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Characterization of a Novel TtLEA2 Gene From Tritipyrum and Its Transformation in Wheat to Enhance Salt Tolerance.

ABSTRACT: Late embryogenesis-abundant (LEA) proteins are critical in helping plants cope with salt stress. "Y1805" is a salt-tolerant Tritipyrum. We identified a "Y1805"-specific LEA gene that was expressed highly and sensitively under salt stress using transcriptome analysis. The novel group 2 LEA gene (TtLEA2-1) was cloned from "Y1805." TtLEA2-1 contained a 453 bp open reading frame encoding an 151-amino-acid protein that showed maximum sequence identity (77.00%) with Thinopyrum elongatum by phylogenetic analysis. It was mainly found to be expressed highly in the roots by qRT-PCR analysis and was located in the whole cell. Forty-eight candidate proteins believed to interact with TtLEA2-1 were confirmed by yeast two-hybrid analysis. These interacting proteins were mainly enriched in "environmental information processing," "glycan biosynthesis and metabolism," and "carbohydrate metabolism." Protein-protein interaction analysis indicated that the translation-related 40S ribosomal protein SA was the central node. An efficient wheat transformation system has been established. A coleoptile length of 2 cm, an Agrobacteria cell density of 0.55-0.60 OD₆₀₀, and 15 KPa vacuum pressure were ideal for common wheat transformation, with an efficiency of up to 43.15%. Overexpression of TaLEA2-1 in wheat "1718" led to greater height, stronger roots, and higher catalase activity than in wild type seedlings. TaLEA2-1 conferred enhanced salt tolerance in transgenic wheat and may be a valuable gene for genetic modification in crops.

SUBMITTER: Yang Z

PROVIDER: S-EPMC9014267 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

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Characterization of a Novel TtLEA2 Gene From Tritipyrum and Its Transformation in Wheat to Enhance Salt Tolerance.

Yang Zhifen Z Mu Yuanhang Y Wang Yiqin Y He Fang F Shi Luxi L Fang Zhongming Z Zhang Jun J Zhang Qingqin Q Geng Guangdong G Zhang Suqin S

Frontiers in plant science 20220404

Late embryogenesis-abundant (LEA) proteins are critical in helping plants cope with salt stress. "Y1805" is a salt-tolerant Tritipyrum. We identified a "Y1805"-specific LEA gene that was expressed highly and sensitively under salt stress using transcriptome analysis. The novel group 2 LEA gene (TtLEA2-1) was cloned from "Y1805." TtLEA2-1 contained a 453 bp open reading frame encoding an 151-amino-acid protein that showed maximum sequence identity (77.00%) with Thinopyrum ...[more]

PMID: 35444677

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Project description:BackgroundCommon wheat (Triticum aestivum L.) is a worldwide cereal crop, which is an integral part of the diets of many countries. In addition, the MYB gene of wheat plays a role in the response to salt stress.Results"Y1805" is a Tritipyrum variety that is relatively tolerant to salt. We used transcriptome analysis to show that the "Y1805" MYB gene was both highly expressed and sensitive to salt stress. Compared with control roots, the level of MYB expression during salt stress was higher, which rapidly decreased to control levels during the recovery process. MYB gene relative expression showed the highest levels in "Y1805" roots during salt stress, with the stems and then leaves being the next highest stressed tissues. The novel MYB gene (TtMYB1) was successfully cloned from "Y1805". It showed a coding sequence length of 783 bp with 95.79% homology with Tel2E01G633100 from Thinopyrum elongatum. TtMYB1 and MYB from Th. elongatum were clustered in the same branch using phylogenetic analysis, which indicated high similarities. The TtMYB1 gene is located in the nucleus. The coleoptile method was employed when a TtMYB1 overexpression vector was used during transformation into "1718" (common wheat). Under high salt stress, TtMYB1 leaves of overexpression lines had decreased wilting, when compared with wild-type (WT) plants. During normal conditions, salt stress, and recovery, the lengths of the roots and the heights of seedlings from the overexpression lines were found to be significantly greater than roots and seedlings of WT plants. In addition, during high salt stress, the overexpression lines showed that proline and soluble sugar levels were higher than that of WT plants, but with lower malondialdehyde levels. Forty-three proteins that interacted with TtMYB1 were identified using the yeast two-hybrid assay. Protein-protein interaction analyses indicated that most were SANT domain-containing and Wd repeat region domain-containing proteins. Among these proteins, ribosomal proteins were the main node. Abiotic stress-related terms (such as "carbonate dehydratase activity", "protein targeting peroxisomes", and "glutathione peroxidase activity") were enriched in GO analysis. In KEGG analysis, "carbohydrate metabolism", "environmental information processing", "genetic information processing", "signaling and cell precursors", and "energy metabolism" pathways were enriched.ConclusionThe TtMYB1 gene might enhance salt tolerance by increasing proline and soluble sugar content and antioxidase activity in transgenic wheat. It therefore has the potential to enhance high salt tolerance in plants.

Project description:NAC transcription factors are widely distributed in the plant kingdom and play an important role in the response to various abiotic stresses in plant species. Tritipyrum, an octoploid derived from hybridization of Triticum aestivum (AABBDD) and Thinopyrum elongatum (EE), is an important genetic resource for integrating the desirable traits of Th. elongatum into wheat. In this study, we investigated the tissue distribution and expression of Tritipyrum NAC genes in the whole genomes of T. aestivum and Th. elongatum after obtaining their complete genome sequences. Based on phylogenetic relationships, conserved motifs, gene synthesis, evolutionary analysis, and expression patterns, we identified and characterized 732 Tritipyrum NAC genes. These genes were divided into six main groups (A, B, C, D, E, and G) based on phylogenetic relationships and evolutionary studies, with members of these groups sharing the same motif composition. The 732 TtNAC genes are widely distributed across 28 chromosomes and include 110 duplicated genes. Gene synthesis analysis indicated that the NAC gene family may have a common ancestor. Transcriptome data and quantitative polymerase chain reaction (qPCR) expression profiles showed 68 TtNAC genes to be highly expressed in response to various salt stress and recovery treatments. Tel3E01T644900 (TtNAC477) was particularly sensitive to salt stress and belongs to the same clade as the salt tolerance genes ANAC019 and ANAC055 in Arabidopsis. Pearson correlation analysis identified 751 genes that correlated positively with expression of TtNAC477, and these genes are enriched in metabolic activities, cellular processes, stimulus responses, and biological regulation. TtNAC477 was found to be highly expressed in roots, stems, and leaves in response to salt stress, as confirmed by real-time PCR. These findings suggest that TtNAC477 is associated with salt tolerance in plants and might serve as a valuable exogenous gene for enhancing salt tolerance in wheat.

Project description:Soybean is one of the most important food crops, breeding salt-tolerant soybean varieties is of great significance to alleviate soybean shortage. In this study, the F-box protein family homologous gene GmFBX322 was cloned from the soybean variety Williams 82 and overexpressed in the Shennong 9 soybean variety to further study and explore the physiological mechanism of soybean salt tolerance. GmFBX322 was constructed on the vector pTF101:35S, and integrated into the genome of Shennong 9 soybean variety by Agrobacterium EHA101-mediated cotyledonary node transformation technology, and 4 overexpressed transgenic lines were obtained, molecular assays were performed on the transformed plants. The expression of GmFBX322 was detected by qRT-PCR and it was found that the leaves of the 4 transgenic lines increased by 2.49, 2.46, 2.77, 2.95 times compared with the wild type; after salt treatment for 12 hours, it was found that the expression of wild type Shennong 9 Inducible expression of GmFBX322. After 72 hours of salt treatment, the leaves of wild-type Shennong 9 soybean plants showed obvious wilting and chlorosis, while the leaves of GmFBX322 plants overexpressing GmFBX322 showed no obvious changes. The leaves were taken at 0, 6, 12, 24, and 48 hours of salt stress to determine the antioxidant activity. Ability and osmotic adjustment level, etc. The results showed that the catalase activity in the leaves of the transgenic lines 2265, 2267, 2269, and 2271 was 2.47, 2.53, 3.59, 2.96 times that of the wild-type plant after 48 hours of salt treatment; the soluble sugar content was 1.22, 1.14, and 1.22 of the wild-type plant. 1.14, 1.57 times; the proline content is 2.20, 1.83, 1.65, 1.84 times of the wild type. After comparing the physiological indicators determined by the experiment, the transgenic lines performed better than the control group, indicating that overexpression of GmFBX322 can enhance the salt tolerance of soybean plants. To verify the function of GmFBX322 gene related to stress resistance, add it to the candidate gene of stress resistance, and provide scientific basis for the selection and breeding of salt-tolerant varieties.

Project description:IntroductionThe transcription factor WRKY is widespread in the plant kingdom and plays a crucial role in diverse abiotic stress responses in plant species. Tritipyrum, an octoploid derived from an intergeneric cross between Triticum aestivum (AABBDD) and Thinopyrum elongatum (EE), is a valuable germplasm resource for introducing superior traits of Th. elongatum into T. aestivum. The recent release of the complete genome sequences of T. aestivum and Th. elongatum enabled us to investigate the organization and expression profiling of Tritipyrum WRKY genes across the entire genome.ResultsIn this study, 346 WRKY genes, from TtWRKY1 to TtWRKY346, were identified in Tritipyrum. The phylogenetic analysis grouped these genes into three subfamilies (I-III), and members of the same subfamilies shared a conserved motif composition. The 346 TtWRKY genes were dispersed unevenly across 28 chromosomes, with 218 duplicates. Analysis of synteny suggests that the WRKY gene family may have a common ancestor. Expression profiles derived from transcriptome data and qPCR demonstrated that 54 TtWRKY genes exhibited relatively high levels of expression across various salt stresses and recovery treatments. Tel1E01T143800 (TtWRKY256) is extremely sensitive to salt stress and is on the same evolutionary branch as the salt-tolerant A. thaliana genes AtWRKY25 and AtWRKY33. From 'Y1805', the novel AtWRKY25 was cloned. The Pearson correlation analysis identified 181 genes that were positively correlated (R>0.9) with the expression of TtWRKY256, and these genes were mainly enriched in metabolic processes, cellular processes, response to stimulus, biological regulation, and regulation of biological. Subcellular localization and qRT-PCR analysis revealed that TtWRKY256 was located in the nucleus and was highly expressed in roots, stems, and leaves under salt stress.DiscussionThe above results suggest that TtWRKY256 may be associated with salt stress tolerance in plants and may be a valuable alien gene for improving salt tolerance in wheat.

Dataset Information

Characterization of a Novel TtLEA2 Gene From Tritipyrum and Its Transformation in Wheat to Enhance Salt Tolerance.

Publications

Characterization of a Novel <i>TtLEA2</i> Gene From <i>Tritipyrum</i> and Its Transformation in Wheat to Enhance Salt Tolerance.

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