Application of Benincasa hispida CqbZIP7 gene and protein encoded thereby in improving cold tolerance of plants
By overexpressing the CqbZIP7 gene in tobacco, it enhances its tolerance to low temperature stress, solves the problem of cold stress in the seedlings period and improves the cold resistance of plants.
Patent Information
- Application Number
- CN202510050818.2
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2025-01-13
- Publication Date
- 2025-07-11
- Estimated Expiration
- 2045-01-13
AI Technical Summary
In the early spring and winter production in South China, melons are susceptible to cold stress in seedling stage, resulting in slow growth, aging and even death, affecting premature maturity and high yields. The existing technology is difficult to effectively improve its resistance to low temperatures.
By cloning the CqbZIP7 gene on the pCAMBIA1300 vector, the overexpression vector was constructed and transformed tobacco, and the CqbZIP7 transgenic plant was obtained, which increased the expression of the CqbZIP7 gene in the plant and enhanced the tolerance to low temperature stress.
Transgenic tobacco overexpressing the CqbZIP7 gene is less affected under low temperature stress, leaves wilting are reduced, hydrogen peroxide production is reduced, enzyme activity is increased, and cell membrane damage is reduced, which significantly improves resistance to low temperature.
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Figure CN119614617B_ABST
Abstract
Description
Technical Field
[0001] The present invention belongs to the technical field of plant genetic engineering. Specifically, the present invention relates to the application of a Chieh-qua CqbZIP7 gene, the encoded protein, an overexpression vector, and a recombinant bacterium in improving the cold stress tolerance of plants. Background Art
[0002] Chieh-qua (Benincasa hispida Cogn. var. Chieh-qua How), also known as hairy melon and hairy Chieh-qua, is a variety of the wax gourd species in the genus Benincasa of the Cucurbitaceae family. It is rich in nutrients such as vitamins and malonic acid, and is a major vegetable crop exported in South China. Chieh-qua originated in subtropical and tropical regions, and the suitable temperature range for its growth is 20°C to 30°C. It likes warmth and is not cold-tolerant. However, in the early spring and winter production in South China, it is easily damaged by cold stress at the seedling stage. When the seedlings grow in cold conditions, the leaves turn yellow, showing water-soaked spots, the plant growth is slow, and it may even age and die, seriously restricting its early maturity and high yield.
[0003] With the increasing demand for year-round production of Chieh-qua, especially in early spring and winter, improving the cold stress resistance of Chieh-qua seedlings at the seedling stage has become an urgent problem to be solved. Exploring the key genes responsive to cold stress and molecular-assisted selection of excellent germplasms resistant to cold at the seedling stage are the fundamental ways to solve the current problem. Summary of the Invention
[0004] Based on this, the purpose of the present invention is to provide the application of the Chieh-qua CqbZIP7 gene and the protein encoded thereby in improving the cold resistance of plants.
[0005] The specific technical solutions for achieving the above invention purposes are as follows.
[0006] In the first aspect of the present invention, there is provided the application of the Chieh-qua CqbZIP7 gene in improving the cold resistance of plants or improving plant cold-resistant germplasm resources, and the nucleotide sequence of the Chieh-qua CqbZIP7 gene is as shown in SEQ ID NO:1.
[0007] In the second aspect of the present invention, there is provided the application of the protein encoded by the Chieh-qua CqbZIP7 gene in improving the cold resistance of plants or improving plant cold-resistant germplasm resources, and the amino acid sequence of the protein is as shown in SEQ ID NO:2.
[0008] In the third aspect of the present invention, there is provided the application of the overexpression vector of the Chieh-qua CqbZIP7 gene in improving the cold resistance of plants or improving plant cold-resistant germplasm resources, and the nucleotide sequence of the Chieh-qua CqbZIP7 gene is as shown in SEQ ID NO:1.
[0009] In the fourth aspect of the present invention, there is provided the use of a recombinant bacterium transformed with an overexpression vector of the CqbZIP7 gene of wax gourd in improving the cold resistance of plants or improving the cold-resistant germplasm resources of plants, wherein the nucleotide sequence of the CqbZIP7 gene of wax gourd is as shown in SEQ ID NO:1.
[0010] In the fifth aspect of the present invention, there is provided a preparation for improving the cold resistance of plants, and the active ingredient of the preparation is a recombinant bacterium transformed with an overexpression vector of the CqbZIP7 gene of wax gourd, wherein the nucleotide sequence of the CqbZIP7 gene of wax gourd is as shown in SEQ ID NO:1.
[0011] In the sixth aspect of the present invention, there is provided a method for improving the cold resistance of plants, comprising the following steps: increasing the expression of the CqbZIP7 gene of wax gourd in plants, wherein the nucleotide sequence of the CqbZIP7 gene of wax gourd is as
[0012] shown in SEQ ID NO:1.
[0013] In the present invention, by cloning the CqbZIP7 gene of wax gourd into the pCAMBIA1300 vector, the fusion vector pCAMBIA1300-CqbZIP7 (i.e., the overexpression vector of the CqbZIP7 gene of wax gourd) is obtained. After transforming tobacco with Agrobacterium tumefaciens, CqbZIP7 transgenic tobacco plants are obtained. The T2 generation plants of CqbZIP7 transgenic tobacco and wild-type tobacco plants are simultaneously placed at 4 °C for low-temperature treatment for 1 day and 7 days. Phenotypic observation shows that the growth of transgenic plants is less affected by low-temperature stress, while the growth of wild-type plants is inhibited and the leaves show wilting. DAB staining shows that the transgenic tobacco produces less hydrogen peroxide than the wild type. After 1 day of low-temperature treatment, compared with the wild type, the relative conductivity of the leaves of transgenic tobacco is higher, the enzyme activities of PAO and POD increase significantly, and the MDA content decreases significantly. Along with the increase in the expression level of the CqbZIP7 gene of wax gourd, the cold resistance of transgenic tobacco is correspondingly improved. These results indicate that the CqbZIP7 gene of wax gourd is involved in responding to low-temperature stress, and overexpression of the CqbZIP7 gene of wax gourd can enhance the tolerance of plants to low-temperature stress.
[0014] The discovery of the cold-resistant function of the CqbZIP7 gene of wax gourd provides new gene targets and resources for cultivating cold-tolerant wax gourd varieties, has important significance for the study of the cold-tolerant molecular mechanism of wax gourd, lays a certain theoretical foundation for the study of the mechanism of plant response to low-temperature stress and the molecular mechanism of resisting adverse environments, and has great significance for agricultural production. Description of the Drawings
[0015] Figure 1Detection results of tobacco plants overexpressing the CqbZIP7 gene of wax gourd in Example 1 of the present invention; among them, lanes 1, 3, 4, 6-32 are transgenic tobacco positive plants; lanes 2, 5 are transgenic tobacco negative plants; "+" is the positive control; "-" is the negative control.
[0016] Figure 2 Detection results of low temperature tolerance of tobacco plants (OE6 and OE12) overexpressing the CqbZIP7 gene of wax gourd and wild-type tobacco plants in Example 3 of the present invention; among them, A is the phenotypic diagram before and after low temperature stress; B is the DAB staining diagram under normal and low temperature stress for 24 hours; C is the detection result of the CqbZIP7 gene expression level of wild-type and transgenic plants; D is the relative conductivity results of wild-type and transgenic plants under normal temperature (RT) and low temperature (LT) treatments; E is the PAO activity results of wild-type and transgenic plants under normal temperature (RT) and low temperature (LT) treatments; F is the POD activity results of wild-type and transgenic plants under normal temperature (RT) and low temperature (LT) treatments; G is the MDA content results of wild-type and transgenic plants under normal temperature (RT) and low temperature (LT) treatments. Detailed implementation manners
[0017] To facilitate the understanding of the present invention, the present invention will be described more comprehensively below. The present invention can be implemented in many different forms and is not limited to the embodiments described herein. On the contrary, these embodiments are provided to make the understanding of the disclosed content of the present invention more thorough and comprehensive.
[0018] Unless otherwise defined, all technical and scientific terms used in the present invention have the same meaning as commonly understood by those skilled in the technical field to which the present invention belongs. The terms used in the description of the present invention are only for the purpose of describing specific embodiments and are not used to limit the present invention. The term "and / or" used in the present invention includes any and all combinations of one or more of the related listed items.
[0019] If not specifically specified, the embodiments are all carried out under conventional experimental conditions, such as those in the Molecular Cloning Experimental Manual by Sambrook et al. (Sambrook J & Russell DW, Molecular Cloning: a Laboratory Manual, 2013), or according to the conditions recommended by the manufacturer's instructions.
[0020] In some embodiments of the present invention, the application of the CqbZIP7 gene of wax gourd in improving the cold resistance of plants is disclosed, and the nucleotide sequence of the CqbZIP7 gene of wax gourd is as shown in SEQ ID NO: 1.
[0021] In some other embodiments of the present invention, the application of the Chieh-qua CqbZIP7 gene in improving plant cold-resistant germplasm resources is disclosed, and the nucleotide sequence of the Chieh-qua CqbZIP7 gene is as shown in SEQ ID NO:1.
[0022] In some other embodiments of the present invention, the application of the protein encoded by the Chieh-qua CqbZIP7 gene in improving plant cold resistance is disclosed, and the amino acid sequence of the encoded protein is as shown in SEQ ID NO:2.
[0023] In some other embodiments of the present invention, the application of the encoding gene of the Chieh-qua CqbZIP7 gene in improving plant cold-resistant germplasm resources is disclosed, and the amino acid sequence of the encoded protein is as shown in SEQ ID NO:2.
[0024] In some other embodiments of the present invention, the application of the overexpression vector of the Chieh-qua CqbZIP7 gene in improving plant cold resistance is disclosed, and the nucleotide sequence of the Chieh-qua CqbZIP7 gene is as shown in SEQ ID NO:1.
[0025] In some other embodiments of the present invention, the application of the overexpression vector of the Chieh-qua CqbZIP7 gene in improving plant cold-resistant germplasm resources is disclosed, and the nucleotide sequence of the Chieh-qua CqbZIP7 gene is as shown in SEQ ID NO:1.
[0026] In some other embodiments of the present invention, the application of the recombinant bacterium transformed with the overexpression vector of the Chieh-qua CqbZIP7 gene in improving plant cold resistance is disclosed, and the nucleotide sequence of the Chieh-qua CqbZIP7 gene is as shown in SEQ ID NO:1.
[0027] In some other embodiments of the present invention, the application of the recombinant bacterium transformed with the overexpression vector of the Chieh-qua CqbZIP7 gene in improving plant cold-resistant germplasm resources is disclosed, and the nucleotide sequence of the Chieh-qua CqbZIP7 gene is as shown in SEQ ID NO:1.
[0028] In some other embodiments of the present invention, a preparation for improving plant cold resistance is disclosed, and the active ingredient of the preparation is the above-mentioned recombinant bacterium transformed with the overexpression vector of the Chieh-qua CqbZIP7 gene.
[0029] In some other embodiments of the present invention, a method for improving plant cold resistance is disclosed, including the following steps: increasing the expression of the Chieh-qua CqbZIP7 gene in plants, and the nucleotide sequence of the Chieh-qua CqbZIP7 gene is as shown in SEQ ID NO:1.
[0030] In some of these embodiments, the plant is a cucurbitaceous plant or tobacco.
[0031] In some of these embodiments, the cucurbitaceous plant is Benincasa hispida var. chieh-qua.
[0032] It should be understood that considering the degeneracy of codons, modifying the base sequences involved in the following embodiments without changing the amino acid sequence also falls within the protection scope of the present invention.
[0033] The present invention will be described in detail below with reference to the accompanying drawings and specific embodiments.
[0034] Example 1 Obtaining of CqbZIP7 Gene of Benincasa hispida var. chieh-qua
[0035] In this example, the total RNA of young leaves of Benincasa hispida var. chieh-qua at the seedling stage was reverse transcribed into cDNA, and the CqbZIP7 gene of Benincasa hispida var. chieh-qua was obtained by PCR amplification using the cDNA as a template. The specific steps are as follows:
[0036] 1. RNA extraction and cDNA obtaining
[0037] Approximately 1 g of seedling leaves of 'Yueguang Benincasa hispida var. chieh-qua' (from the Vegetable Research Institute of Guangdong Academy of Agricultural Sciences) was selected. After adding liquid nitrogen, the tissues and cells were lysed in a mortar and quickly transferred into a 2.0 mL centrifuge tube. The plant total RNA extraction kit (RNAprep pure Tissue Kit, TIANGEN) was used to extract the leaf RNA. The concentration of RNA was detected with a spectrophotometer, and the integrity of RNA was detected by agarose gel electrophoresis at the same time.
[0038] Next, using the reverse transcription kit of Takara Biotechnology (Dalian) Co., Ltd. (Prime Script Reverse Transcriptase kit, Takara), 1 μL of Oligo dT Primer, 1 μL of PrimeScript RT Enzyme MixI, 4 μL of 5X PrimeScript Buffer, and 1000 ng of template RNA were mixed (the specific volume was calculated according to the concentration of template RNA), and RNase Free dH2O was added to make up the total volume to 20 μL. It was reverse transcribed into cDNA at 37 °C for 15 minutes and 85 °C for 5 seconds, and stored in a -20 °C refrigerator for standby.
[0039] 2. Amplification of CqbZIP7 gene
[0040] Using the total cDNA after reverse transcription as a template, and using CqbZIP7-F (SEQ ID NO:3): ATGAACTCATCATCCACTAAA and CqbZIP7-R (SEQ ID NO:4): CTATGCAGGCTCACAA GGACG as the forward primer and reverse primer, PCR amplification was carried out using the high-fidelity enzyme (PrimeSTAR Max DNA Polymerase, Takara) from TaKaRa Biotechnology (Dalian) Co., Ltd.
[0041] The PCR reaction system was as follows: 1 μL of cDNA template, 0.5 μL of CqbZIP7-F, 0.5 μL of CqbZIP7-R, 25 μL of PrimeSTAR Max Premix (2X), and 23 μL of dH2O.
[0042] The PCR reaction program was as follows: pre-denaturation at 94 °C for 5 minutes; denaturation at 94 °C for 30 seconds, annealing at 58 °C for 2 minutes, extension at 72 °C for 45 seconds, for 34 cycles; extension at 72 °C for 8 minutes.
[0043] The PCR amplification product was sequenced, and the sequence of the CqbZIP7 gene of wax gourd was obtained. Its nucleotide sequence is shown in SEQ ID NO:1, and the amino acid sequence of the encoded protein is shown in SEQ ID NO:2.
[0044] SEQ ID NO:1:
[0045] ATGAACTCATCATCCACTAAATTTTACAATCCAGGAAGGATGGGATTGTATG
[0046] AACCTATCCACCATATTGGAATGTGGGGAGAAACTTTTAGAACCAATGCCA
[0047] ATTTAGATCCACCATCCTCCTTTATTATTGAAGCTGATACCAAGCTAGAAAA
[0048] TCAGTCCGATGATGCTTCTCTCGGCTCACTTGGAGATCCACATGTATTTGAT
[0049] CAAGATGATACCAAACGCATTGATAAGATTCAAAGACGCCTGGCACAAAAT
[0050] CGGGAAGCAGCTCGCAAAAGTCGCTTGCGGAAAAAGGCTTATATTAAGCA
[0051] ATTGGAAACAAGCCGCTTAAAACTCATTCAATTGGAGCAAGAGCTTGAAA
[0052] AGGCAAGACAACAAGGTCTCTTGGCTGGATCCCGATTTGATAATAACCAGT
[0053] TGGGTTTATCAGGAACCACAAATTCAGGCATCTCTGCATTTGAATCAGAGT
[0054] ATGAGCAATGGGTGGAGGAGCAGAACAGGCAGATTTGCGATCTGAGGACT
[0055] GCTGTGCATGCCGATATTACTGATATCGAGCTTCGAATACTCGTAGAAAATG
[0056] CAATGAGACACTACTTTAAATTTTTTCGCATGAAAGCTAAAGCTGCAAAAG
[0057] CCGATGTCTCTTACATAATGTCAGGCATGTGGAAAACATCAGCCGAAAGAC
[0058] TTTTCTTATGGATAGGAGGATTTCGCCCTTCAGAACTTCTCAAGGTTTTGAT
[0059] ACCTCAACTGGAGACATTAACTGAACAACAAATTTCGGAAACTGGTAGCC
[0060] TTAGGAAATCTTGTCTACAAGCAGAAGATGCTCTTAGACAAGGTATGGAAA
[0061] AACTACAACAAAATCTATTTGATAGCGTAGTGGCTGGTCAGCTGGGAGAAG
[0062] GAAGTCATCCTCTACAAATGACTGCTGCAATGGAGAGATTAGAAGCGCTCA
[0063] TTAGCTTTGTGAATCAGGCTGACCATCTACGACAGGAAACATTGCAACAGA
[0064] TGTACAAAATTCTAACGACTCGACAATCTGCTCAAGGCCTTCTTACTCTCG
[0065] GGGAGTTTTTCCAACGACTCCGAGCATTGAGCTCGCTTTGGGCTAATCGTC
[0066] CTTGTGAGCCTGCATAG
[0067] SEQ ID NO:2:
[0068] MNSSSTKFYNPGRMGLYEPIHHIGMWGETFRTNANLDPPSSFIIEADTKL
[0069] ENQSDDASLGSLGDPHVFDQDDTKRIDKIQRRLAQNREAARKSRLRKKAY
[0070] IKQLETSRLKLIQLEQELEKARQQGLLAGSRFDNNQLGLSGTTNSGISAF
[0071] ESEYEQWVEEQNRQICDLRTAVHADITDIELRILVENAMRHYFKFFRMKA
[0072] KAAKADVSYIMSGMWKTSAERLFLWIGGFRPSELLKVLIPQLETLTEQQI
[0073] SETGSLRKSCLQAEDALRQGMEKLQQNLFDSVVAGQLGEGSHPLQMTAAM
[0074] ERLEALISFVNQADHLRQETLQQMYKILTTRQSAQGLLTLGEFFQRLRAL Example 2 Construction of an overexpression vector of the CqbZIP7 gene of Chieh-qua and obtaining transgenic tobacco plants
[0075] It includes the following steps:
[0076] 1. Connect the CqbZIP7 gene of Chieh-qua to the pCAMBIA1300 plasmid by In-fusion technology, select positive clones for sequencing after transformation, and the correct plasmid is the overexpression vector of the CqbZIP7 gene of Chieh-qua.
[0077] 2. By using the liquid nitrogen freeze-thaw method, the overexpression vector was transformed into Agrobacterium tumefaciens GV3101. Rifampicin at 50 mg / L and Kanamycin at 50 mg / L were used for resistance screening, and the obtained monoclonal colonies were identified by PCR to obtain positive clones.
[0078] 3. Obtain transgenic tobacco plants
[0079] (1) Pre-culture: Tobacco seeds were disinfected with 75% ethanol for 30 s, washed with sterile water for 1 min, then disinfected with 84 disinfectant for 3 - 5 min, and washed with sterile water 3 times, 1 min each time. The disinfected tobacco seeds were sown on MS medium (MS MEDIUM w / VITAMINS&SUCROSE, Phyto-Technology), and cultured at 23°C with a 16 h / 8 h light / dark cycle for 4 weeks. The sterile tobacco leaves were cut into small pieces with a scalpel and inoculated on MS medium.
[0080] (2) Agrobacterium infection and co-culture: Use the infection solution (1.07 g of MES, 1.02 g of MgCl₂·6H₂O, add acetosyringone with a final concentration of 200 μM, add water to a final volume of 500 mL, and adjust the pH value to 5.0 - 5.4). The Agrobacterium tumefaciens prepared in step 2 above was made into a suspension with an OD 600 = 0.2. The tobacco leaves pre-cultured for 3 days were inoculated into the Agrobacterium suspension for 15 min, and the infected tobacco leaves were placed on filter paper, air-dried and then placed on the co-culture medium (MS + 50 mg / L Kan), and cultured in the dark for 72 h.
[0081] (3) Induction and screening: The tobacco leaves after 2 days of co-culture were transferred to the induction medium (MS + 0.1 mg / L IAA + 1.5 mg / L 6 - BA + 50 mg / L Kan + 500 mg / L Carb) to induce callus for about 10 days until callus grew out. Select vigorously growing callus for subculture (MS + 0.1 mg / L IAA + 1.5 mg / L 6 - BA + 50 mg / L Kan + 500 mg / L Carb), 4 - 5 callus per petri dish, and culture at 23°C with a 16 h / 8 h light / dark cycle for 20 days.
[0082] (4) Differentiation and rooting: Select vigorously growing callus for subculture (MS + 0.1 mg / L IAA + 1.5 mg / L 6 - BA + 50 mg / L Kan + 500 mg / L Carb) until seedlings were formed, and then transfer them to the rooting medium (MS + 0.5 mg / L IAA) to grow for 7 - 10 days.
[0083] 4. Detect transgenic tobacco plants
[0084] The genomic DNA of tobacco was extracted by the CTAB method and subjected to PCR detection. The T0 generation of tobacco was planted in sterilized peat soil and cultured in an incubator with a light intensity of 120 μmol / (m 2 ·s) to 150 μmol / (m 2 ·s), a photoperiod of 16 h light and 8 h dark. The T1 generation of seeds was collected and the T1 seeds were continuously planted. The T1 tobacco plants were detected by detection primers. Detection primer Bar-F (SEQ ID NO:5): CGGCGA CGAGCCAGGGATA; Detection primer Bar-R (SEQ ID NO:6): GCACCATCGTCAA CCACTACAT. The results are as Figure 1 shown. The results showed that 30 T1 tobacco plants were positive plants.
[0085] Example 3 Detection of Low Temperature Tolerance of Transgenic Plants Overexpressing the CqbZIP7 Gene of Wax Gourd
[0086] Six pots of wild-type and T2 generation tobacco seedlings (OE6 and OE12) overexpressing the CqbZIP7 gene of wax gourd with good growth and consistent growth were selected and subjected to different temperature treatments (light intensity of 500 - 600 μmol / (m²·s), relative humidity of 65% - 75%) in two artificial climate chambers. One group was the control (RT) with a temperature set at 25 °C; the other group was the low temperature treatment (LT) with a temperature set at 4 °C, and the low temperature stress was 24 h. Three biological replicates were set for each treatment. Excel 2019 was used for data calculation and processing, SPSS software was used for significant difference analysis (P < 0.05), and Graphpad Prism 9.5 was used for chart drawing.
[0087] 1. Plant phenotype
[0088] The growth of transgenic tobacco OE12 and OE6 overexpressing the CqbZIP7 gene of wax gourd was stronger than that of wild-type plants under low temperature stress, and the leaves of wild-type plants were significantly withered ( Figure 2 A in).
[0089] 2. DAB staining
[0090] The leaves of the control (RT) and the low temperature treatment (LT) for 24 hours were collected and placed in a glass bottle for tissue culture, and an appropriate amount of DAB staining solution (1 mg / mL DAB, water-soluble, pH 3.8) was poured in. After overnight staining, the background was decolorized with 95% ethanol.
[0091] The DAB staining results showed that the hydrogen peroxide produced by transgenic tobacco OE12 and OE6 was significantly less than that of wild-type plants ( Figure 2 B in).
[0092] 3. Expression level of CqbZIP7 gene
[0093] Take the leaves of positive plants and wild - type plants before low - temperature stress treatment, extract total RNA respectively, and reverse - transcribe it into cDNA (the same as in Example 1). Use qPCR reagent premix (PerfectStart Green qPCR SuperMix, Transgen) to detect the expression level of CqbZIP7 gene in transgenic tobacco by q - PCR.
[0094] The reaction system is: 0.5 μL of cDNA template, 0.2 μL each of CqbZIP7 - F and CqbZIP7 - R, 5 μL of qPCR SuperMix (2X), and make up to 10 μL with dH2O.
[0095] The PCR reaction program is: 30 seconds at 94 °C; 5 seconds at 94 °C, 30 minutes at 60 °C, for 40 cycles.
[0096] The results are as shown in Figure 2 C in it. It can be seen from the figure that the expression level of CqbZIP7 gene of Benincasa hispida in transgenic tobacco OE12 and OE6 is significantly higher than that in wild - type tobacco plants, and the expression level of CqbZIP7 gene of Benincasa hispida in transgenic tobacco OE6 is higher than that in transgenic tobacco OE12.
[0097] 4. Physiological and biochemical indexes
[0098] Collect the leaves of the control (RT) and those treated with low - temperature (LT) for 24 hours, and measure the relative electrical conductivity (REC), as well as the activities of peroxidase (POD), polyamine oxidase (PAO) and the content of malondialdehyde (MDA) (kit, Solarbio). The above indexes are measured 3 times and the average value is taken.
[0099] After 24 hours of low - temperature treatment, compared with WT, OE6 and OE12, the relative electrical conductivity of the leaves is higher, indicating that over - expression reduces the damage and destruction of the cytoplasmic membrane structure of plants by low - temperature ( Figure 2 D in it); compared with RT, the enzyme activities of PAO and POD in LT increase significantly, and the MDA content decreases significantly. Compared with WT, the PAO activities of OE6 and OE12 increase by 69.9% and 50.8% respectively ( Figure 2 E in it), the POD activities of OE6 and OE12 increase by 35.3% and 82.4% respectively ( Figure 2 F in it), and the MDA contents of OE6 and OE12 decrease by 55.0% and 27.5% respectively ( Figure 2 G in it).
[0100] The results of this example show that overexpressing the CqbZIP7 gene of Benincasa hispida can improve the cold stress resistance of tobacco, and with the increase in the expression level of the CqbZIP7 gene of Benincasa hispida, the cold resistance of transgenic tobacco is also correspondingly improved. Therefore, the cold resistance of Benincasa hispida can be improved by overexpressing the CqbZIP7 gene of Benincasa hispida, and the cold-resistant germplasm resources of Benincasa hispida can be improved.
[0101] The technical features of the above-described embodiments can be combined arbitrarily. For the sake of concise description, not all possible combinations of the technical features in the above-described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, it should be considered that the scope described in this specification.
[0102] The above-described embodiments only express several implementation manners of the present invention, and the description is relatively specific and detailed, but it cannot be understood as a limitation on the scope of the invention patent. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present invention, several modifications and improvements can still be made, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the invention patent shall be subject to the appended claims.
Claims
1. Application of wax gourd CqbZIP7 gene in improving plant cold resistance, characterized in that, The nucleotide sequence of the wax gourd CqbZIP7 gene is shown in SEQ ID NO:1; the plant is wax gourd or tobacco.
2. Application of the CqbZIP7 gene of wax gourd in improving plant cold-resistant germplasm resources, characterized in that, The nucleotide sequence of the wax gourd CqbZIP7 gene is shown in SEQ ID NO:1; the plant is wax gourd or tobacco.
3. Use of the overexpression vector of the CqbZIP7 gene of Benincasa hispida in improving plant cold resistance or improving plant cold-resistant germplasm resources, characterized in that, The nucleotide sequence of the wax gourd CqbZIP7 gene is shown in SEQ ID NO:1; the plant is wax gourd or tobacco.
4. A method for improving the cold resistance of plants, characterized in that, Comprising the following steps: Enhancing the expression of the wax gourd CqbZIP7 gene in plants, the nucleotide sequence of the wax gourd CqbZIP7 gene is shown in SEQ ID NO:1; the plant is wax gourd or tobacco.