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CRISPR/Cas9-mediated targeted mutagenesis of GmLHY genes alters plant height and internode length in soybean
BACKGROUND: Soybean (Glycine max) is an economically important oil and protein crop. Plant height is a key trait that significantly impacts the yield of soybean; however, research on the molecular mechanisms associated with soybean plant height is lacking. The CRISPR (clustered regularly interspaced...
Autores principales: | , , , , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2019
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6921449/ https://www.ncbi.nlm.nih.gov/pubmed/31852439 http://dx.doi.org/10.1186/s12870-019-2145-8 |
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author | Cheng, Qun Dong, Lidong Su, Tong Li, Tingyu Gan, Zhuoran Nan, Haiyang Lu, Sijia Fang, Chao Kong, Lingping Li, Haiyang Hou, Zhihong Kou, Kun Tang, Yang Lin, Xiaoya Zhao, Xiaohui Chen, Liyu Liu, Baohui Kong, Fanjiang |
author_facet | Cheng, Qun Dong, Lidong Su, Tong Li, Tingyu Gan, Zhuoran Nan, Haiyang Lu, Sijia Fang, Chao Kong, Lingping Li, Haiyang Hou, Zhihong Kou, Kun Tang, Yang Lin, Xiaoya Zhao, Xiaohui Chen, Liyu Liu, Baohui Kong, Fanjiang |
author_sort | Cheng, Qun |
collection | PubMed |
description | BACKGROUND: Soybean (Glycine max) is an economically important oil and protein crop. Plant height is a key trait that significantly impacts the yield of soybean; however, research on the molecular mechanisms associated with soybean plant height is lacking. The CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9 (CRISPR-associated system 9) system is a recently developed technology for gene editing that has been utilized to edit the genomes of crop plants. RESULTS: Here, we designed four gRNAs to mutate four LATE ELONGATED HYPOCOTYL (LHY) genes in soybean. In order to test whether the gRNAs could perform properly in transgenic soybean plants, we first tested the CRISPR construct in transgenic soybean hairy roots using Agrobacterium rhizogenes strain K599. Once confirmed, we performed stable soybean transformation and obtained 19 independent transgenic soybean plants. Subsequently, we obtained one T(1) transgene-free homozygous quadruple mutant of GmLHY by self-crossing. The phenotypes of the T(2)-generation transgene-free quadruple mutant plants were observed, and the results showed that the quadruple mutant of GmLHY displayed reduced plant height and shortened internodes. The levels of endogenous gibberellic acid (GA3) in Gmlhy1a1b2a2b was lower than in the wild type (WT), and the shortened internode phenotype could be rescued by treatment with exogenous GA3. In addition, the relative expression levels of GA metabolic pathway genes in the quadruple mutant of GmLHY were significantly decreased in comparison to the WT. These results suggest that GmLHY encodes an MYB transcription factor that affects plant height through mediating the GA pathway in soybean. We also developed genetic markers for identifying mutants for application in breeding studies. CONCLUSIONS: Our results indicate that CRISPR/Cas9-mediated targeted mutagenesis of four GmLHY genes reduces soybean plant height and shortens internodes from 20 to 35 days after emergence (DAE). These findings provide insight into the mechanisms underlying plant height regulatory networks in soybean. |
format | Online Article Text |
id | pubmed-6921449 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2019 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-69214492019-12-30 CRISPR/Cas9-mediated targeted mutagenesis of GmLHY genes alters plant height and internode length in soybean Cheng, Qun Dong, Lidong Su, Tong Li, Tingyu Gan, Zhuoran Nan, Haiyang Lu, Sijia Fang, Chao Kong, Lingping Li, Haiyang Hou, Zhihong Kou, Kun Tang, Yang Lin, Xiaoya Zhao, Xiaohui Chen, Liyu Liu, Baohui Kong, Fanjiang BMC Plant Biol Research Article BACKGROUND: Soybean (Glycine max) is an economically important oil and protein crop. Plant height is a key trait that significantly impacts the yield of soybean; however, research on the molecular mechanisms associated with soybean plant height is lacking. The CRISPR (clustered regularly interspaced short palindromic repeat)/Cas9 (CRISPR-associated system 9) system is a recently developed technology for gene editing that has been utilized to edit the genomes of crop plants. RESULTS: Here, we designed four gRNAs to mutate four LATE ELONGATED HYPOCOTYL (LHY) genes in soybean. In order to test whether the gRNAs could perform properly in transgenic soybean plants, we first tested the CRISPR construct in transgenic soybean hairy roots using Agrobacterium rhizogenes strain K599. Once confirmed, we performed stable soybean transformation and obtained 19 independent transgenic soybean plants. Subsequently, we obtained one T(1) transgene-free homozygous quadruple mutant of GmLHY by self-crossing. The phenotypes of the T(2)-generation transgene-free quadruple mutant plants were observed, and the results showed that the quadruple mutant of GmLHY displayed reduced plant height and shortened internodes. The levels of endogenous gibberellic acid (GA3) in Gmlhy1a1b2a2b was lower than in the wild type (WT), and the shortened internode phenotype could be rescued by treatment with exogenous GA3. In addition, the relative expression levels of GA metabolic pathway genes in the quadruple mutant of GmLHY were significantly decreased in comparison to the WT. These results suggest that GmLHY encodes an MYB transcription factor that affects plant height through mediating the GA pathway in soybean. We also developed genetic markers for identifying mutants for application in breeding studies. CONCLUSIONS: Our results indicate that CRISPR/Cas9-mediated targeted mutagenesis of four GmLHY genes reduces soybean plant height and shortens internodes from 20 to 35 days after emergence (DAE). These findings provide insight into the mechanisms underlying plant height regulatory networks in soybean. BioMed Central 2019-12-18 /pmc/articles/PMC6921449/ /pubmed/31852439 http://dx.doi.org/10.1186/s12870-019-2145-8 Text en © The Author(s). 2019 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Research Article Cheng, Qun Dong, Lidong Su, Tong Li, Tingyu Gan, Zhuoran Nan, Haiyang Lu, Sijia Fang, Chao Kong, Lingping Li, Haiyang Hou, Zhihong Kou, Kun Tang, Yang Lin, Xiaoya Zhao, Xiaohui Chen, Liyu Liu, Baohui Kong, Fanjiang CRISPR/Cas9-mediated targeted mutagenesis of GmLHY genes alters plant height and internode length in soybean |
title | CRISPR/Cas9-mediated targeted mutagenesis of GmLHY genes alters plant height and internode length in soybean |
title_full | CRISPR/Cas9-mediated targeted mutagenesis of GmLHY genes alters plant height and internode length in soybean |
title_fullStr | CRISPR/Cas9-mediated targeted mutagenesis of GmLHY genes alters plant height and internode length in soybean |
title_full_unstemmed | CRISPR/Cas9-mediated targeted mutagenesis of GmLHY genes alters plant height and internode length in soybean |
title_short | CRISPR/Cas9-mediated targeted mutagenesis of GmLHY genes alters plant height and internode length in soybean |
title_sort | crispr/cas9-mediated targeted mutagenesis of gmlhy genes alters plant height and internode length in soybean |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6921449/ https://www.ncbi.nlm.nih.gov/pubmed/31852439 http://dx.doi.org/10.1186/s12870-019-2145-8 |
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