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Mutation of GmIPK1 Gene Using CRISPR/Cas9 Reduced Phytic Acid Content in Soybean Seeds

Authors :
Ji Hyeon Song
Gilok Shin
Hye Jeong Kim
Saet Buyl Lee
Ju Yeon Moon
Jae Cheol Jeong
Hong-Kyu Choi
In Ah Kim
Hyeon Jin Song
Cha Young Kim
Young-Soo Chung
Source :
International Journal of Molecular Sciences; Volume 23; Issue 18; Pages: 10583
Publication Year :
2022
Publisher :
Multidisciplinary Digital Publishing Institute, 2022.

Abstract

Phytic acid (PA) acts as an antinutrient substance in cereal grains, disturbing the bioavailability of micronutrients, such as iron and zinc, in humans, causing malnutrition. GmIPK1 encodes the inositol 1,3,4,5,6-pentakisphosphate 2-kinase enzyme, which converts myo-inopsitol-1,3,4,5,6-pentakisphosphate (IP5) to myo-inositol-1,2,3,4,5,6-hexakisphosphate (IP6) in soybean (Glycine max L.). In this study, for developing soybean with low PA levels, we attempted to edit the GmIPK1 gene using the CRISPR/Cas9 system to introduce mutations into the GmIPK1 gene with guide RNAs in soybean (cv. Kwangankong). The GmIPK1 gene was disrupted using the CRISPR/Cas9 system, with sgRNA-1 and sgRNA-4 targeting the second and third exon, respectively. Several soybean Gmipk1 gene-edited lines were obtained in the T0 generation at editing frequencies of 0.1–84.3%. Sequencing analysis revealed various indel patterns with the deletion of 1–9 nucleotides and insertions of 1 nucleotide in several soybean lines (T0). Finally, we confirmed two sgRNA-4 Gmipk1 gene-edited homozygote soybean T1 plants (line #21-2: 5 bp deletion; line #21-3: 1 bp insertion) by PPT leaf coating assay and PCR analysis. Analysis of soybean Gmipk1 gene-edited lines indicated a reduction in PA content in soybean T2 seeds but did not show any defects in plant growth and seed development.

Details

Language :
English
ISSN :
14220067
Database :
OpenAIRE
Journal :
International Journal of Molecular Sciences; Volume 23; Issue 18; Pages: 10583
Accession number :
edsair.doi.dedup.....671b19a47a07e9ec8b9fdafb9672e817
Full Text :
https://doi.org/10.3390/ijms231810583