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Stem Vacuole-targeted Sucrose Isomerase Enhances Sugar Accumulation in Sorghum
Guoquan Liu
Centre for Crop Science, Queensland Alliance for Agriculture and Food Innovation, The University of Queensland;
Corresponding Author
ORCiD: https://orcid.org/0000-0001-5979-785X
License:
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Background: Sugar accumulation is critically important in determining sugar crop productivity. However, improvement in sugar content has been stagnant among sugar crops for decades. Sorghum, especially sweet sorghum with high biomass, has shown great potential for biofuel. In this study, sorghum was investigated as a C4 diploid model for crops with more complicated genomes such as maize and sugarcane. To enhance sugar accumulation, the sucrose isomerase (SI) gene, driven by stem-specific promoters (A2 or LSG) with a vacuole-targeted signal peptide, was transformed into the sorghum inbred line (Tx430).
Results: The study demonstrated that transgenic lines of grain sorghum, containing 50-60% isomaltulose, accumulated sevenfold (804 mM) more total sugar than the control Tx430 did (118 mM) in stalks. Subsequently, the elite engineered lines (A5, and LSG9) were crossed with sweet sorghum (R9188, and Rio). Total sugar contents (over 750 mM), were significantly higher in F1, and F2 progenies than the control Rio (480 mM). The sugar contents of the engineered lines (over 750 mM), including T0, T1, F1, and F2, are higher than that of the field-grown sugarcane (normal range 600-700 mmol/L). Additionally, physiological characterization demonstrated that the superior progenies had notably higher rates of photosynthesis, sucrose transport, and sink strength than the controls.
Conclusions: The genetic engineering approach has significantly enhanced total sugar content in grain sorghum (T0, and T1) and hybrid sorghum (F1, and F2), demonstrating that sorghum can accumulate sugar contents as high or higher than sugarcane. This research puts sorghum in the spotlight and frontier as a biofuel crop, particularly as it is a shorter duration crop. The substantial increase in sugar content would lead to enormous financial benefits for industrial utilization. This study could have a substantial impact on renewable bioenergy. More importantly, our results demonstrated that the phenotype of high sugar accumulation is inheritable and shed light on improvement for other sugar crops.
Keywords
Isomaltulose, Sorghum, sucrose isomerase, sugar accumulation, renewable energy, photosynthesis, sugarcane, genetic engineering
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- SupplementaryMaterials.docx
Supplementary materials Fig. S1. Transgenic sorghum lines were grown in a PC2 glasshouse. (a) one week; (b) two weeks; (c) three weeks; (d) five weeks; (e) seven weeks; (f) ten weeks in the glasshouse. Fig. S2. The Construct Used for Gene Transformation. Stem-specific Promoter: either A1 resulting in high gene expression in the mature stalk or LSG2 resulting in high gene expression in the loading sucrose section of the stalk. Vacuole leading: encoding a propeptide to guide SI gene products to vacuole where sucrose accumulates. Non-silence target gene: sucrose isomerase without motifs gene silencing in plants. Multiple terminators: three recombined terminators complex to guarantee the proper termination of gene transcription. Fig. S3 PCR screening of sucrose isomerase gene in transgenic lines. An example of agarose gel running the genomic PCR products to test the sucrose isomerase gene in the transgenic plantlets. M: DNA ladder; NC: negative control; Number 1 to 18 are transgenic samples. Fig. S4 Total sugar content in the T1 lines of L9 . Sugars, including isomaltulose, were measured 20 days post-anthesis in the middle section of internode 4 (counted from top). L9 is one of the top lines in T0 transgenic lines. IL9-2, 9-3, 9-6, 9-7, 9-11, 9-12 are isomaltulose positive samples. Nil-IL9 is the null-segregant in T1 generation. Tx430 is the non-transformed control. Fig. S5 PCR screening of sucrose isomerase gene in hybrid lines (Rio X L9). An example of agarose gel running the genomic PCR products to test the sucrose isomerase gene in hybrid lines. M: DNA ladder Number 1 to 12 are hybrid samples. Table S1 Sugar profile of controls and positive transgenic lines with isomaltulose Table S2 Sugar profile of F1 hybrid lines of R9188 X L9
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This preprint is under consideration at Biotechnology for Biofuels. A preprint is a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Learn more about In Review
Research
Stem Vacuole-targeted Sucrose Isomerase Enhances Sugar Accumulation in Sorghum
Guoquan Liu
Centre for Crop Science, Queensland Alliance for Agriculture and Food Innovation, The University of Queensland;
Corresponding Author
ORCiD: https://orcid.org/0000-0001-5979-785X
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Under Review
Version 1
Posted 30 Dec, 2020
No community comments so far
Reviewer #2 agreed
On 03 Jan, 2021
Reviewer #1 agreed
On 01 Jan, 2021
Reviewers invited
Invitations sent on 31 Dec, 2020
Editor invited
On 22 Dec, 2020
Editor assigned
On 22 Dec, 2020
Submission checks complete
On 22 Dec, 2020
First submitted
On 21 Dec, 2020
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
Subject Areas
Biotechnology and Bioengineering
License:
This work is licensed under a CC BY 4.0 License. Read Full License
Background: Sugar accumulation is critically important in determining sugar crop productivity. However, improvement in sugar content has been stagnant among sugar crops for decades. Sorghum, especially sweet sorghum with high biomass, has shown great potential for biofuel. In this study, sorghum was investigated as a C4 diploid model for crops with more complicated genomes such as maize and sugarcane. To enhance sugar accumulation, the sucrose isomerase (SI) gene, driven by stem-specific promoters (A2 or LSG) with a vacuole-targeted signal peptide, was transformed into the sorghum inbred line (Tx430).
Results: The study demonstrated that transgenic lines of grain sorghum, containing 50-60% isomaltulose, accumulated sevenfold (804 mM) more total sugar than the control Tx430 did (118 mM) in stalks. Subsequently, the elite engineered lines (A5, and LSG9) were crossed with sweet sorghum (R9188, and Rio). Total sugar contents (over 750 mM), were significantly higher in F1, and F2 progenies than the control Rio (480 mM). The sugar contents of the engineered lines (over 750 mM), including T0, T1, F1, and F2, are higher than that of the field-grown sugarcane (normal range 600-700 mmol/L). Additionally, physiological characterization demonstrated that the superior progenies had notably higher rates of photosynthesis, sucrose transport, and sink strength than the controls.
Conclusions: The genetic engineering approach has significantly enhanced total sugar content in grain sorghum (T0, and T1) and hybrid sorghum (F1, and F2), demonstrating that sorghum can accumulate sugar contents as high or higher than sugarcane. This research puts sorghum in the spotlight and frontier as a biofuel crop, particularly as it is a shorter duration crop. The substantial increase in sugar content would lead to enormous financial benefits for industrial utilization. This study could have a substantial impact on renewable bioenergy. More importantly, our results demonstrated that the phenotype of high sugar accumulation is inheritable and shed light on improvement for other sugar crops.
Figure 1
Figure 2
Figure 3
Figure 4
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Figure 9