Abstract
Genome sequencing is important for discovering critical genes in crops and improving crop breeding efficiency. Generally, fresh, young leaves are used for DNA extraction from plants. However, seeds, the storage form, are more efficient because they do not require cultivation and can be ground at room temperature. Yet, only a few DNA extraction kits or methods suitable for seeds have been developed to date. In this study, we introduced an improved (IMP) Boom method that is relatively low-cost, simple to operate, and yields high-quality DNA that can withstand long-read sequencing. The method successfully extracted approximately 8 µg of DNA per gram of seed weight from soybean seeds at an average concentration of 48.3 ng/µL, approximately 40-fold higher than that extracted from seeds using a common extraction method kit. The A260/280 and A260/230 values of the DNA were 1.90 and 2.43, respectively, which exceeded the respective quality thresholds of 1.8 and 2.0. The DNA also had a DNA integrity number value (indicating the degree of DNA degradation) of 8.1, higher than that obtained using the kit and cetyltrimethylammonium bromide methods. Furthermore, the DNA showed a read length N50 of 20.96 kbp and a maximum read length of 127.8 kbp upon long-read sequencing using the Oxford Nanopore sequencer, with both values being higher than those obtained using the other methods. DNA extracted from seeds using the IMP Boom method showed an increase in the percentage of the nuclear genome with a decrease in the relative ratio of chloroplast DNA. These results suggested that the proposed IMP Boom method can extract high-quality and high-concentration DNA that can be used for long-read sequencing, which cannot be achieved from plant seeds using other conventional DNA extraction methods. The IMP Boom method could also be adapted to crop seeds other than soybeans, such as pea, okra, maize, and sunflower. This improved method is expected to improve the efficiency of various crop-breeding operations, including seed variety determination, testing of genetically modified seeds, and marker-assisted selection.
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Claims 10
Claim · model-written summary
The method yielded approximately 8 µg DNA per gram of soybean seed.
- Taxon
- soybean
Glycine maxe0 - Outcome
- DNA per gram of seed weighte0
- Value
- approximately 8 µge0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“approximately 8 µg of DNA per gram of seed weight from soybean seeds”
Claim · model-written summary
The method gave an average soybean seed DNA concentration of 48.3 ng/µL, about 40-fold above a common kit.
- Taxon
- soybeanGlycine maxe1
- Outcome
- concentratione0
- Direction
- highere0
- Value
- 48.3 ng/µLe0
- Qualifier
- approximately 40-folde0
- Comparator
- common extraction method kite0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“at an average concentration of 48.3 ng/µL, approximately 40-fold higher than that extracted from seeds using a common extraction method kit”
e1“soybean seeds”
Claim · model-written summary
The extracted DNA had an A260/280 value of 1.90 above the quality threshold.
- Taxon
- soybeanGlycine maxe1
- Outcome
- A260/280e0
- Direction
- exceedede0
- Value
- 1.90e0
- Qualifier
- 1.8e0
- Comparator
- quality thresholdse0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“The A260/280 and A260/230 values of the DNA were 1.90 and 2.43, respectively, which exceeded the respective quality thresholds of 1.8 and 2.0”
e1“soybean seeds”
Claim · model-written summary
The extracted DNA had an A260/230 value of 2.43 above the quality threshold.
- Taxon
- soybeanGlycine maxe1
- Outcome
- A260/230e0
- Direction
- exceedede0
- Value
- 2.43e0
- Qualifier
- 2.0e0
- Comparator
- quality thresholdse0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“The A260/280 and A260/230 values of the DNA were 1.90 and 2.43, respectively, which exceeded the respective quality thresholds of 1.8 and 2.0”
e1“soybean seeds”
Claim · model-written summary
The DNA integrity number was 8.1, above the kit and CTAB methods.
- Taxon
- soybeanGlycine maxe1
- Outcome
- DNA integrity number valuee0
- Direction
- highere0
- Value
- 8.1e0
- Comparator
- the kit and cetyltrimethylammonium bromide methodse0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“DNA integrity number value (indicating the degree of DNA degradation) of 8.1, higher than that obtained using the kit and cetyltrimethylammonium bromide methods”
e1“soybean seeds”
Claim · model-written summary
Long-read sequencing gave a read length N50 of 20.96 kbp, above other methods.
- Taxon
- soybeanGlycine maxe1
- Outcome
- read length N50e0
- Direction
- highere0
- Value
- 20.96 kbpe0
- Comparator
- the other methodse0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“read length N50 of 20.96 kbp and a maximum read length of 127.8 kbp upon long-read sequencing using the Oxford Nanopore sequencer, with both values being higher than those obtained using the other methods”
e1“soybean seeds”
Claim · model-written summary
Long-read sequencing gave a maximum read length of 127.8 kbp, above other methods.
- Taxon
- soybeanGlycine maxe1
- Outcome
- maximum read lengthe0
- Direction
- highere0
- Value
- 127.8 kbpe0
- Comparator
- the other methodse0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“read length N50 of 20.96 kbp and a maximum read length of 127.8 kbp upon long-read sequencing using the Oxford Nanopore sequencer, with both values being higher than those obtained using the other methods”
e1“soybean seeds”
Claim · model-written summary
The method increased the percentage of nuclear genome reads.
- Taxon
- soybeanGlycine maxe1
- Outcome
- percentage of the nuclear genomee0
- Direction
- increasee0
- Comparator
- seeds using the IMP Boom methode0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“DNA extracted from seeds using the IMP Boom method showed an increase in the percentage of the nuclear genome with a decrease in the relative ratio of chloroplast DNA”
e1“soybean seeds”
Claim · model-written summary
The method decreased the relative ratio of chloroplast DNA.
- Taxon
- soybeanGlycine maxe1
- Outcome
- relative ratio of chloroplast DNAe0
- Direction
- decreasee0
- Comparator
- seeds using the IMP Boom methode0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“DNA extracted from seeds using the IMP Boom method showed an increase in the percentage of the nuclear genome with a decrease in the relative ratio of chloroplast DNA”
e1“soybean seeds”
Claim · model-written summary
The method could also be adapted to pea, okra, maize and sunflower seeds.
- Taxon
- pea
Lathyrus oleraceuse0 - Outcome
- adapted to crop seeds other than soybeanse0
- Comparator
- soybeanse0
Not reportedValue
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
e0“The IMP Boom method could also be adapted to crop seeds other than soybeans, such as pea, okra, maize, and sunflower”