Selenium binding protein gene BpSBP1 for regulating and controlling selenium metabolism of paper mulberry and application of selenium binding protein gene BpSBP1

Abstract

The invention belongs to the technical field of genetic engineering identification, and particularly relates to a selenium binding protein gene BpSBP1 for regulating and controlling selenium metabolism of broussonetia papyrifera and application of the selenium binding protein gene BpSBP1. The invention provides a selenium binding protein gene BpSBP1 for regulating and controlling selenium metabolism of paper mulberry and application of the selenium binding protein gene BpSBP1. The paper mulberry BpSBP1 gene can improve the selenium resistance of plants; bpSBP1 is related to synthesis of paper mulberry selenomethionine, excessive selenium accumulation of paper mulberry can be inhibited, the synthesis proportion of organic selenium can be adjusted, the root growth capability and biomass of paper mulberry in a selenium stress environment can be improved, and the paper mulberry BpSBP1 is used as a target gene for improving the selenium resistance of paper mulberry. The nutritional value of the broussonetia papyrifera serving as a feed raw material in a high-selenium area is balanced through a genetic engineering means, the problems of toxicity control and yield stability of the selenium-rich broussonetia papyrifera feed are solved, the selenium-rich broussonetia papyrifera within a safety threshold value is cultivated in the high-selenium area to serve as a livestock and poultry feed raw material, and then the requirement for selenium supplement in daily diet of people is met.

Description

Technical Field

[0001] The present invention belongs to the field of genetic engineering identification technology, and specifically relates to a selenium-binding protein gene that regulates selenium metabolism in paper mulberry. BpSBP1 and its applications. Background Art

[0002] Selenium (Se) is an essential trace element for plants and animals, involved in antioxidant defense, immune regulation, and metabolic processes. However, the safe utilization range of selenium is extremely narrow, and excessive intake can cause significant toxicity, seriously endangering the health of animals and plants and the safety of agricultural production. In high-selenium environments, plants are prone to symptoms such as impaired seed germination, inhibited growth, yellowing leaves, root deformities, and even death; animals are prone to "selenium poisoning," manifested by hair and hoof loss, swollen joints, loss of appetite, anemia, and reproductive disorders. In the current production of selenium-enriched feed, the amount of selenium added to the soil or from exogenous sources is difficult to accurately control, which can easily lead to excessive poisoning of animals and plants. Therefore, developing technologies that efficiently regulate selenium metabolism and achieve "safe selenium enrichment" is an urgent need to ensure the sustainable development of the ecology and industry.

[0003] Selenium-binding proteins (SBPs) are a class of important proteins found throughout eukaryotes and specifically binding to selenium. Plant SBPs, as key molecules in selenium metabolism, have garnered significant attention in recent years. Protein interactomics studies have revealed that SBP1 physically interacts with selenocysteine ​​cleavage enzymes (SELENOP) and glutathione synthetase (GS), suggesting a close involvement in selenium metabolism. While the multifunctionality of SBPs in plant selenium metabolism and stress responses has been preliminarily elucidated, their specific molecular mechanisms and potential for application in crop improvement require further exploration.

[0004] Paper mulberry is a multi-purpose economic tree species with application value in papermaking, ecological restoration and feed production, and has a certain ability to accumulate selenium. In paper mulberry leaves treated with exogenous selenium, selenomethionine (SeMet) is the main form of organic selenium, indicating that paper mulberry is expected to become a candidate variety for selenium-enriched feed. However, high concentrations of selenium can have a toxic effect on its growth, limiting its potential for selenium enhancement. Previous studies have focused more on how to improve the selenium accumulation capacity of plants, and rarely mentioned that selenium is not an essential element for plants and can be toxic to growth. Therefore, screening and identifying genes that reduce selenium enrichment are of great value for improving the ecological adaptability of paper mulberry. Selenium-binding protein gene ( BpB 1) Its role in selenium metabolism in paper mulberry is still unclear, so the analysis BpSBP1 The function of the gene is of great significance to improving the selenium tolerance, organic selenium content and feeding value of paper mulberry. Summary of the Invention

[0005] The purpose of the present invention is to provide a selenium-binding protein gene that regulates selenium metabolism in paper mulberry to solve the existing problems. BpSBP1 and its applications.

[0006] The present invention is achieved through the following technical solutions: A selenium-binding protein gene regulating selenium metabolism in paper mulberry BpSBP1 The nucleotide sequence is shown in SEQ ID NO.1, and the amino acid sequence of the protein encoded by it is shown in SEQ ID NO.2.

[0007] SEQ ID NO.1: SEQ ID NO.2: MSWVYNPQYTWGMYSCITVFYIAEDWDFVSLCRTVSVKYLAGNENMRAVIGGPTDYIRLLGKSWGDGAERYGKYLIGEELSDLTPLGGAGHYKGKSLSFSFSFSSLFSLEFIVAVLFFVRFLERESERVLERESYGYLVCKRWSTARVESRNGRGMRQRTRRAAGKWGGREYGLASQGHVWSPEKLSTHHIYVQLRTGIQKPDYLATVDVDPSSPTYSKVIHRLPVPYLGDELHHTGWNSCSSCHGDPSADRRFLIVPSLVSGRIYVVDTKTNPKAPSLHKVVHSEEIVQKTNLGYPHTSHCLASGDVMVSCLGDKDGNAEGNGFLLLDSEFNVKGRWEKPGHSPLFGYDFWYQPRHKTMISSSWGAPAAFTKGFSLDDVANGLYGRHLYVYSWPGGELKQTLDLGNTGLLPLEIRFLHDPSKDTGFVGSALSSNMVRFFKNQDDSWSHEVAIPVKPLKVQNWILPEMPGLITDFLISLDDRYLYFVNWLHGDVRQYNIEDPKNPKLTGQVWTGGLIQKGSPILAEGEDGKTWQFDVPEVQGHKLRGGPQMIQLSLDGKRLYVTNSLFSTWDRQFYPDLPKKGSHMLQIDVDTEKGGLAINPNFFVDFGAEPDGPSLAHEMRYPGGDCTSDIWI* Furthermore, its 2000 bp upstream promoter sequence is as shown in SEQ ID NO.3.

[0008] SEQ ID NO.3: A selenium-binding protein gene regulating selenium metabolism in paper mulberry BpSBP1 Or the application of protein in improving the selenium tolerance of plants.

[0009] A selenium-binding protein gene regulating selenium metabolism in paper mulberry BpSBP1 Or the application of protein in inhibiting excessive selenium accumulation in plants.

[0010] A selenium-binding protein gene regulating selenium metabolism in paper mulberry BpSBP1 Or the application of protein in regulating the ratio of organic selenium.

[0011] A selenium-binding protein gene regulating selenium metabolism in paper mulberry BpSBP1 Or the application of protein in cultivating selenium-rich plants within the safety threshold in high-selenium areas.

[0012] Furthermore, the plant includes Nicotiana tabacum (K326).

[0013] Compared with the prior art, the present invention has the following advantages: The present invention provides a selenium-binding protein gene for regulating selenium metabolism in paper mulberry BpSBP1 and its application, paper mulberry BpSBP1 Genes can improve a plant's selenium tolerance; BpSBP1 It is related to the synthesis of selenomethionine in paper mulberry. It can inhibit the excessive accumulation of selenium in paper mulberry and adjust the synthesis ratio of organic selenium, thereby improving the root growth ability and biomass of paper mulberry under selenium stress environment. BpSBP1 In order to improve the target genes of the selenium tolerance of paper mulberry, genetic engineering is used to balance the nutritional value of paper mulberry as a feed raw material in high-selenium areas, solve the problems of toxicity control and yield stability of selenium-rich paper mulberry feed, and cultivate selenium-rich paper mulberry within the safety threshold in high-selenium areas as a feed raw material for livestock and poultry, thereby meeting people's needs for selenium supplementation in their daily diet. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] Figure 1 The secondary structure and tertiary structure of the BpSPB1 protein provided by the present invention; Figure 2 The present invention provides BpSPB1 response elements of genes; Figure 3 The present invention provides BpSBP1 Subcellular localization map of Figure 4 This is the PCR detection diagram of the transgenic tobacco provided by the present invention; Figure 5 A map of the pNC-Cam1304-MCS-35S::BpSBP1 recombinant plasmid provided by the present invention; Figure 6This is a microscopic observation photo of GUS staining of Nicotiana tabacum provided by the present invention; Figure 7 The growth conditions of different tobacco strains provided by the present invention after being cultured on plates containing sodium selenate; Figure 8 This is a graph showing the total selenium and selenoamino acid contents of selenium-treated tobacco provided by the present invention; Figure 9 The different tobacco strains provided by the present invention NtAPS Relative expression level graph; Figure 10 The different tobacco strains provided by the present invention NtAPR expression level graph; Figure 11 The different tobacco strains provided by the present invention NtSAT expression level graph; Figure 12 The tobacco provided by the present invention NT3C Gene expression level graphs; Figure 13 The tobacco provided by the present invention NtMHT Gene expression levels. DETAILED DESCRIPTION

[0015] In order to further explain the present invention, it is described below with reference to the following specific embodiments.

[0016] The present invention provides paper mulberry BpSBP1 Application of genes in improving plant selenium tolerance. BpSBP1 The roots of the genetically modified tobacco were significantly longer than those of the wild type.

[0017] The present invention also provides paper mulberry BpSBP1 The application of genes in inhibiting excessive selenium accumulation in plants. BpSBP1 The total selenium content of the genetically modified Nicotiana tabacum was lower than that of the wild type.

[0018] The present invention also provides BpSBP1 Application of regulating organic selenium accumulation. BpSBP1 The ratio of selenocysteine ​​to methylselenocysteine ​​in the gene-producing Nicotiana tabacum was higher than that in the wild type.

[0019] In the present invention, the plant includes Nicotiana tabacum.

[0020] The present invention also provides paper mulberry BpSBP1 The application of genes in cultivating selenium-rich plants within safety thresholds.

[0021] The paper mulberry tree of the present invention is described below in conjunction with specific embodiments. BpSBP1 The application of genes is further introduced in detail.

[0022] Example 1 Plant material processing: Nicotiana benthamiana and Nicotiana tabacum (K326) were used for subcellular localization and transgenic experiments, respectively. Seeds were pre-chilled at 4°C before sowing and planted in a nutrient medium (peat:vermiculite:perlite = 7:2:1). When seedlings developed three true leaves, they were transplanted into small pots and grown in a light incubator for two weeks before subcellular localization assays. For Nicotiana tabacum, seeds were sown in the same nutrient medium and grown in a light incubator until six fully expanded true leaves were observed. Plants at this point were used for genetic transformation experiments. Tobacco growth conditions were 28°C (day) and 24°C (night) with a photoperiod of 14 h light / 10 h dark.

[0023] 1. Bioinformatics Analysis The Blast Zone tool of Tbtools was used to establish a local database of Arabidopsis thaliana. BpSBP1 The physicochemical properties of BpSBP1 protein were analyzed through the ExPasy website; the secondary structure of BpSBP1 protein was analyzed using the online website SPOMA; the amino acid sequence of BpSBP1 in Broussonetia papyrifera was predicted using SWISS-MODEL to construct the three-dimensional structure model of the protein; the amino acid sequence of BpSBP1 in Broussonetia papyrifera was predicted using the PlantCare online website BpSBP1 The promoter regions of genes were subjected to cis-element analysis.

[0024] The results showed that the BpSBP1 The gene ID is Bp13G1838, and the gene was compared using Tbtools. BpSBP1 yes AtSBP1 Analysis of the physicochemical properties of the BpSBP1 protein using the ExPasy website revealed a total length of 1932 bp, 644 amino acids, a molecular weight of 71.39 kDa, and a theoretical isoelectric point (PI) of 6.50, indicating an acidic protein. The total number of negatively charged residues (Asp+Glu) is 71, and the total number of positively charged residues (Arg+Lys) is 66. The instability index is 32.89, less than 40, indicating an unstable protein. The aliphatic index is 77.74, and the grand average of hydrophilicity is -0.371, indicating a hydrophilic protein. Figure 1A shows that the secondary structure of the protein is composed of three main types, among which α-helix (Hh) accounts for 13.41%, extended chain (Ee) accounts for 27.13%, and random coil (Cc) accounts for 59.49%, which is the largest proportion. This shows that random coil is an important component in the composition of the secondary structure of BpSBP1 protein. The tertiary structure of BpSBP1 protein is a stable three-dimensional configuration formed by complex spatial rearrangement on the basis of secondary structural elements such as α-helix and β-sheet. The protein structure contains multiple α-helical regions maintained by hydrogen bond networks. These helical structures are connected to each other through turns and extension regions caused by residues such as proline, forming a specific topological framework ( Figure 1 B). Figure 2 show BpSBP1 It contains 9 types of hormone-related abscisic acid response elements (ABRE), methyl jasmonate response elements (CGTCA-motif), light response elements (G-box), anaerobic inducible response elements (ARE), and WRKY transcription factor response elements W-box, totaling 39 response element sites. BpSBP1 Multiple ABRE sites were detected in the promoter region, suggesting that the gene may be involved in the ABA-mediated stress response pathway; the presence of CGTCA-motif indicates BpSBP1 It is also possible that it responds to jasmonic acid signals and participates in the regulation of plant stress resistance.

[0025] 2. BpSBP1 Gene cloning and sequencing (1) RNA extraction and reverse transcription: Total RNA was extracted using the RNA prep Pure Plant Kit DP432 (Tiangen Biochemical Technology Co., Ltd., Beijing, China) and reverse transcribed according to the manufacturer's instructions of the HiScript II 1st Strand cDNA Synthesis Kit (Nanjing Novozyme Biotechnology Co., Ltd.). Specific primers were designed using Primer Premier 6 (Premier Biosoft, UK), and Ubiquitin (UBI11) was used as an internal reference gene. -ΔΔCT Methods The relative expression levels of genes were calculated.

[0026] (2) BpSBP1 Cloning and vector construction: Select transcriptome analysis to identify genes that are highly correlated with selenium content BpSBP1 Functional verification was performed. The complete ORF was amplified by PCR using the primers listed in Table 1. The PCR product was purified by gel extraction and cloned into the pMD19-T vector (Takara), which was then transformed into Escherichia coli DH5α. Positive clones were verified by colony PCR and Sanger sequencing.

[0027] 3. Construction of the plant overexpression vector pNC-Cam1304-MCS-35S::BpSBP1 To overexpress BpSBP1 , re-amplify using one-step cloning primers (Table 1) BpSBP1 The binary vector pNC-Cam1304-MCS-35S was linearized with SfiI and inserted into the vector using the Ultra One Step Cloning Kit (Vazyme, Nanjing, China). BpSBP1 The recombinant plasmid pNC-Cam1304-MCS-35S::BpSBP1 was transformed into DH5α, and positive clones were screened by PCR using primers M13-47 / M13-48.

[0028] Table 1 Primer sequences used in the present invention

[0029] 4. Transformation of pNC-Cam1304-MCS-35S::BpSBP1 plasmid into Agrobacterium Plasmids were isolated from positive clones using the EasyPure Plasmid MiniPrep Kit (TransGen Biotech) and transformed into Agrobacterium tumefaciens GV3101 by electroporation for subsequent plant transformation experiments.

[0030] 5. Leaf disc method for tobacco plant transformation Agrobacterium GV3101 harboring the recombinant vector pNC-Cam1304-MCS-35S::BpSBP1 was revived overnight, subcultured to an OD600 of ≈ 0.8, harvested by centrifugation (4,000 rpm, 10 min), and resuspended in MS medium containing 100 μM acetosyringone (AS) to an OD600 of ≈ 0.4. Sterilized Nicotiana tabacum leaf explants (5 × 5 mm) were immersed in the bacterial suspension for 15 min, smeared, and incubated in the dark for 48 h on MS medium containing 2 mg / L 6-BA, 0.5 mg / L IAA, and 100 μM AS. The explants were then transferred to shoot induction medium (MS supplemented with 2 mg / L 6-BA, 0.5 mg / L IAA, 20 mg / L hygromycin, and 200 mg / L ceftriaxone) under a 16-h light / 8-h dark cycle. New shoots (1-2 cm) were excised and rooted on MS medium supplemented with 1 mg / L IBA, 20 mg / L hygromycin, and 200 mg / L cephalosporin. Regenerated plants were identified by PCR and GUS staining before acclimation in the greenhouse.

[0031] 5.1 Tobacco PCR Identification Take the transgenic tobacco leaves grown in a light incubator for about one month and use the CTAB method to obtain crude DNA. Then use the DNA as a template and use the BpSBP1-OE-F and BpSBP1-OE-R primers to detect whether the transgenic tobacco plants are positive ( Figure 4 ).

[0032] The results showed that the three tobacco lines were detected BpSBP1 The presence of genes ( Figure 4 ).

[0033] 5.2 GUS staining Wild-type and transgenic tobacco seedlings grown in MS medium for approximately two weeks were placed in centrifuge tubes. Premixed GUS stain was added according to the Gusbluekit (Huayueyang, Beijing) kit instructions to submerge the leaves. The leaves were incubated in a 37°C incubator for 12 hours. The stain was then removed and decolorized with 75% ethanol, with the alcohol replaced every 8 hours until the leaves became white and transparent. The staining was observed and photographed using a stereomicroscope (Leica M165FC, Leica). Figure 6 ).

[0034] GUS staining was performed on wild-type and transgenic tobacco leaves, and GUS activity was detected in transgenic tobacco leaves ( Figure 6 ), speculated BpSBP1 Actively expressed in leaves.

[0035] 6. Subcellular Localization The pCAMBIA1304-35S::BpSBP1-eGFP fusion vector, the eGFP-only control vector, and the nuclear marker vector pCAMBIA2300-35S-H2B-mCherry-OCS (Biolabs) were prepared and transformed into Agrobacterium tumefaciens GV3101 by electroporation. Fluorescence visualization was performed using a Leica TCSSP8 confocal laser scanning microscope (Leica Microsystems, Germany). Figure 3 ).

[0036] The BpSBP1-GFP fusion vector under the CaMV35S promoter was transiently expressed in Nicotiana benthamiana. Confocal microscopy revealed that the GFP signal extensively overlapped with the nuclear marker mCherry, and weaker fluorescence was detected in the cytoplasm ( Figure 3 ),show BpSBP1 May have functions in both the nucleus and the cytoplasm.

[0037] Example 2 The results of exogenous sodium selenate application on tobacco were compared with those on wild-type and transgenic lines, including root length, seedling height, total selenium content and selenium morphology. BpSBP1 Genetic influences on the selenium accumulation capacity of plants.

[0038] 1. Verification of Selenium Tolerance in Transgenic Tobacco Will BpSBP1 Seeds of overexpressing tobacco lines (OE01, OE02, and OE03) and wild-type strains were sown on MS agar plates supplemented with 75 mg / L sodium selenate (selected from preliminary experiments). The plates were placed vertically in a growth chamber at 25°C / 23°C (day / night) with a 14 h light / 10 h dark photoperiod. After 15 days, the root length and height of each seedling were measured.

[0039] The results are as follows Figure 7 The results showed that on MS medium supplemented with 75 mg / L Na2SeO4, wild-type plants exhibited typical selenium toxicity symptoms, such as leaf chlorosis and severely stunted root development (8.09 cm). In contrast, OE lines maintained green leaves and had root lengths of 18.85, 22.43, and 19.49 cm, respectively, which were 1.32-1.96 times longer than those of the wild type.

[0040] 2. Verification of Selenium Accumulation in Transgenic Tobacco Using 75 mg / L sodium selenate BpSBP1 Overexpression and wild-type tobacco lines were foliarly sprayed. Spraying was performed every 5 days for a total of 3 times. Leaves were collected on day 15, flash-frozen in liquid nitrogen, and stored at -80°C for selenium content and gene expression analysis. Total RNA was extracted as described above and qRT-PCR was performed using specific primers to quantify BpSBP1 and endogenous tobacco selenium metabolism gene transcripts.

[0041] 3. Determination of Total Selenium and Selenium Species 3.1 Total selenium content detection For total selenium determination, 0.2 g of dried sample was digested with nitric acid and hydrogen peroxide in a microwave digester (YMW-HSP100, Yonglekang Instrument, Changsha, China), adjusted to 10 mL with distilled water, and measured using a hydride generation atomic fluorescence spectrometer (AFS8510, Haiguang Instrument, Beijing, China). For selenium speciation, 0.2 g of lyophilized sample was incubated with proteinase XIV (4 mg / mL) at 37°C for 16 h. The resulting supernatant was filtered through a 0.45 μm membrane before analysis. For total selenium determination, 0.2 g of dried sample was digested with nitric acid and hydrogen peroxide in a microwave digester (YMW-HSP100, Yonglekang Instrument, Changsha, China), adjusted to 10 mL with distilled water, and measured using a hydride generation atomic fluorescence spectrometer (AFS8510, Haiguang Instrument, Beijing, China).

[0042] Under the condition of foliar spraying of the same concentration of sodium selenate, the selenium accumulation in transgenic leaves (48.87-63.88 mgkg-1DW) was significantly lower than that in wild-type leaves (93.43 mgkg-1DW, Figure 8 ).

[0043] 3.2 Selenium speciation determination Selenium species were detected using LC-AFS. Protease XIV was prepared at 8 mg / mL in 0.01 mol / L phosphate buffer. 0.2 g of tobacco powder was weighed and added to 8 mL of Protease XIV in a 10 mL centrifuge tube. The mixture was incubated at 37°C with shaking for 16 h, followed by ultrasonic extraction at room temperature for 30 min and centrifugation at 10,000 rpm for 10 min at 4°C. The supernatant was filtered through a 0.45 μm aqueous filter and used for analysis. Mobile phases A and B were prepared separately: mobile phase A consisted of 5 mM citric acid (pH 4.6) and mobile phase B consisted of 40 mM potassium dihydrogen phosphate (pH 6.0). The gradient program was as follows: phase A for 8 min, A->B for 1 min, phase B for 6 min, and B->A for 1 min. SeCys2, SeMeCys, Se (IV), SeMet, and Se (VI) standard solutions were prepared and diluted to a gradient of 40, 80, 120, 160, and 200 μg / L (in terms of Se). A Hamilton PRP-X (Hamilton, OH, USA) cathode column was used for chromatography, with a column temperature of 25°C, argon (99.999%) as the carrier gas, a flow rate of 600 mL / min, a negative high voltage of 320 V, and a total lamp current of 80 mA. Three biological replicates were designed for each sample group, and each sample was measured three times. Selenium speciation analysis showed that ( Figure 8 ), OE strain (6.51-15.28 mgkg -1 DW) compared with the wild type (41.54 mg / kg -1SeMet in DW was significantly reduced, the proportion of selenomethionine in organic selenium decreased from 83.1% to 35.5%-44.5%, and the proportion of SeCys2 and MeSeCys was adjusted from 9.1% to 7.5%-21.2%, indicating that BpB Overexpression inhibited the conversion of inorganic Se to organic Se and adjusted the ratio of organic Se.

[0044] Example 3 Real-time quantitative fluorescence PCR was used to detect selenium metabolism-related genes in selenium-treated tobacco, and the selenium metabolism-related genes in paper mulberry were analyzed. BpSBP1 Effects of genes on plant selenium metabolism and synthesis pathway genes.

[0045] 1. RNA Extraction and Reverse Transcription from Nicotiana tabacum The method of step 2 in Example 1 was used to extract RNA from leaves of the WT and three transgenic tobacco lines treated with sodium selenate foliar spray in Example 2 and reverse transcribed to obtain cDNA.

[0046] 2. Real-time quantitative PCR detection of selenium metabolism genes in Nicotiana tabacum The tobacco genome data was downloaded from EnsemblPlants (https: / / plants.ensembl.org / Arabidopsis_thaliana / Info / Index). Based on the plant selenium metabolic pathway, adenosine triphosphate sulfurylase (NtAPS), adenosine triphosphate sulfate reductase (NtAPR), serine O-acetyltransferase (SAT), cysteine ​​synthetase (CS), and (HMT) in the selenium metabolic pathway were selected from the tobacco genome. The CDS sequences of the corresponding genes were extracted using Tbtools, and PrimerPremier 6.0 was used to design the gene expression vectors including BpSBP1 Gene quantitative detection primers. Perform qRT-PCR detection using the cDNA of each tobacco strain reverse-transcribed in step 1 as template and β-Action as the internal reference gene.

[0047] The results showed that qRT-PCR of selenium metabolism genes showed differential regulation in OE strains: sulfate-activated genes NtAPS1 , NtAPS2 and NtAPS6 Significantly downregulated ( Figure 9 ). At the same time, compared with the wild type , NtAPR1 Upregulated expression, other NtAPR ( NtAPR2 、 NtAPR3 and NtAPR4 ) gene down-regulated expression ( Figure 10 ). Serine O-acetyltransferase NtSAT1 Cysteine ​​synthase NtCS2 Upregulation ( Figure 11, Figure 12 Among the SeMet synthesis genes, Nt HMT1 The expression level of Nt HMT6 The expression level of other homologous genes was significantly increased; the expression of other homologous genes in OE lines was different ( Figure 13 ).

[0048] In summary, BpSBP1 The selenium tolerance of transgenic tobacco was improved, and the proportion of organic selenium was adjusted while the total selenium content of the plant was reduced. BpSBP1 It is a key gene involved in selenium metabolism and can improve the tolerance of plants to high-selenium areas.

[0049] The above description is only a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any modifications, equivalent substitutions and improvements made by any technician familiar with this technical field within the technical scope disclosed by the present invention and within the spirit and principles of the present invention should be covered by the scope of protection of the present invention.

Claims

1. A selenium-binding protein gene that regulates selenium metabolism in paper mulberry BpSBP1 , characterized in that, The nucleotide sequence thereof is shown as SEQ ID NO.1, and the amino acid sequence of the protein encoded by the same is shown as SEQ ID NO.

2.

2. The selenium-binding protein gene for regulating paper mulberry selenium metabolism according to claim 1 BpSBP1 Or the application of protein in improving the selenium tolerance of plants.

3. The selenium-binding protein gene for regulating selenium metabolism in paper mulberry according to claim 1 BpSBP1 Or the application of protein in inhibiting excessive selenium accumulation in plants.

4. The selenium-binding protein gene for regulating paper mulberry selenium metabolism according to claim 1 BpSBP1 Or the application of protein in regulating the ratio of organic selenium.

5. The selenium-binding protein gene for regulating selenium metabolism in paper mulberry according to claim 1 BpSBP1 Or the application of protein in cultivating selenium-rich plants within the safety threshold in high-selenium areas.

6. The use according to any one of claims 2 to 5, characterized in that The plants include Nicotiana tabacum.

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