Abstract
BACKGROUND AND AIMS: Vitamin E (tocochromanol) is a lipid-soluble antioxidant and an essential nutrient for human health. Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E, which includes both tocopherols and tocotrienols. Although the vitamin E biosynthetic pathway has been characterized in dicots, such as Arabidopsis, which only accumulate tocopherols, knowledge regarding vitamin E biosynthesis in monocots is limited because of the lack of functional mutants. This study aimed to obtain gene knockout mutants to elucidate the genetic control of vitamin E composition in barley. METHODS: Targeted knockout mutations of HvHPT and HvHGGT in barley were created with CRISPR/Cas9-enabled genome editing. High-performance liquid chromatography (HPLC) was performed to analyse the content of tocochromanol isomers in transgene-free homozygous Hvhpt and Hvhggt mutants. KEY RESULTS: Mutagenesis efficiency among T0 regenerated plantlets was 50-65 % as a result of two simultaneously expressed guide RNAs targeting each gene; most of the mutations were stably inherited by the next generation. The transgene-free homozygous mutants of Hvhpt and Hvhggt exhibited decreased grain size and weight, and the HvHGGT mutation led to a shrunken phenotype and significantly lower total starch content in grains. HPLC analysis revealed that targeted mutation of HvHPT significantly reduced the content of both tocopherols and tocotrienols, whereas mutations in HvHGGT completely blocked tocotrienol biosynthesis in barley grains. Transient overexpression of an HvHPT homologue in tobacco leaves significantly increased the production of γ- and δ-tocopherols, which may partly explain why targeted mutation of HvHPT in barley grains did not eliminate tocopherol production. CONCLUSIONS: Our results functionally validated that HvHGGT is the only committed gene for the production of tocotrienols, whereas HvHPT is partly responsible for tocopherol biosynthesis in barley.
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Claims 11
Claim · model-written summary
Transgene-free homozygous Hvhpt and Hvhggt mutants had decreased grain size and weight.
- Taxon
- Hordeum vulgare
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Hordeum vulgaree0 - Material
- transgene-free homozygous mutants of Hvhpt and Hvhggte1
- Organ / stage
- graine1
- Treatment
- Targeted knockout mutationse2
- Gene
- Hvhpte1
- Outcome
- grain size and weighte1
- Direction
- decreasede1
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
.jpg?width=500)
Verbatim
e0“Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E”
e1“The transgene-free homozygous mutants of Hvhpt and Hvhggt exhibited decreased grain size and weight”
e2“Targeted knockout mutations of HvHPT and HvHGGT in barley were created with CRISPR/Cas9-enabled genome editing”
Claim · model-written summary
The HvHGGT mutation significantly lowered total starch content in grains.
- Taxon
- Hordeum vulgareHordeum vulgaree0
- Organ / stage
- grainse2
- Treatment
- the HvHGGT mutatione2
- Gene
- HvHGGTe2
- Outcome
- total starch content in grainse2
- Direction
- lowere2
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E”
e2“the HvHGGT mutation led to a shrunken phenotype and significantly lower total starch content in grains”
Claim · model-written summary
The HvHGGT mutation led to a shrunken grain phenotype.
- Taxon
- Hordeum vulgareHordeum vulgaree0
- Treatment
- the HvHGGT mutatione2
- Gene
- HvHGGTe2
- Outcome
- shrunken phenotypee2
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E”
e2“the HvHGGT mutation led to a shrunken phenotype and significantly lower total starch content in grains”
Claim · model-written summary
Targeted mutation of HvHPT significantly reduced the content of both tocopherols and tocotrienols in barley grains.
- Taxon
- Hordeum vulgareHordeum vulgaree0
- Treatment
- targeted mutation of HvHPTe3
- Gene
- HvHPTe3
- Outcome
- content of both tocopherols and tocotrienolse3
- Direction
- reducede3
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E”
e3“HPLC analysis revealed that targeted mutation of HvHPT significantly reduced the content of both tocopherols and tocotrienols”
Claim · model-written summary
Mutations in HvHGGT completely blocked tocotrienol biosynthesis in barley grains.
- Taxon
- Hordeum vulgareHordeum vulgaree0
- Organ / stage
- barley grainse4
- Treatment
- mutations in HvHGGTe4
- Gene
- HvHGGTe4
- Outcome
- tocotrienol biosynthesise4
- Direction
- blockede4
- Qualifier
- completelye4
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E”
e4“mutations in HvHGGT completely blocked tocotrienol biosynthesis in barley grains”
Claim · model-written summary
Transient overexpression of an HvHPT homologue significantly increased the production of γ- and δ-tocopherols in tobacco leaves.
- Taxon
- tobacco
Nicotiana tabacume5 - Organ / stage
- leavese5
- Treatment
- Transient overexpression of an HvHPT homologuee5
- Gene
- HvHPT homologuee5
- Outcome
- production of γ- and δ-tocopherolse5
- Direction
- increasede5
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e5“Transient overexpression of an HvHPT homologue in tobacco leaves significantly increased the production of γ- and δ-tocopherols”
Claim · model-written summary
HvHPT may partly explain why its targeted mutation in barley grains did not eliminate tocopherol production.
- Taxon
- barleyHordeum vulgaree6
- Gene
- HvHPTe6
- Outcome
- tocopherol productione6
- Qualifier
- may partly explaine6
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E”
e6“which may partly explain why targeted mutation of HvHPT in barley grains did not eliminate tocopherol production”
Claim · model-written summary
HvHGGT is the only committed gene for tocotrienol production in barley.
- Taxon
- Hordeum vulgareHordeum vulgaree0
- Gene
- HvHGGTe7
- Outcome
- production of tocotrienolse7
- Qualifier
- only committed genee7
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E”
e7“HvHGGT is the only committed gene for the production of tocotrienols”
Claim · model-written summary
HvHPT is partly responsible for tocopherol biosynthesis in barley.
- Taxon
- Hordeum vulgareHordeum vulgaree0
- Gene
- HvHPTe1
- Outcome
- tocopherol biosynthesise1
- Qualifier
- partly responsiblee1
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E”
e1“HvHPT is partly responsible for tocopherol biosynthesis in barley”
Claim · model-written summary
Mutagenesis efficiency among T0 regenerated plantlets was 50-65 %.
- Taxon
- Hordeum vulgareHordeum vulgaree0
- Material
- T0 regenerated plantletse1
- Outcome
- Mutagenesis efficiencye1
- Value
- 50-65 %e1
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E”
e1“Mutagenesis efficiency among T0 regenerated plantlets was 50-65 %”
Claim · model-written summary
Most of the induced mutations were stably inherited by the next generation.
- Taxon
- Hordeum vulgareHordeum vulgaree0
- Outcome
- stably inheritede1
- Qualifier
- by the next generatione1
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“Among cereal crops, barley (Hordeum vulgare) contains a high level of vitamin E”
e1“most of the mutations were stably inherited by the next generation”
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See all 14 papers →Claims are grouped by organism, treatment and what was measured or targeted. A paper counts as positive when any of its claims in the group is positive.