Soybean Gm-SEIPIN2 family gene and application thereof in improving oil content of plant seeds
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A technology of plant seeds, oil content, applied in the field of genetic engineering
Inactive Publication Date: 2020-03-13
JILIN UNIV
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However, there is no functional research on the SEIPIN f
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Embodiment 1
[0034] Embodiment 1, soybean gene Gm-SEIPIN2A and Gm-SEIPIN2B gene phylogenetic tree
[0035] For analysis, input the amino acid sequences of soybean Gm-SEIPIN2A and Gm-SEIPIN2B genes into NCBI for blast, and download the sequences with high amino acid similarity The amino acid sequence of the At-SEIPIN gene was downloaded, and the phylogenetic tree analysis was performed with MEGA7 software. Analysis results (see figure 1 ) shows that Arabidopsis has one SEIPIN1 gene compared to two Gm-SEIPIN2 genes in soybean, but this phenomenon also exists in other crops such as plants such as alfalfa, peanut, pepper, etc.
Embodiment 2
[0036] Example 2, the expression results of Gm-SEIPIN2A and Gm-SEIPIN2B genes in soybean different tissues
[0037] Roots, stems, leaves, flowers and developing seeds were harvested from two soybeans "Williams82" grown in the same natural environment in Jilin University, Changchun City. The above material was collected at various times and immediately frozen in liquid nitrogen and stored at -80°C after processing for RNA extraction.
[0038] The total RNA of the processed material was extracted with reference to the column type plant total RNA extraction and purification kit of Sangon Company, and the integrity of the RNA was detected by 1% agarose electrophoresis. cDNA was synthesized according to Reverse Transcriptase M-MLV (RNase H - ) instructions. Real-time fluorescent quantitative PCR was used to detect the expression of Gm-SEIPIN2A and Gm-SEIPIN2B genes in different soybean tissues and different embryo developmental stages. The experimental operation was carried out ...
Embodiment 3
[0047] Embodiment 3, the cloning of soybean gene Gm-SEIPIN2A and Gm-SEIPIN2B
[0048] Select 3 soybean seeds with full grains, and sow the seeds in the flower soil for potting. The culture conditions are: 16 hours of light, temperature of 26°C, humidity of 65%, light intensity of 30000 lux. Sampling soybean seeds 25 days after flowering, extracting total RNA, reverse transcription into cDNA, designing primers according to the gene sequence published on soybase, the primers are:
[0049]
[0050] Add reaction components according to Table 1, and perform PCR according to the procedure in Table 2:
[0051] Table 1 PCR reaction system
[0052]
[0053] Table 2 PCR program
[0054]
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Abstract
The invention relates to a soybean Gm-SEIPIN2 family gene and application of the soybean Gm-SEIPIN2 family gene in increasing the oil content of plant seeds, belonging to the technical field of gene engineering. The soybean Gm-SEIPIN2 family gene comprises two genes, namely, a Gm-SEIPIN2A gene and a Gm-SEIPIN2B gene. The soybean Gm-SEIPIN2A gene and the soybean Gm-SEIPIN2B gene of soybean Gm-SEIPIN2 family gene are subjected to functional verification by using a yeast mutant transformation technology and a transgenic Arabidopsis thaliana technology; the results of oil content determination ofand fatty acid ratio determination through a gas chromatography technology prove that the Gm-SEIPIN2A gene and the Gm-SEIPIN2B gene can improve the oil content to different degrees; the results of determination of fatty acid ratios in yeasts show that the above two genes can improve the content of oleic acid; and Arabidopsis oil content determination results show that the above two genes can obviously improve the oil content of Arabidopsis seeds and obviously reduce the content of palmitic acid. The two genes of the Gm-SEIPIN2 family are of great significance in increasing the oil content of plant seeds and increasing the oleic acid content, especially in cultivating high-oil and high-oleic-acid soybean varieties.
Description
technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to soybean Gm-SEIPIN2 family genes and applications thereof. Background technique [0002] Soybean (Glycine max (Linn.) Merr.) is widely grown worldwide and is an important legume crop that provides dietary protein and oils for humans and livestock. In 2017, soybean production accounted for 59% of oilseed production and 29% of vegetable oil consumption, making it the most important oilseed crop in the world. Soybean oil mainly contains five fatty acids, followed by palmitic acid (16:0, PA), stearic acid (18:0, ST), oleic acid (18:0, ST), linoleic acid (18:1, OL ), linolenic acid (18:3, LN). Among them, OL, LI, and LN are unsaturated fatty acids, which can prevent cardiovascular and cerebrovascular diseases and some cancers. However, polyunsaturated fatty acids are very unstable and easily oxidized, and are easily hydrogenated to produce trans fatty acids d...
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