2-phosphoglycerate; acetyl-coA. *
 13C/12C: The ratio of ^13^C-labelled to ^12^C-unlabelled metabolite. M1:
 one carbon molecule of above metabolite is ^13^C-labeled; M2: two carbon
 molecules of above metabolite are ^13^C-labeled; M3: three carbon
 molecules of above metabolite are ^13^C-labeled; M4: four carbon molecules
 of above metabolite are ^13^C-labeled; M5: five carbon molecules of above
 metabolite are ^13^C-labeled; M6: six carbon molecules of above metabolite
 are ^13^C-labeled. - AVG: average value of 13C/12C ratios from 6
 biological replicates of each line. * SE: standard errors of of 13C/12C
 ratios from 6 biological replicates of each line. - 13C/12C (total): Sum
 of 13C /12C ratios for each line for statistical analysis of significance
 difference between WT and McG lines. Statistical analysis:
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value (2)
 "Fig4B.csv" file contains data acquired from GC-MSMS analytic
 experiments, which are used for determining the contents of the
 photorespiration metabolites. Number of variables: 5 Variables: *
 genotypes: WT, McG_1, McG_2, McG_3, McG_4 - bio-rep: 5-6 biological
 replicates * metabolite/ribitol ratio: the peak area of the metabolite to
 the peak area of the ribitol. Metabolites are glycolate, glyoxylate,
 serine, glycin, glycerate, and pyruvate. - AVG: average value
 of metabolite/ribitol ratios of biological replicates * SE: standard error
 of metabolite/ribitol ratios of biological replicates Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value **Fig5 folder:** (1) The first folder (Fig. 5A Nile red
 staining leaves) contains images acquired from a confocal microscopy
 experiment for visualizing lipid droplets in leaves. (2) The folder "
 Fig5B Leaf TEM LD" contains transmission electron micrographs (TEM)
 of the leaves for visualizing lipid droplets (LD) in the WT and McG lines.
 (3) The folder "Fig5C green seed TEM" contains TEMs acquired
 from green seeds of each line for visualizing their seed LDs. (4) The file
 " Fig5D.csv" contains the concentrations of different lipid
 classes in the leaves of the WT and the McG
 plants."LipidBarByClass.R", is used to calculate the
 concentration of each lipid class. Number of variables: 3 Variables: *
 genotypes: WT, McG_1, McG_2, McG_3, McG_4 - bio-rep: 5-6 biological
 replicates * Lipid class: TAG: triacylglycerol;
 MGDG: monogalactosyldiacylglycerol; DGDG: digalactosyldiacylglycerol; FA:
 fatty acid. Concentration unit: ug g-1 Fw Statistical analysis: 
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05)
 Dunnett's T3 multiple comparisons test for adjusted-P-value (5)  The
 file "Fig5E.csv" contains the data for analyzing chain length
 distribution in triacylglycerol (TAG) by R coding file
 "LipidFADis.R". Number of variables: 2 * genotype and biological
 replicates: WT_1 to WT_6 (six biological replicates for WT), McG1_1,
 McG1_3, McG1_4, McG1_5, McG1_6 (five biological replicates for McG_1
 line), McG2_1 to McG2_6 (six biological replicates for McG_2 line), McG3_1
 to McG3_6 (six biological replicates for McG_3 line), McG4-1 to McG4-6
 (six biological replicates for McG_4 line). - TG chain lengths:
 concentrations of different chain lengths in each triacylglycerol (TG) (6)
 The file "Fig5F.csv" contains the data for the concentrations of
 different lipid classes in the seeds of the WT and the McG
 plants."LipidBarByClass.R", is used to calculate the
 concentration of each lipid class. Number of variables: 3 Variables: *
 genotypes: WT, McG_1, McG_2, McG_3, McG_4 - bio-rep: 5-6 biological
 replicates * Lipid class: TAG: triacylglycerol; DAG: diacylglycerol; PC:
 phosphatidylcholine; FA: fatty acid. Concentration unit: ug g-1 Fw
 Statistical analysis: Welch's ANOVA test for One-Way-ANOVA test
 (P-value &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value (7) The file "Fig5G.csv" contains the data for
 analyzing chain length distribution in triacylglycerol (TAG) of the seeds
 from WT and McG lines, by using R coding file "LipidFADis.R".
 Number of variables: 2 * genotype and biological replicates: WT_1 to WT_6
 (six biological replicates for WT), McG1_1 to McG_1_6 (six biological
 replicates for McG_1 line), McG_2_1 to McG_2_6 (six biological replicates
 for McG_2 line), McG_3_1 to McG_3_6 (six biological replicates for McG_3
 line), McG_4-1 to McG_4-6 (six biological replicates for McG_4 line). - TG
 chain lengths: concentrations of different chain lengths in each
 triacylglycerol (TG) 
 
 **FigS2 folder**: (1) The file "FigS2A.csv" showed that
 expression of MTK fusion protein and MCL can rescue the growth defect of
 the acetyl-coA auxotrophic ˆstrain (LYN2-1), which cannot metabolize
 glucose to acetyl-coA for growth. Number of variables: 3 Variables: *
 genotypes: *E. coli* LYN2-1strain and the *E. coli* LYN2-1strain
 expressing with MTK fusion protein and MCL  (1, 12, 17, 19) - incubation
 time (h: hour): 0, 24, 48, 72h * OD600: absorption at 600 nm wavelength at
 different incubation time point (0, 24, 48, 72h) (2) The file
 "FigS2B_Mc_plasmid_pCambia1300-35S-1B-3FLAG-PPC_Acc65I-35S__1B-A-B_35S__1B-Mcl-Xba1 Correct nanopor rapid.fasta" and "FigS2B_G_plamid_map_pEarleygate103-U10p-1B-CrGDH-1B-Gcl-1B-Tsr.fasta" contain the maps of the Mc and G constructs. (3) The files "FigS2C_GDH_GCL_TSR_Untitled 55", "FigS2C_PPC_MTK_MCL_Untitled 42", and "FigS2C_loading_CBS_IMG_5768" are the images acquired from immunoblotting to show protein expressions of the McG transgenes in the Mc and G transgenic lines. **FigS3 folder**: (1) The file "FigS3A.csv" demonstrated the transcript levels of the McG transgenes in the McG lines. Total RNA isolated from 2-week-old WT and McG plants and used for digital qPCR to quantitate the transcript levels of PPC, MtKA-B, Mcl, GDH, Gcl and Tsr. The expression of the endogenous granule-bound starch synthase (GBSS) gene was used as the internal control. Number of variables: 5 Variables: * genotype: WT. McG_1, McG_2, McG_3, McG_4 - bio-rep: 3 biological replicates for each line * Transcript levels of *PPC*, *MtkA_B, Mcl, GDH, Gcl,* and *Tsr *to the transcript level of endogenous GBSS: PPC/GBSS, MTK/GBSS, MCL/GBSS, GDH/GBSS, GCL/GBSS, and TSR/GBSS. - AVG: average transcriptional level of three biological replicates * SE: standard error of mean of transcriptional level of three biological replicates. (2) The file "FigS3B_individual_McG_plants_Untitled-1", "FigS3B_loading_CBS_IMG_4358" showed the protein expression levels of the McG transgenes in the WT and McG lines. (3) The folder "FigS3C_nonopore WT McG" contains raw reads (.fasta files) acquired from Nanopore whole genome sequencing for the WT and the McG lines. **FigS4 folder**: The file "FigS4_McG-GDH_Untitled 45-1.tif" contains immunoblotting results for the detection of heterologous PPC, MTK, MCL, GCL, and TSR protein expression in McG-GDH lines. The file "FigS4_loading_CBS_IMG_5771_1.tif" for loading control.  **FigS5 folder**: This folder contains photographs of the WT and McG plants for 4 weeks, 6 weeks, and 7 weeks. **FigS6 folder**: This folder contains photographs of WT (white label), selected F3 homozygous McG_1 (pink), McG_2 (yellow/orange), McG_3 (green), McG_4 (blue) plants. **FigS7 folder**: (1) The subfolders " FigS7A_10d", " FigS7A_6_7wk", "FigS7A_9wk", "Fig.S7B", "FigS7C_4wk", "FigS7C_6wk", and "FigS7D_6wk" contain images for showing growth phenotypes of WT and McG plants in FigS7A, FigS7B, FigS7C, and FigS7D. (2) The file "FigS7C_6wk.csv" in the "FigS7C_6wk" folder contains the fresh weight (FW) and the dry weight (DW) of 6-week-old WT and McG plants. Number of variables: 8 Variables: * genotype: WT. McG_1, McG_2, McG_3, McG_4 - bio-rep: 9-13 biological replicates for each line * FW (g): fresh weight (gram) - FW AVG: average FW of biological replicates * FW SE: standard error of mean FW of biological replicates - DW (mg): dry weight (milligram) * DW AVG: average DW of biological replicates - DW SE: standard error of mean DW of biological replicates Statistical analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05). Dunnett's T3 multiple comparisons test for adjusted-P-value (3) The file "FigS7D.csv" in the "FigS7D_6wk" folder contains the fresh weight (FW) and the dry weight (DW) of 6-week-old WT and McG plants. Number of variables: 8 Variables: * genotype: WT. McG_1, McG_2, McG_3, McG_4 - bio-rep: 8 biological replicates for each line * FW (g): fresh weight (gram) - FW AVG: average FW of biological replicates * FW SE: standard error of mean FW of biological replicates - DW (mg): dry weight (milligram) * DW AVG: average DW of biological replicates - DW SE: standard error of mean DW of biological replicates Statistical analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05). Dunnett's T3 multiple comparisons test for adjusted-P-value. **FigS8 folder**: This folder contains photographs of the WT and McG plants for 4, 6, and 9 weeks under greenhouse conditions. **FigS9 folder**: This folder contains photographs of the WT and McG plants grown under 8h light/16h dark (8_16) short day or 16h light/ 8h dark (16_8) long day conditions. **FigS10 folder**: (1) The file "FigS10A_leaf primordia.pptx" contains scanning electron micrographs of shoot apical meristems from the WT and McG plants. (2) The file "FigS10B_longitudinal SAM_Highlight phenotype Arabidopsis shoot apical meristem LM.pptx" contains light micrographs of longitudinal shoot apical meristems for the WT and the McG plants. (3) The file "FigS10C.csv" contains raw data for calculating leaf emergence rates of the WT and McG lines. Number of variables: 4 Variables:  * genotype: WT. McG_1, McG_2, McG_3, McG_4 - bio-rep: 5-7 biological replicates for each lines * DAG: days after germination - leaf number: rosette leaf number per plant 
 
 **FigS12 folder**: (1) The folder "FigS12AB_opensilique_seeds"
 contains light micrographs of green open siliques and their seeds from the
 WT and McG lines. (2) The folder "FigS12C_green seed
 blightfield" contains light micrographs of the green seeds from the
 WT and McG lines. (3) The folder "FigS12D_mature seed BF"
 contains the micrographs of the mature seeds from the WT and the McG
 lines. (4) The file "FigS12E_csv" contains raw data for
 calculating seed germination rates of the WT and McG lines. Number of
 variables: 4 Variables:  * genotype: WT. McG_1, McG_2, McG_3, McG_4 -
 bio-rep: 4 plate replicates for each lines * initial seed number: number
 of seeds on each plate - hr after sow: hour after sowing seeds
 under 12hr/12hr light/dark cycle, light intensity of 100 𝜇mol m^-2^ s^-1^
 * germination number: number of seeds are germinated - %: germination
 rate: % of germination number/initial seed number * AVG: average
 germination rate from 4 plate replicates of each line - SE: standard error
 of mean of germination rate from 4 plates of each line **FigS13 folder**:
 This folder contains photographs for 9-week-old and 3-month-old WT and McG
 plants **FigS14 folder**: (1) The folder "FigS14A_leaf cross
 section" contains cross-section light micrographs of 2-week-old
 leaves from the WT and McG lines.  (2) The file "FigS14B.csv"
 contains raw data for calculating the proportion of each ploidy number
 (2C-16C) of the WT and McG lines. Number of variables: 17 Variables: *
 genotype: WT. McG_1, McG_2, McG_3, McG_4 - bio-rep: 4 biological
 replicates for each lines * proportion of each ploidy: proportion of 2C
 (diploid), proportion of 4C (diploid number of 2), proportion of 8C
 (diploid number of 4), proportion of 16C (diploid number of 8), and
 proportion of 32C (diploid number of 16). - 2C AVG: averaged diploid
 number of 4 biological replicates for each line * 2C SE: standard error of
 mean of diploid number of 4 biological replicates for each line - 4C
 AVG: averaged 4C of 4 biological replicates for each line * 4C SE:
 standard error of mean of 4C of 4 biological replicates for each line - 8C
 AVG: averaged 8C of 4 biological replicates for each line * 8C SE:
 standard error of mean of 8C of 4 biological replicates for each line -
 16C AVG: averaged 16C of 4 biological replicates for each line * 16C SE:
 standard error of mean of 16C of 4 biological replicates for each line -
 32C AVG: averaged 32C of 4 biological replicates for each line * 32C SE:
 standard error of mean of 32C of 4 biological replicates for each line
 **FigS15 folder**: (1) The folder "FigS15A_14d root" contains
 photographs of 14-day-old seedlings of the WT and McG lines. (2) The file
 "FigS15B.csv" contains raw data for root weights of the WT and
 McG lines. Number of variables: 5 Variables:  * genotype: WT. McG_1,
 McG_2, McG_3, McG_4 - no: 6 biological replicates for each line * mg:
 milligram of root weight of each plant - AVG: average of root weight of 6
 biological replicates * SE: standard error of mean of root weight of 6
 biological replicates Statistical analysis: Welch's ANOVA test for
 One-Way-ANOVA test (P-value&lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value. (3) The
 file "FigS15C_IMG_5395" represents 7-week-old root phenotypes of
 the soil-grown WT and McG lines. **FigS16 folder**: This folder contains
 two replicate experiments of Fig. S15A **FigS17 folder**: (1) The file
 "FigS17_ABCD.csv_contains raw data of the fresh weight (FW) and dry
 weight (DW) of 4-week-old WT, McG, Mc, G, and McG-GDH lines. Number of
 variables: 8 Variables: * genotype: WT, McG_1, McG_2, McG_3, McG_4, Mc_1,
 Mc_2, Mc_3, G_1, G_2, G_3, McG-GDH_1, McG-GDH_2, McG-GDH_3 - no: 6
 biological replicates for each line * FW mg: milligram of fresh weight of
 a plant - FW AVG: average fresh weight of 6 biological replicates * FW SE:
 standard error of mean of FW of 6 biological replicates - DW mg: milligram
 of dry weight of a plant * DW AVG: average dry weight of 6 biological
 replicates - DW SE: standard error of mean of DW of 6 biological
 replicates Statistical analysis: Welch's ANOVA test for One-Way-ANOVA
 test (P-value &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (2) The file "FigS17BD statistics.ppt"
 contains adjusted p-values between multiple lines by using One-way Welch’s
 ANOVA test, followed by Dunnett’s T3 multiple pairwise comparison. (3) The
 images "FigS17E_2wk_IMG_5696.jpeg",
 "FigS17E_3wk_IMG_5724.jpeg", and
 "FigS17E_4wk_IMG_5752" represent the growth phenotypes of the
 lines for 2-week, 3-week, and 4-week-old under ambient CO~2~ condition.
 **FigS18 folder**: There are four replicate experiments of Fig.S17,
 labeled as "FigS18A_1_3wk", "FigS18B_1_3wk",
 "FigS18C_4_7wk", and "FigS18D_4wk". (1) These four
 folders contain images showing growth phenotypes of WT, McG, Mc, G, and
 McG-GDH lines under ambient CO~2~ condition. (2) The file
 "FigS18D.csv" in the FigS18D_4wk contains FW and DW of
 32-day-old WT, McG, Mc, G, and McG-GDH plants under ambient CO~2~, light
 intensity of 100 𝜇mol m^-2^ s^-1^ (100uE). Number of variables: 8
 Variables:  * genotype: WT, McG_1, McG_2, McG_3, McG_4, Mc_1, Mc_2, G_1,
 G_2, McG-GDH_1, McG-GDH_2 - bio-rep: 8 biological replicates for each line
 * FW g: gram of fresh weight of a plant - FW AVG: average fresh weight of
 8 biological replicates * FW SE: standard error of mean of FW of 8
 biological replicates - DW mg: milligram of dry weight of a plant * DW
 AVG: average dry weight of 8 biological replicates - DW SE: standard error
 of mean of DW of 8 biological replicates Statistical analysis:
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value. **FigS19
 folder**: (1) The file "FigS19ABCD.csv" contains raw data of the
 FW and DW of the WT, McG, Mc, G, and McG-GDH lines under high CO~2~ (3000
 ppm) conditions. Number of variables: 8 Variables:  * genotype: WT, McG_1,
 McG_2, McG_3, McG_4, Mc_1, Mc_2, Mc_3, G_1, G_2, G_3, McG-GDH_1,
 McG-GDH_2. McG-GDH_3 - bio-rep: 6 biological replicates for each line * FW
 mg: milligram of fresh weight of a plant - FW AVG: average fresh weight of
 6 biological replicates * FW SE: standard error of mean of FW of 6
 biological replicates - DW mg: milligram of dry weight of a plant * DW
 AVG: average dry weight of 6 biological replicates - DW SE: standard error
 of mean of DW of 6 biological replicates Statistical analysis:
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value. Table of
 adjusted-P-value for FW Table of adjusted-P-value for DW (2) The folders
 "FigS19E_2wk_111224 3000" and "FigS19E_4wk_271224
 3000" contain photographs of the WT, McG, Mc, G, and McG-GDH lines
 for 2-week-old and 4-week-old plants. **FigS20 folder**: The replicate
 experiment of Fig. S19E for growth phenotype observations. **FigS21
 folder**: (1) The folder "FigS21A_150E" contains photographs of
 the plants grown under 150 uE light intensity for 2-4 weeks. (2) The
 folder "FigS21B_300E" contains photographs of the plants grown
 under 300 uE light intensity for 2-4 weeks. (3) The file
 "FigS21A_150E.csv" in the 4weekold subfolder of the FigS21A_150E
 folder contains FW and DW of the 4-week-old plants under light intensity
 of 150 𝜇mol m^-2^ s^-1^ (150 μE). Number of variables: 8 Variables:  *
 genotype: WT, McG_1, McG_3, Mc_1, Mc_2, G_1, G_2, McG-GDH_1, McG-GDH_2. -
 bio-rep: 8 biological replicates for each line * FW g: gram of fresh
 weight of a plant - FW AVG: average fresh weight of 8 biological
 replicates * FW SE: standard error of mean of FW of 8 biological
 replicates - DW mg: milligram of dry weight of a plant * DW AVG: average
 dry weight of 8 biological replicates - DW SE: standard error of mean of
 DW of 8 biological replicates Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. Table of adjusted-P-value for FW Table
 of adjusted-P-value for DW (4) The file "FigS21B_300E.csv" in
 the 4weekold subfolder of the FigS21B_300E folder contains FW and DW of
 the 4-week-old plants under light intensity of 300 𝜇mol m^-2^ s^-1^ (300
 μE) Number of variables: 8 Variables:  * genotype: WT, McG_1, McG_3, Mc_1,
 Mc_2, G_1, G_2, McG-GDH_1, McG-GDH_2. - bio-rep: 8 biological replicates
 for each line * FW g: gram of fresh weight of a plant - FW AVG: average
 fresh weight of 8 biological replicates * FW SE: standard error of mean of
 FW of 8 biological replicates - DW mg: milligram of dry weight of a plant
 * DW AVG: average dry weight of 8 biological replicates - DW SE: standard
 error of mean of DW of 8 biological replicates Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. Table of adjusted-P-value for FW Table
 of adjusted-P-value for DW **FigS22 folder**: (1) The
 "FigS22A.csv" file contains net carbon assimilation rates (A400
 [CO~2~ µmol m⁻² s⁻¹] ) of 4-week-old WT and McG plant leaves under ambient
 CO~2~ (400 ppm) Number of variables: 3 Variables:  * genotype: WT, McG_1,
 McG_2, McG_3, McG_4 - leaf number: 10th, 12th, 14th leaves of WT and McG
 plants are used for A400 measurements * A400 (µmol m⁻² s⁻¹): net carbon
 assimilation rate at ambient CO~2~ (400 ppm) Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (2) The "FigS22B.csv" file contains leaf mass
 per area (LMA; g m^-2^) of 10th, 12th, 14th leaves from 4-week-old WT and
 McG plants grown under ambient CO~2~ (400 ppm) Number of variables: 3
 Variables:  * genotype: WT, McG_1, McG_2, McG_3, McG_4 - leaf number:
 10th, 12th, 14th leaves of WT and McG plants * LMA (g m⁻²¹): leaf mass per
 area (LMA; g m^-2^) Statistical analysis: Welch's ANOVA test for
 One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value. (3) The
 "FigS22C.csv" file contains chlorophyll content (g m^-2^) of
 10th, 12th, 14th leaves from 4-week-old WT and McG plants grown under
 ambient CO~2~ (400 ppm) Number of variables: 3 Variables:  * genotype: WT,
 McG_1, McG_2, McG_3, McG_4 - leaf number: 10th, 12th, 14th leaves
 of 4-week-old WT and McG plants * chlorophyll content (g m⁻²): chlorophyll
 content of 10th, 12th, 14th leaves of 4-week-old WT and McG plants
 Statistical analysis: Welch's ANOVA test for One-Way-ANOVA test
 (P-value &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (4) The "FigS22D.csv" file contains A400 (
 CO~2~ µmol m⁻² s⁻¹) normalized to LMA (g m^-2^) of 10th, 12th, 14th leaves
 from 4-week-old WT and McG plants grown under ambient CO~2~ (400 ppm) as
 to A400/LMA (CO~2~ µmol g⁻1 s⁻¹) Number of variables: 3 Variables:  *
 genotype: WT, McG_1, McG_2, McG_3, McG_4 - leaf number: 10th, 12th, 14th
 leaves of 4-week-old WT and McG plants * A400/LMA : A400 normalized to LMA
 of 10th, 12th, 14th leaves of 4-week-old WT and McG plants Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (5) The "FigS22E.csv" file contains A400 (
 CO~2~ µmol m⁻² s⁻¹) normalized to chlorophyll content (g m^-2^) of 10th,
 12th, 14th leaves from 4-week-old WT and McG plants grown under ambient
 CO~2~ (400 ppm) as to A400/chll (CO~2~ µmol g⁻1 s⁻¹) Number of variables:
 3 Variables:  * genotype: WT, McG_1, McG_2, McG_3, McG_4 - leaf number:
 10th, 12th, 14th leaves of 4-week-old WT and McG plants * A400/Chll : A400
 normalized to chlorophyll contents of 10th, 12th, 14th leaves
 of 4-week-old WT and McG plants Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. **FigS23 folder**: (1) The file "FigS23A.csv"
 contains net carbon assimilation rates (A400 [CO~2~ µmol m⁻² s⁻¹] ) of
 5-week-old WT and McG plant leaves under ambient CO~2~ (400 ppm) Number of
 variables: 3 Variables:  * genotype: WT, McG_1, McG_2, McG_3, McG_4 - leaf
 number: 10th, 12th, 14th, 16th leaves of WT and McG plants are used for
 A400 measurements * A400 (µmol m⁻² s⁻¹): net carbon assimilation rate at
 ambient CO~2~ (400 ppm) Statistical analysis: Welch's ANOVA test for
 One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value. (2) The
 "FigS23B.csv" file contains leaf mass per area (LMA; g-1 m-2) of
 10th, 12th, 14th leaves from 5-week-old WT and McG plants grown under
 ambient CO~2~ (400 ppm) Number of variables: 3 Variables:  * genotype: WT,
 McG_1, McG_2, McG_3, McG_4 - leaf number: 10th, 12th, 14th, 16th leaves of
 WT and McG plants * LMA (g m⁻²¹): leaf mass per area (LMA; g-1 m-2)
 Statistical analysis: Welch's ANOVA test for One-Way-ANOVA test
 (P-value &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (3) The "FigS23C.csv" file contains
 chlorophyll content (g-1 m-2) of 10th, 12th, 14th, 16th leaves from
 5-week-old WT and McG plants grown under ambient CO~2~ (400 ppm) Number of
 variables: 3 Variables:  * genotype: WT, McG_1, McG_2, McG_3, McG_4 - leaf
 number: 10th, 12th, 14th, 16th leaves of 5-week-old WT and McG plants *
 chlorophyll content (g m⁻²): chlorophyll content of 10th, 12th, 14th, 16th
 leaves of 5-week-old WT and McG plants Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (4) The "FigS23D.csv" file contains A400 (
 CO~2~ µmol m⁻² s⁻¹) normalized to LMA (g-1 m-2) of 10th, 12th, 14th, 16th
 leaves from 5-week-old WT and McG plants grown under ambient CO~2~ (400
 ppm) as to A400/LMA (CO~2~ µmol g⁻1 s⁻¹) Number of variables: 3
 Variables:  * genotype: WT, McG_1, McG_2, McG_3, McG_4 - leaf number:
 10th, 12th, 14th leaves of 5-week-old WT and McG plants * A400/LMA : A400
 normalized to LMA of 10th, 12th, 14th, 16th leaves of 5-week-old WT and
 McG plants Statistical analysis: Welch's ANOVA test for One-Way-ANOVA
 test (P-value &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (5) The "FigS23E.csv" file contains A400 (
 CO~2~ µmol m⁻² s⁻¹) normalized to chlorophyll content (g-1 m-2) of 10th,
 12th, 14th, 16th leaves from 5-week-old WT and McG plants grown under
 ambient CO~2~ (400 ppm) as to A400/chll (CO~2~ µmol g⁻1 s⁻¹) Number of
 variables: 3 Variables:  * genotype: WT, McG_1, McG_2, McG_3, McG_4 - leaf
 number: 10th, 12th, 14th, 16th leaves of 5-week-old WT and McG plants *
 A400/Chll : A400 normalized to chlorophyll contents of 10th, 12th, 14th,
 16th leaves of 5-week-old WT and McG plants Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. **FigS24 folder**: (1) The "FigS24A.csv" file
 contains raw data for net carbon assimilation rates of the WT and McG
 lines under various CO~2~ concentrations, and light intensity of 100 uE.
 Number of variables: 8 Variables:  * genotype: WT, McG_1, McG_2, McG_3,
 McG_4 - biological replicates: three replicates for WT, four replicates
 for McG_1, three replicates for McG_2, and four replicates for McG_3 and
 McG_4 * Ci (CO2 𝜇mol mol-1): intercellular CO2 concentrations (Ci) - A
 (CO2 𝜇mol m-2s-1): net carbon assimilation rate * AVG Ci (CO2 𝜇mol mol-1):
 average Ci value of biological replicates - AVG A (CO2 𝜇mol m-2s-1):
 average A value of biological replicates * SE Ci (CO2 𝜇mol mol-1):
 standard error of Ci mean of biological replicates - SE A (CO2 𝜇mol
 m-2s-1): standard error of A mean of biological replicates (2) The
 "FigS24B.csv" file shows the net carbon assimilation rates under
 ambient CO~2~ (400 ppm) as to A400 of  WT and McG plants in the
 FigS24A.csv. Number of variables: 4 Variables:  * genotype and biological
 replicates : three replicates for WT, four replicates for McG_1, three
 replicates for McG_2, and four replicates for McG_3 and McG_4 - A400 (CO2
 𝜇mol m-2s-1) under 400ppm CO2: net carbon assimilation rate under 400ppm
 CO2 * AVG A400 (CO2 𝜇mol m-2s-1): average A value of biological replicates
 - SE A400 (CO2 𝜇mol m-2s-1): standard error of A mean of biological
 replicates Statistical analysis: Welch's ANOVA test for One-Way-ANOVA
 test (P-value &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (3) The "FigS24C.csv" contains Water use
 efficiency (WUE) of WT and McG lines under light intensity of 100 𝜇mol
 m-2s-1, 400 𝜇mol mol-1 of CO~2~ Number of variables: 6 Variables:  *
 genotype and biological replicates : three replicates for WT, four
 replicates for McG_1, three replicates for McG_2, and four replicates for
 McG_3 and McG_4 - A400 (CO2 𝜇mol m-2s-1) under 400ppm CO2: net carbon
 assimilation rate under 400ppm CO2 * Emm: transpiration rate (mmol m⁻²
 s⁻¹) under 400pm CO2 - WUE (A/E) at 400ppm: water use efficiency is
 calculated as A400/Emm (µmol mmol-1) * AVG: average WUE value of
 biological replicates - SE: standard error of WUE mean of biological
 replicates Statistical analysis: Welch's ANOVA test for One-Way-ANOVA
 test (P-value &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (4) The "FigS24D.csv" file contains the day
 respiration rate of WT and McG lines by using the slope-intercept
 approach, which has been published by Walker et al, 2016 (reference 47).
 Number of variables: 4 Variables:  * genotype and biological replicates :
 three replicates for WT, McG_1, McG_2, McG_3, and McG_4 - Rd (umol m-2
 s-1): day respiration rate * AVG: average Rd of biological replicates -
 SE: standard error of Rd mean of biological replicates Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (5) The "FigS24E.csv" file contains net carbon
 assimilation rates of whole plants under various CO2 concentrations.
 Number of variables: 8 Variables:  * genotype and biological replicates :
 four replicates for WT, McG_1, McG_2, McG_3, and McG_4 - rosette area m2:
 whole rosette area of a plant (m2) * CO2_a (µmol mol⁻¹): ambient CO~2~
 concentration - A (µmol m⁻² s⁻¹): net carbon assimilation rate per leaf
 area * AVG Ca: average Ca of biological replicates - SE Ca: standard error
 of Ca mean of biological replicates * AVG A: average net carbon
 assimilation rate per leaf area of biological replicates - SE A: standard
 error of A mean per leaf area of biological replicates **FigS25 folder**:
 (1) The file "FigS25_A.csv" contains net CO2 assimilation rates
 for the WT and Mc plants under various CO2 concentrations. Number of
 variables: 7 Variables:  * genotype and replicates: three biological
 replicates for WT, Mc_1, Mc_2, and Mc_3 lines - Ci (µmol mol⁻¹):
 intercellular CO2 concentrations * A µmol m⁻² s⁻¹: net carbon assimilation
 rate - Ci AVG: average of Ci of biological replicates * Ci SE: standard
 error of Ci mean of biological replicates - A AVG: average of A of
 biological replicates * A SE: standard error of A mean of biological
 replicates (2) The file "FigS25_B.csv" contains net CO2
 assimilation rates for the WT and G plants under various CO2
 concentrations. Number of variables: 7 Variables:  * genotype and
 replicates: three biological replicates for WT, G_1, G_2, and G_3 lines -
 Ci (µmol mol⁻¹): intercellular CO2 concentrations * A µmol m⁻² s⁻¹: net
 carbon assimilation rate - Ci AVG: average of Ci of biological replicates
 * Ci SE: standard error of Ci mean of biological replicates - A AVG:
 average of A of biological replicates * A SE: standard error of A mean of
 biological replicates (3) The file "FigS25C.csv" contains net
 CO2 assimilation rates under various CO2 concentrations for the WT and
 McG-GDH plants. Number of variables: 7 Variables:  * genotype and
 replicates: three biological replicates for WT, McG~-GDH~_1, McG~-GDH~_2,
 and McG~-GDH~_3 lines - Ci (µmol mol⁻¹): intercellular CO2 concentrations
 * A µmol m⁻² s⁻¹: net carbon assimilation rate - Ci AVG: average of Ci of
 biological replicates * Ci SE: standard error of Ci mean of biological
 replicates - A AVG: average of A of biological replicates * A SE: standard
 error of A mean of biological replicates (4) The "FigS25D.csv"
 file contains A400 values for three replicates of WT and Mc_1, Mc_2, and
 Mc_3 lines showed in "FigS25A.csv" Number of variables: 4
 Variables:  * genotype and replicates: three biological replicates for WT,
 Mc_1, Mc_2, Mc_3 - A400 (CO2 𝜇mol m-2s-1): net carbon assimilation rate
 under 400ppm CO2 * AVG: average of A400 of replicates - SE: standard error
 of A400 mean of replicates Statistical analysis: Welch's ANOVA test
 for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value. (5) The
 "FigS25E.csv" file contains A400 values for three replicates of
 WT and G_1, G_2, and G_3 lines showed in "FigS25B.csv" Number of
 variables: 4 Variables:  * genotype and replicates: three biological
 replicates for WT, G_1, G_2, and G_3 line - A400 (CO2 𝜇mol m-2s-1): net
 carbon assimilation rate under 400ppm CO2 * AVG: average of A400 of
 replicates - SE: standard error of A400 mean of replicates Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value (6) The "FigS25F.csv" file contains A400 values
 for three replicates of WT and McG~-GDH~_1, McG~-GDH~_2, and McG~-GDH~_3
 lines showed in "FigS25C.csv" Number of variables: 4 Variables: 
 * genotype and replicates: three biological replicates for WT,
 McG~-GDH~_1, McG~-GDH~_2, McG~-GDH~_3 - A400 (CO2 𝜇mol m-2s-1): net carbon
 assimilation rate under 400ppm CO2 * AVG: average of A400 of replicates -
 SE: standard error of A400 mean of replicates Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. **FigS26 folder**: (1) The file " FigS26A.csv"
 contains the quantum efficiency of PSII under various light intensities
 (from 1500 μE to 25uE) at 400ppm CO2 for the WT and McG plants grown under
 ambient CO2 condition. Number of Variables: 5 Variables: * genotypes and
 replicates: four biological replicates for WT, McG_1, McG_2, McG_3, McG_4
 - Qin Light µmol m⁻² s⁻¹: light intensity (µmol m⁻² s⁻¹) * PhiPSII:
 quantum efficiency of photosystem II - AVG: average of PhiPSII of
 biological replicates * SE: standard error of PhiPSII mean of biological
 replicates (2)The file " FigS26B.csv" contains the quantum
 efficiency of PSII under various light intensities (from 1500 μE to 25uE)
 at 1500ppm CO2 for the WT and McG plants grown under high CO2 (3000ppm)
 condition. Number of Variables: 5 Variables: * genotypes and replicates:
 four biological replicates for WT, McG_1, McG_2, McG_3, McG_4 - Qin Light
 µmol m⁻² s⁻¹: light intensity (µmol m⁻² s⁻¹) * PhiPSII: quantum efficiency
 of photosystem II - AVG: average of PhiPSII of biological replicates * SE:
 standard error of PhiPSII mean of biological replicates **FigS27 folder**:
 (1) The "FigS27AB_photosynthesisHeatmap.csv" contains data for
 generating heatmaps shown in FigS27A and FigS27B by using
 "photosynthesisHeatmapSeparate.R" Variables: Accession: Uniport
 ID protein name group: a: proteins involved in photosystem II; b: proteins
 involved in cytochrome b6/f and electron transfer; c: proteins involved in
 photosystem I; d: proteins involved in F-type ATP synthase; e: proteins
 involved in Calvin-Benson-Bassham (CBB) cycle genotype and replicates:
 four biological replicates for WT, McG_1, McG_2, McG_3, and McG_4 lines.
 (2) The "FigS27C.csv" file contains RuBisCO activity
 measurements of WT and four McG lines grown under ambient CO2 or High CO2
 (3000 ppm) condition. Number of Variables: 5 Variables: * genotype and
 replicates: four biological replicates are used for WT and four McG lines.
 - growth condition: plants were grown under ambient CO2 or High CO2 (3000
 ppm) * Unit mg-1 protein: Unit enzyme activity per mg protein extract. -
 AVG: average unit enzyme activity per mg protein extract of biological
 replicates * SE: standard errors of mean of unit enzyme activity per mg
 protein extract of biological replicates Statistical analysis:
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value. **FigS28
 folder**: (1) The file "FigS28A.csv" contains quantum efficiency
 of PSII  of WT and McG lines under light intensity of 100 µmol m⁻² s⁻¹,
 ambient CO2 (400 ppm). Number of variables: 4 Variables: * genotype and
 replicates: four biological replicates for WT and four McG lines -
 PhiPSII: quantum efficiency of PSII  * Average: average of PhiPSII - SE:
 standard error of PhiPSII mean of biological replicates Statistical
 analysis: Welch's ANOVA test for One-Way-ANOVA test (P-value
 &lt;0.05). Dunnett's T3 multiple comparisons test for
 adjusted-P-value. (2) The file "FIgS28B_LM ppt.ppt" contains
 light micrographs of the WT and McG leaves. (3) The file
 "FigS28C.csv" is used to calculate chloroplast number per
 mesophyll cell of the WT and McG lines. Number of variables: 2 Variables:
 * genotype and replicates: 80 mesophyll cell replicates for WT and four
 McG line - Chloroplast number per mesophyll cell Statistical analysis:
 Welch's ANOVA test for One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value. **FigS29
 folder**: (1) The folders "FigS29A\_ WT",
 "FigS29B_McG_1", "FigS29C_McG*_*2",
 "FigS29D_McG_3", and "FigS29E_McG_4" contain
 transmission electron micrographs of cross-section leaves for WT and each
 McG line accordingly.  (2) The file "FigS29A-E_thylakoid.pptx"
 contains representative images for the WT and McG chloroplast, which are
 used in FigS29A-E. (3) The file "FigS29F.csv" shows the
 thylakoid grana number per chloroplast of WT and four McG lines. Number of
 variables: 4 Variables: * genotypes and their chloroplast replicates: 10
 of chloroplast replicates of WT and four McG lines are used for
 calculating the thylakoid grana number per chloroplast - Numbers:
 thylakoid grana number in a chloroplast * AVG: average thylakoid grana
 number of chloroplast replicates - SE: standard error of mean of thylakoid
 grana number Statistical analysis: Welch's ANOVA test for
 One-Way-ANOVA test (P-value &lt;0.05).
 Dunnett's T3 multiple comparisons test for adjusted-P-value. **FigS30
 folder**: (1) The file "MetaboAnalyst_R_PCA.R" and "
 FigS30A_leaf_Conc_PCA.csv" are used to generate the PCA plot (Fig.
 S30A) for visualizing the difference between the WT and McG leaf lipids.
 The FigS30A_leaf_Conc_PCA.csv contains detected lipid concentrations in WT
 and four McG lines. Number of variables:  3 Variables: * metabolite name:
 lipid name and its chain length - concentrations (𝜇g g-1FW) of each lipid
 in leaves * genotypes and replicates: six biological replicates for WT,
 McG_2, McG_3, and McG_4 lines. Five replicates for McG_1 line. (2) The
 file "HCL.R" is used to generate the "FigS30B_Statistical
 Analysis_Conc_Heatmap.csv" and the heatmap shown in Fig. S30B.
 FigS30B_Statistical Analysis_Conc_Heatmap.csv contains log2-transformed
 ratio to the WT average of each lipid Number of variables:  3 Variables: *
 genotypes and replicates: six biological replicates for WT, McG_2, McG_3,
 and McG_4 lines. Five replicates for McG_1 line. - metabolite name: lipid
 name and its chain length * log2-transformed ratio to the WT average of
 each lipid (4) The file
 "FigS30AB_20240305_analysis_PCA_heatmap.ppt" contains the PCA
 and the heatmap used in FigS30AB. **FigS31 folder**: (1) The
 file "MetaboAnalyst_R_PCA" and
 "FigS31A_Seed_Conc_PCA.csv" are used to generate the PCA plot in
 FigS31A. The"FigS31A_Seed_Conc_PCA.csv" contains detected lipid
 concentrations in WT and four McG seeds. Number of variables:  3
 Variables: * metabolite name: lipid name and its chain length -
 concentrations (𝜇g g-1FW) of each lipid in seeds * genotypes and
 replicates: six biological replicates for WT, McG_1, McG_2, McG_3, and
 McG_4 lines. (2) The file "FigS31B_Statistical
 Analysis_Seed.csv" and "HCL.R" are used to generate the
 heatmap in FigS31B. The file "FigS31B_Statistical
 Analysis_Seed.csv" contains log2-transformed ratio to the WT average
 of each lipid in seeds Number of variables:  3 Variables: * genotypes and
 replicates: six biological replicates for WT, McG_1, McG_2, McG_3, and
 McG_4 line - metabolite name: lipid name and its chain length *
 log2-transformed ratio to the WT average of each lipid (3) The file
 "FigS31AB_20240305_analysis_PCA_heatmap.ppt" contain figures
 transferred from the file "FigS31B_Statistical Analysis_Seed"
 used for FigS31A-B. **FigS32 folder**:  (1) In folder
 "FigS32A_Leaf_Conc_chainlength",  the FigS32A_leaf_Conc.csv and
 LipidFAD.R are used for generating the .svg files showing concentrations
 of chain lengths of each lipid shown in Fig.S32A. The
 "FigS32A_leaf_Conc.csv" contains detected lipid concentrations
 (𝜇g g-1FW) in leaves of WT and four McG lines. Number of variables:  2
 Variables: * metabolite name: lipid name and its chain length -
 concentrations (𝜇g g-1FW) of each lipid in leaves (2) In folder
 "FigS32B_Seed_Conc_chainlength",
 "FigS32B_Seed_Conc.csv" and LipidFAD.R are used for generating
 the .svg files showing concentrations of chain lengths of each lipid shown
 in Fig.S32B. The  "FigS32B_Seed_Conc.csv" contains detected
 lipid concentrations (𝜇g g-1FW) in seeds of WT and four McG lines. Number
 of variables:  2 Variables: * metabolite name: lipid name and its chain
 length - concentrations (𝜇g g-1FW) of each lipid in seeds.
 **TableS1_energy cost of CBB MCG.csv**: This file is used for Table
 S1"Comparison of carbon conversion into acetyl-coA synthesis between
 the native CBB-PDH pathway and CBB-McG dual-cycle" Variables: CBB +
 PDH: Calvin-Benson-Bassham cycle and pyruvate dehydrogenase pathway CBB +
 McG: Calvin-Benson-Bassham cycle with McG cycle NAD(P)H consumption:
 number of NADPH molecules is required for one acetyl-coA synthesis. ATP
 consumption: number of ATP molecules are required for one acetyl-coA
 synthesis. RubisCo turnover per C2: number of RubisCo carboxylation
 reaction is required for one acetyl-coA synthesis The theoretical carbon
 yield per C3: carbon use efficiency of a C3 for an acetyl-coA formation
 **TableS2_folder:** This folder contains auxin (Indole-3-acetic acid),
 cytokinin (trans-zeatin), gibberellins (GA), and brassinosteroids (BR)
 measurements of WT and McG plants. It is used to generate Table S2
 "Phytohormone measurements". (1) "TableS2_BR.csv" file
 contains BR standard curve and for measuring brassinosteroids (BR) in WT
 and McG plant leaves.  Number of variables: 6 Variables:  * standard BR
 concentrations: 50ul solutions of 0, 15.6, 31.25, 62.5, 125, 250, and 500
 pmole/L BR - OD450: absorption at 450nm * genotypes and their biological
 replicates: four biological replicates for each genotype - BR pg g-1FW:
 concentrations of BR in WT and McG plant leaves * AVG: average BR
 concentration of replicates - SE: standard error of mean of BR
 concentrations of replicates (2) "TableS2.csv" file contains GA
 measurements of WT and McG plant leaves. Variables:  * standard GA
 concentrations: 50ul solutions of 0, 15.6, 31.25, 62.5, 125, 250, and 500
 pmole/L BR - OD450: absorption at 450nm * genotypes and their biological
 replicates: four biological replicates for each genotype - GA pg g-1FW:
 concentrations of GA in WT and McG plant leaves * AVG: average GA
 concentration of replicates - SE: standard error of mean of GA
 concentrations of replicates (3)"TableS2_IAA.csv" file
 contains Indole-3-acetic acid (IAA) measurements of WT and McG plant
 leaves Number of Variables: Variables: 11 * genotype: WT and four McG
 lines - bio-rep samples: biological replicates for WT and each line *
 tech-rep: 3 technical replicate measurements are performed for each
 biological replicate - Area of IAA: detected peak area of IAA in samples *
 Area of 13C6-IAA: detected peak area of spiked-in standard13C6-IAA
 (10ng/10mL extract solution) - IAA/13C6-IAA ratio: peak area ratio
 of IAA/13C6-IAA * mean: mean of peak area ratio of IAA/13C6-IAA of three
 technical replicates - 13C6-IAA (ng): concentration of 13C6-IAA in sample
 extract solution * FW (g): sample fresh weight (g) - ng g-1 IAA:
 concentration of IAA of each sample * mean: average IAA concentration of 6
 biological replicates (4)"TableS2_trans_Zeatin.csv" file
 contains Cytokinin (trans-Zeatin) concentrations in WT and McG plant
 leaves Number of variables:12 Variables: * genotype: WT and four McG lines
 - bio-rep: 6 biological replicates for WT and each McG line * tech-rep: 3
 technical replicate measurements are performed for each biological
 replicate - FW g: fresh weight (g) of each biological replicate * ml
 extract bf: milliliter (ml) extraction buffer used for each biological
 replicate - ng d5-tz: nanogram (ng) of d5-trans-Zeatin spiked-in ml
 extract bf * d5-tZ (s) Area: peak area of spiked in d5-tZ in each sample -
 tZ (s) Area: peak area of trans-Zeatin  (tZ) in each sample * tZ (s) ng
 g-1 FW: concentration of trans-Zeatin (ng g-1 FW) of each sample - tech
 replicatesAVG: average tZ concentration of three technical replicate for
 each biological replicate * bio replicate AVG: average tZ concentration of
 six biological replicates - bio replicate SE: standard error of  mean of
 tZ concentration of six biological replicates **TableS3.csv**: This file
 contains chlorophyll contents of WT and McG rosette leaves. Number of
 variables:14 Variables: * genotype: WT and four McG lines - bio-rep: 6
 biological replicates * OD663: absorption at 663nm - OD646: absorption at
 646nm * OD645:absorption at 645nm - Chla ug g-1FW: chlorophyll a content
 of each biological replicate * Chlb ug g-1FW: chlorophyll b content of
 each biological replicate - Total chl ug g-1FW: total chlorophyll content
 of each biological replicate * AVG Chla: average chlorophyll a content -
 SE Chla: standard error of mean of chlorophyll a content * AVG Chlb:
 average chlorophyll b content - SE Chlb: standard error of mean of
 chlorophyll b content * AVG total Chl: average of total chlorophyll
 content - SE total Chl: standard error of mean of  total chlorophyll
 content **TableS4 folder** This folder contains measurements for quantify
 total protein content and carbohydrate contents of the WT and McG leaves.
 (1) TableS4_gluc.csv contains glucose contents for WT and McG lines Number
 of variables: Variables: 7 * genotype: WT and four McG lines - bio-rep:
 four biological replicates for WT and each line * FW g: leaf fresh weight
 (g) - Glu Area: detected peak area of glucose on HPCL * STD mM: 10ul of
 10-400 mM glucose solution is used for generating standard curve for
 calculating glucose concentrations in WT and each McG plant samples - mM
 in 10ul: Glucose (mM) in 10ul injected sample solution * mg/g: Glucose
 content (mg g-1FW) - AVG: average glucose content of 4 biological
 replicates * SE: standard error of mean of glucose content of 4 biological
 replicates  (2) TableS4_sucrose.csv contains sucrose contents of WT and
 McG lines Variables: 7 * genotype: WT and four McG lines - bio-rep: four
 biological replicates for WT and each line * FW g: leaf fresh weight (g) -
 Suc Area: detected peak area of sucrose on HPCL * STD mM: 10ul of 10-400
 mM sucrose solution is used for generating standard curve for calculating
 glucose concentrations in WT and each McG plant samples - mM in 10ul:
 Sucrose (mM) in 10ul injected sample solution * mg/g: Sucrose content (mg
 g-1FW) - AVG: average sucrose content of 4 biological replicates * SE:
 standard error of mean of sucrose content of 4 biological replicates  (3)
 TableS4_starch.csv contains starch measurements for WT and McG lines
 Variables: 7 * genotype: WT and four McG lines - bio-rep: four biological
 replicates for WT and each line * FW g: leaf fresh weight (g) -
 deltaOD340: measured absorbance at 340nm - blank absorbance at 340nm *
 mg/g: Starch content (mg g-1FW) - AVG: average starch content of 4
 biological replicates * SE: standard error of mean of starch content of 4
 biological replicates  (4) TableS4_water_soluble_glucan.csv contains
 soluble glucan measurements for WT and McG lines. Variables: 7 * genotype:
 WT and four McG lines - bio-rep: four biological replicates for WT and
 each line * FW g: leaf fresh weight (g) - deltaOD340: measured absorbance
 at 340nm - blank absorbance at 340nm * mg/g: water soluble glucan content
 (mg g-1FW) - AVG: average of water soluble glucan content of 4 biological
 replicates * SE: standard error of mean of water soluble glucan content of
 4 biological replicates (5) TableS4_protein_BCA.csv file contains protein
 contents of WT and McG plant leaves. Variables: 7 * genotype: WT and four
 McG lines - bio-rep: four biological replicates for WT and each line * FW
 g: leaf fresh weight (g) - OD562: absorbance at 562nm * mg/g: protein
 content (mg) g-1FW - AVG: average protein content of 4 biological
 replicates * SE: standard error of mean of protein content of 4 biological
 replicates **TableS5.csv**: (1)The file contains concentration of each
 lipid class in the WT and McG plant leaves. Number of variables: 6
 Variables: * lipid class: different lipid class - number: number of lipids
 in each class * genotype and replicates: six biological replicates for WT,
 McG_2, McG_3, and McG_4.  Five biological replicates for McG_1 -
 concentration of lipid shown in each cell: ug g-1FW * AVG: average lipid
 content of biological replicates - SE: standard error of mean of lipid
 contents of biological replicates **TableS6.csv**: This file contains
 contents of different lipid classes in WT and McG plant seeds. Number of
 variables: 6 Variables: * lipid class: different lipid class - number:
 number of lipids in each class * genotype and replicates: six biological
 replicates for WT, McG_1, McG_2, McG_3, and McG_4. - concentration of
 lipid shown in each cell: ug g-1FW * AVG: average lipid content of
 biological replicates - SE: standard error of mean of lipid contents of
 biological replicates **TableS7.csv**: This file contains sequences of the
 primers and probes for detecting transcriptional levels of McG transgenes
 in the McG lines. Number of variables: 5 Variables: * probe dye:
 fluorescence dye for measuring expression levels of transgenes (PPC, MTK,
 MCL, GDH, GCL, TSR) and Arabidopsis endogenous GBSS gene - Gene name *
 Forward: sequence of forward primer - Probe: sequence of probe * Reverse:
 sequence of reverse primer ## Code/software (1) Biorender is used to
 create Fig. 1. (2) Geneious Prim is used to analyze whole genome sequence
 data (.fasta) of the WT and McG lines to identify the insertion sites of
 the McG transgenes in the genome. (3) Excel and R Studio are used to
 analyze raw data and generate figures and tables. The scripts are included
 in the R files, which are described above.   (4) Prism version 10.4.1 is
 used for One-way Welch's ANOVA statistical analyses (p-value&lt;0.05)
 and Dunnett's T3 multiple comparison tests. ## Access information
 Other publicly accessible locations of the data: * NA Data was derived
 from the following sources: * NA