Cationic-Cl-co-transporter coding gene PcCCC1 of pugionium cornutum and application thereof

Abstract

The invention discloses a pugionium cornutum cation-Cl-co-transporter protein coding gene PcCCC1 and application thereof. The nucleotide sequence of the cation-Cl-co-transporter protein coding gene PcCCC1 is as shown in SEQ ID No.1, the amino acid sequence of the coding protein of the cation-Cl-co-transporter protein coding gene PcCCC1 is as shown in SEQ ID No.2, and the sequence of a specific promoter of the cation-Cl-co-transporter protein coding gene PcCCC1 is as shown in SEQ ID No.3. The invention further discloses an application of the cation-Cl-co-transporter protein coding gene PcCCC1. Experiments prove that after the gene is driven by a specific promoter of the gene to be overexpressed in arabidopsis thaliana, accumulation of Na < + > and Cl <-> of the overground part under salt stress can be reduced, so that the salt tolerance of the plant is improved, a gene resource is provided for improvement of the salt tolerance of the plant, and the gene has a wide application value.

Description

Technical Field

[0001] The present invention relates to the technical field of plant molecular biology, and in particular to a Psoralea cochinchinensis cation-Cl- cotransporter encoding gene PcCCC1 and an application thereof. Background Art

[0002] Drought poses a serious threat to the sustainable development of agriculture and animal husbandry and ecological environment construction in northern my country, especially in the northwest region. [1] In arid and semi-arid areas, irrigation has become one of the key measures to ensure agricultural production. [1] However, while this measure improves soil moisture conditions, it also causes deep salt to be transported to the surface along with water under the drive of strong evaporation, resulting in salt overload in the rhizosphere micro-domain, which in turn induces secondary soil salinization, ultimately having an adverse impact on the normal growth of resource plants such as crops and forage in arid areas. [2] It is worth noting that the drought-tolerant plants in desert ecosystems have evolved unique stress resistance and adaptation strategies through long-term natural selection. [3-5] Systematic analysis of the physiological and ecological mechanisms and molecular regulatory networks of these plants in adapting to adversity can provide excellent genetic resources for genetic improvement of high-quality forage and crop resistance. [4, 6] .

[0003] Sand mustard ( Pugionium cornutum ) belongs to the genus Psoralea of ​​the family Brassicaceae ( Pugionium ) is a typical succulent xerophyte, endemic to my country. [7] It is mainly distributed in the desert and semi-desert grassland areas of northern provinces such as Inner Mongolia, Gansu, and Ningxia. It has strong drought and salt tolerance and is an excellent sand-fixing pioneer plant. [8] Our previous study found that under salt treatment, sand mustard can regulate the Cl - and Na + The homeostatic balance of the osmotic regulator improves its osmotic regulation ability and maintains the charge balance in the cell. This mechanism plays an important role in the salt tolerance of sand mustard. [9, 10] Therefore, the Cl contained in sand mustard - and Na + Homeostasis-related gene resources can be used for genetic improvement of plant salt tolerance [9, 10] .

[0004] References: 1. Zhao Wenzhi, Ren Heng, Du Jun, Yang Rong, Yang Qiyue, Liu Hu. Some thoughts and suggestions on the ecological construction and agricultural development of the Hexi Corridor oasis. Bulletin of the Chinese Academy of Sciences, 2023, 38(03): 424-434. 2. Shan Lun. Scientific response to agricultural drought. Agricultural Research in Arid Areas, 2011, 29(2): 1-5. 3. Xie Wengang, Liu Zhipeng, Liu Gongshe, Chen Shuangyan, Hu Xiaowen, Qian Yongqiang, Yang Peizhi, Bai Shiqie, Yan Jiajun, Han Yunhua, Li Chunjie. Current status, key scientific issues and development directions of native grass breeding in my country. National Natural Science Foundation of China, 2023, 37(4): 552-559. 4.Chen X, Zhao CC, Yun P, Yu M, Zhou MX, Chen ZH,Shabala S. Climate-resilient crops: Lessons from xerophytes. Plant Journal, 2024, 117: 1815-1835. 5.Ashraf M. Inducing drought tolerance in plants: Recent advances. Biotechnology Advances, 2010, 28(1): 169-183. 6. Rawat N, Wungrampha S, Singla-Pareek SL, Yu M, Shabala S, Paree A. Rewilding staple crops for the lost halophytism: Toward sustainability and profitability of agricultural production systems. Molecular Plant, 2022, 15:45-64. 7. Zhao Yizhi. Endemic genera and basic characteristics of plants on the Mongolian Plateau. Journal of Inner Mongolia University (Natural Science Edition), 1997, 28(4): 547-552. 8. Wang Juyuan, Zhai Sheng, Hao Lizhen. Research on the current status of research and utilization and development strategy of the sand-dwelling plant Psoralea corylifolia. Agricultural Research in the Arid Areas, 2004, 22(4): 232-235. 9.Cui YN, Li XT, Yuan JZ, Wang FZ, Guo H, Xia ZR, Wang SM, Ma Q.Chloride is beneficial for growth of the xerophyte Pugionium cornutumbyenhancing osmotic adjustment capacity under salt and drought stresses. Journal of Experimental Botany, 2020, 71: 4215-4231. 10.Cui YN, Wang FZ, Yuan JZ, Guo H, Wang SM, Ma Q. Highconcentrations of sodium and chloride ions have opposing effects on the growth of the xerophyte Pugionium cornutum under saline conditions. Journal ofPlant Nutrition and Soil Science, 2021, 184: 88-97. Summary of the Invention

[0005] In view of the shortcomings of the prior art, the present invention aims to provide a sand mustard cation-Cl- cotransporter encoding gene PcCCC1 and its application, and obtain a sand mustard cation-Cl- cotransporter encoding gene PcCCC1 by screening from sand mustard. - The co-transporter encoding gene PcCCC1 was identified, and a plant expression vector was constructed in which its own specific promoter drives the expression of PcCCC1. The vector was then transferred into Arabidopsis for functional verification, and it was found that it can significantly improve the salt tolerance of the plant.

[0006] In order to achieve the above object, the technical solution adopted by the present invention is as follows: In the first aspect, the present invention proposes a Psoralea coparsis cation-Cl- cotransporter encoding gene PcCCC1, the key of which is that the nucleotide sequence of the cation-Cl- cotransporter encoding gene PcCCC1 is shown in SEQ ID No.1, and the amino acid sequence of the protein encoded by its full-length sequence is shown in SEQ ID No.2.

[0007] Furthermore, the self-specific promoter sequence of the cation-Cl- cotransporter encoding gene PcCCC1 is shown in SEQ ID No.3.

[0008] In a second aspect, the present invention further proposes the use of the sand mustard cation-Cl- cotransporter encoding gene PcCCC1 or the protein it encodes or its own specific promoter described in the first aspect in improving plant salt tolerance or in salt-tolerant plant breeding.

[0009] In the third aspect, the present invention proposes a plant expression vector, in which the specific expression of the PcCCC1 gene is driven by the self-specific promoter of the PcCCC1 gene, the nucleotide sequence of the PcCCC1 gene is shown in SEQ ID No.1, the amino acid sequence of the protein encoded by its full-length sequence is shown in SEQ ID No.2, and the sequence of its self-specific promoter is shown in SEQ ID No.3.

[0010] In a fourth aspect, the present invention proposes a use of the plant expression vector described in the third aspect in improving plant salt tolerance or in breeding salt-tolerant plants.

[0011] The significant effect of the present invention is: the present invention provides a sand mustard sand mustard cation-Cl - Co-transporter encoding genes PcCCC1 Experiments have shown that overexpression of this gene driven by its own specific promoter in Arabidopsis can reduce the Na + and Cl - The accumulation of genes significantly improves the salt tolerance of plants, provides genetic resources for improving the salt tolerance of plants, and has broad application value. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] Figure 1 The results are as follows: PcCCC1 Schematic diagram of changes in expression levels; Among them, A and B are the root and aerial parts after treatment with 25 mM NaCl for 0 h, 3 h, 6 h, 12 h, 24 h, and 48 h, respectively. PcCCC1 C and D are the relative expression levels of the roots and shoots after 0 h, 3 h, 6 h, 12 h, 24 h, and 48 h of treatment with 100 mM NaCl. PcCCC1 The relative expression level of Figure 2 yes PcCCC1 Heterologous overexpression lines PcCCC1 The expression level analysis results of Figure 3 After 7 days of treatment with 5 mM and 50 mM NaCl PcCCC1 Growth status of heterologous overexpression lines (O1 and O2) and wild type (WT); Figure 4 The control condition and NaCl treatment after 7 days PcCCC1 Comparison chart of growth indicators of heterologous overexpression lines (O1 and O2) and wild type (WT); Wherein, A is tissue fresh weight; B is tissue dry weight; C is aboveground water content; D is leaf relative membrane permeability; Figure 5 The control condition and NaCl treatment after 7 days PcCCC1 The Na content in roots and shoots of heterologous overexpression lines (O1 and O2) and wild type (WT) was significantly different. + and Cl - Concentration test result diagram; Where A is Na + concentration; B is Cl - concentration. DETAILED DESCRIPTION

[0013] The specific embodiments and working principles of the present invention are further described in detail below with reference to the accompanying drawings. The instruments, equipment, reagents, and test methods involved in the following examples are all conventional methods unless otherwise specified.

[0014] The seeds of the following examples were collected from Taole Town, Pingluo County, Shizuishan City, Ningxia Hui Autonomous Region, my country; The Arabidopsis wild type Col-0 described in the following examples was previously preserved by our research group.

[0015] Example 1: Sand mustard cation-Cl - Co-transporter encoding genes PcCCC1 Screening and cloning Using Psoralea cochinchinensis as the material, transcriptome and bioinformatics analysis revealed a cation-Cl- cotransporter gene, PcCCC1. Fluorescence quantitative PCR was further used to verify the expression pattern of this gene at different time periods under different salt treatments. It was found that under 25 mM NaCl treatment, the expression levels of PcCCC1 in both the shoots and roots of Psoralea cochinchinensis were significantly inhibited (see Figure 1 A and B); Under 100 mM NaCl treatment, the expression level of the gene PcCCC1 in the roots of P. serrata still decreased significantly, but the expression level of the gene PcCCC1 in the aerial parts of P. serrata was significantly increased. It can be seen that the expression of the gene PcCCC1 in the aerial parts of P. serrata is significantly induced by high concentration salt treatment (see Figure 1 C and D). The cation -Cl - The nucleotide sequence of the co-transporter encoding gene PcCCC1 is shown in SEQ ID No. 1, and the amino acid sequence of the protein encoded by the full-length sequence is shown in SEQ ID No. 2.

[0016] Example 2: Sand mustard cation-Cl - Acquisition of a self-specific promoter for the cotransporter encoding gene PcCCC1 The promoter of the gene PcCCC1 was identified based on the genome sequence of P. serrata. Amplification primers P1 and P2 were designed based on this sequence. The sequence of P1 is CACAGCGACACTAAGTATTTATTTG, and the sequence of P2 is TTCTCCCGCCGTATTTGGAC. Genomic DNA extracted from P. serrata leaves was used as a template for PCR amplification using the above primers. The PCR amplification protocol was as follows: 98°C for 30 s, followed by 30 cycles of 98°C for 10 s, 60°C for 30 s, and 72°C for 80 s, with an extension at 72°C for 10 min. A 2048-bp self-specific promoter of the gene PcCCC1 was successfully isolated. The nucleotide sequence is shown in SEQ ID No. 3.

[0017] Example 3: Sand mustard cation-Cl - Construction of a plant expression vector driving the expression of the cotransporter encoding gene PcCCC1 by its own specific promoter The PCR product of the PcCCC1 gene self-specific promoter (hereinafter abbreviated as ProPcCCC1) in "Example 2" was purified and ligated into the pMD19-T vector to obtain the pMD19-T-ProPcCCC1 vector. Using this vector plasmid as a template, PCR amplification was performed using primers P3 and P4, wherein the sequence of P3 is: TGATTACGCCAAGCTTCACAGCGACACTAAGTATTTATTTG, and the sequence of P4 is: GGACTGACCACCCGGGTTCTCCCGCCGTATTTGGAC. This yielded a PcCCC1-specific promoter fragment containing an EcoRI restriction site at the 5' end and a NocI restriction site at the 3' end. Simultaneously, the plant expression vector pCambia3301 plasmid was double-digested with the restriction endonucleases EcoRI and NocI to obtain a linearized pCambia3301 vector. Finally, the ProPcCCC1 fragment was ligated with the pCambia3301 linearized vector using In-Fusion HD clonase to construct the pCambia3301-ProPcCCC1 recombinant vector.

[0018] RNA was extracted from leaves of P. pumila (L.) and reverse transcribed to obtain cDNA. Using this cDNA as a template, PCR amplification was performed using primers P5 (sequence: GGCGGGAGAACCATGGATAGCGGCGATATTGAAGAAG) and P6 (sequence: CTGTAATTCACGTGCTATGTAAACAAAGTTACAACATCTCT). This yielded a PCR product containing the full-length coding sequence of PcCCC1, containing a NocI restriction site at the 5' end and a PmlI restriction site at the 3' end. The pCambia3301-ProPcCCC1 plasmid was double-digested with the restriction endonucleases NocI and PmlI to obtain a linearized vector. Finally, the PcCCC1 fragment was ligated with the linearized vector pCambia3301-ProPcCCC1 using In-Fusion HD Clonase, successfully constructing the plant expression vector pCambia3301-ProPcCCC1-PcCCC1, which expresses the PcCCC1 gene driven by its own specific promoter.

[0019] Example 4: Sand mustard cation-Cl - Functional verification of the cotransporter encoding gene PcCCC1 in improving plant salt tolerance The plant expression vector pCambia3301-ProPcCCC1-PcCCC1 obtained in Example 3 was transformed into Arabidopsis thaliana using the inflorescence infection method, and resistance screening was performed using 1‰ (v / v) Basta solution. After three consecutive generations of screening, T3 generation homozygous transgenic lines were obtained. The transcription level of the gene PcCCC1 in the root and aerial tissues of 10 lines was analyzed by RT-qPCR. The two lines with the highest expression levels (line 3 and line 10) were selected for subsequent research and named O1 and O2, respectively. Figure 2 shown.

[0020] Under the control and 5 mM NaCl treatments, there was no significant difference in the growth of transgenic lines O1 and O2 compared with the WT. After treatment with 50 mM NaCl, the growth of the aboveground parts of all lines was significantly inhibited, but O1 and O2 had relatively strong salt tolerance, and their growth state was significantly better than that of the wild type. In addition, the O2 line with a higher expression level of the gene PcCCC1 showed better growth than the O1 line, as shown in Figure 2. Figure 3 shown.

[0021] The results of biomass determination showed that under the control conditions and 5 mM NaCl treatment, there were no significant differences in the fresh weight, dry weight and relative water content of the roots and shoots between the transgenic lines (O1 and O2) and the WT. However, under 50 mM NaCl treatment, although there were no significant differences in the root biomass of each line, the fresh weight, dry weight and relative water content of the shoots of the transgenic lines were significantly higher than those of the WT (e.g. Figure 4 A, B, and C show that under control conditions and 5 mM NaCl treatment, there was no significant difference in the relative plasma membrane permeability of leaves of each strain; however, under 50 mM NaCl treatment, the relative plasma membrane permeability of WT leaves was significantly higher than that of transgenic strains O1 and O2 (as shown in Figure 2). Figure 4 D), indicating that salt stress caused more severe cell membrane damage in WT than in transgenic lines.

[0022] Under control conditions and 5 mM NaCl treatment, Na + and Cl - The concentrations were not significantly different from those of WT ( Figure 5 This indicates that under normal conditions and low salt concentration treatment, heterologous overexpression of the gene PcCCC1 does not affect the Na + and Cl - Under 50 mM NaCl treatment, the accumulation of Na in the aboveground part of the transgenic line + and Cl - The concentrations of Na in the aboveground parts of strains O1 and O2 were significantly lower than those in WT. + The concentrations of WT decreased by 26% and 45%, respectively ( Figure 5 A), Cl - The concentrations were significantly reduced by 51% and 64%, respectively ( Figure 5 B), while there was no significant difference in the concentrations of Na+ and Cl- in the roots of each strain ( Figure 5 A and B). It can be seen that under high salt stress, after the heterologous overexpression of the gene PcCCC1, the Na + and Cl - The accumulation was significantly reduced.

[0023] The above results indicate that overexpression of PcCCC1 in Arabidopsis thaliana driven by its own specific promoter can effectively improve plant salt tolerance. This suggests that the PcCCC1 gene encoding the cation-Cl- cotransporter from Arabidopsis thaliana described herein can significantly enhance plant salt tolerance, providing a gene resource for improving plant salt tolerance and possessing broad application value.

[0024] Sequence Listing SEQ ID NO.1 and SEQ ID NO.2 are the nucleotide sequence of the cation-Cl- cotransporter encoding gene PcCCC1 and the amino acid sequence of the encoded protein thereof, respectively, with sequence lengths of 2931 bp and 2928 bp, respectively; SEQ ID NO.3 is the sequence of the self-specific promoter of the cation-Cl- cotransporter encoding gene PcCCC1, as shown in SEQ ID No.3, with a sequence length of 2202 bp.

[0025] SEQ ID NO.1: ATGGATAGCG GCGATATTGA AGAAGCCCGGT GGTAACGGCG AAGAGAAATT CCGGTCAGGA 60 CCTCGTATCG GCGGAGGAAA ATACAGGCCA GTGGTGGCGC ACGATAGGGC GGTTGTCGAA 120 ATGTCTTCCA TCGATCCTGG ATCTTCCACG CTCAAGAACA TAAAAGTAGT TGCACCAGGA 180 GAGGTGAACG CTAGTGCTAG GGAGGGGCCA AGACCAGAAG ATGGAGTCAA TGGCCATCAG 240 AAGGAATCAA AGCTGGAATT ATTTGGTTTC GATTCTCTTG TTAACATTCT TGGTTTGAAG 300 AGTATGACAG GGGAGCAAAT TCCAGCACCA TCTAGCCCTA GAGATGGGGA GGATATCTCC 360 ATCACACAAG GGCACCCACA GCCTTCTGCA TTGAAGATGG GTACAATGAT GGGAGTTTTT 420 GTTCCCTGCT TGCAAAACAT ATTAGGAATT ATATACTATA TCCGTTTCAC ATGGATTGTT 480 GGCATGGCTG GTGTTGGACA AGGCCTAGTA CTCGTATTGC TCTGTGGCAC ATGTACATTC 540 TTGACAACCA TATCTTTGAG TGCTATTGCA ACAAATGGCG CCATGAAGGG TGGAGGACCA 600 TATTACCTCA TTGGTCGCGC TCTTGGTCCG GAGGTTGGGA TTAGCATAGG TTTATGCTTC 660 TTCCTTGGCA ATGCAGTTGC AGGAGCTCTG TACGTTTTGG GTGCTGTGGA GACTTTTCTA 721 AAAGCGTTCC CTGCTGCTGG GATTTTTCGA GAAACTATCA CAAAGGTTAA TGGAACTGCA 781 GTTGCTGAAT CAATACAAAG CCCAAACTCG CATGACTTGC AGATTTATGG AATAGTTGTA 841 ACTATCCTTC TATGCTTCAT TGTGTTTGGC GGCGTTAAGA TGATCAATCG GGTTGCACCT 900 GCTTTCCTCG TGCCCGTTTT GCTCTCCATC TTCTGCATAT TCATTGGGAT ATTTTTGGCA 960 AAGACAGATG ACCCTGACAC TGGAATTACG GGCTTGCGTT TAAAAAGTTT TAAAGATAAC 1020 TGGGGTTCTG CTTATCAGAT GACAAATCAC GCTGGAATTC CTGATCCAAC TGGAGACACG 1080 TACTGGAGTT TCAATGAGTT GGTGGGTCTA TTTTTCCCTG CTGTAACAGG AATTATGGCT 1140 GGCTCAAATC GATCAGCTTC GCTGAAAGAC ACACAAAGAT CAATTCCTCT TGGAACGCTG 1200 GCTGCCACTC TGACTACAAC CTTACTGTAT TTGATCTCTG TGTTGTTTTT TGGAGCTGTT 1260 GCGACCCGCA ACAAACTATT GACTGACAGG CTACTTACTG CTACAATTGC TTGGCCTTTC 1320 CCTGCAATTG TCCATGTTGG AATCATCCTT TCAACCTTAG GGGCTGCTCT CCAGAGTTTG 1380 ACAGGGGCCC CGAGGTTGCT TGCCGCTATA GCAAATGATG ATATTCTCCC CATCCTGAAT 1440 TATTTTAAAG TTGCAGATAC TAGCGAACCT CACATAGCAA CACTTTTTAC TGCATTGATT 1500 TGCATTGGAT GTGTTGTTAT CGGAAATTTG GATCTTATCA CACCGACTGT GACTATGTTT 1560 TATCTTTTAT GCTATGCGGG AGTAAACTTG TCCTGTTTCC TGCTCGATCT TCTTGATGCT 1620 CCCAGTTGGC GTCCGCGGTG GAAATATCAT CATTGGAGCC TTTCGTTTGT TGGAGCCTCA 1680 CTTTGCATAG TGATCATGTT CTTGATTTCT TGGTCATTCA CTGTGGTTGC CATTGCACTG 1740 GCAAGTCTTA TATACAAATA CGTTGGCTTA AAAGGGAAGG CCGGGGACTG GGGGGATGGT 1800 TTCAAGAGTG CATATTTTCA GTTGGCCCTT CGAAGTCTCA GGTCACTCGG AGCAAATCAA 1860 GTGCACCCCA AGAATTGGTA TCCAATTCCC CTTGTATTCT GCAGACCGTG GGGACAGCTC 1920 CCGGAGAATG TCCCGTGCCA TCCTAAGCTG GCTGATTTTG CTAACTGTAT GAAGAAAAAA 1980 GGTCGTGGAA TGTCGATTTT TGTCTCAATA TTAGATGGTG ACTATTATGA GTGTGCTGAA 2040 GAAGCAAAGG AAGCCTGCAA ACAACTAGCA ACCTACATTG AGTATAAGCG TTGTGAAGGT 2100 GTAGCTGAAA TCGTTGTAGC TCCTAACATG ACGGAAGGAT TTCGTGGGAT CATCCAGACC 2160 ATGGGACTCG GAAATCTCAA ACCCAACATT GTCGTAATGC GGTACCCAGA GATTTGGCGC 2220 CGAGAAAATC TAACAGAGAT TCCATCCACA TTTGTTGGGA TAATCAATGA CTGCATAACG 2280 GCAAACAAAG CAGTTGTCAT TATCAAAGGG TTAGACGAGT GGCCAAACGA GTACCAAAGA 2340 CAGTATGGAA CAATTGACTT ATACTGGATT GTGAGAGACG GTGGGTTCAT GCTTCTTCTG 2400 TCACAGCTGC TTCTCACAAA AGAGAGCTTT GAGAGCTGCA AAATCCAACT CTTCTGCATA 2460 GCCGAAGAGG ATTCAGATGC GGAAGCATTG AAAGCGGACG TGAAGAAGTT CCTCTACGAT 2520 CTCAGAATGC AAGCCGAAGT AATTGTGGTC AAGATGAAT CATGGGATAT AAGATCTGAA 2580 GGAAACAGCC AAGAAGATTC ATTGGAAGCT TTTGATGCTG CACAGAGACG AATCTCAGAT 2640 TACTTGGGAG AGATCAAGAG ACAAGGTTCG ACTCCGTTGC TGGCGAATGG GAAACCGATG 2700 GTGGTGAATG AGCAACAAGT GGAGAAGTTT CTATACACGA TGCTAAAACT GAACTCGACC 2760 ATACTCAGTT ACTCTAGGAT GGCTGCGGTT GTTCTTGTTA GTCTCCCTCC ACCACCATTG 2820 AACCACCCGG CATATTTCTA TATGGAATAT ATGGATTTGC TGGTGGAGAA TGTACCAAGG 2880 ATGTTGATCG TAAGAGGGTA TCACAGAGAT GTTGTAACTT TGTTTACATA G 2931 SEQ ID NO.2: Met Asp Ser Gly Asp Ile Glu Glu Ala Gly Gly Asn Gly Glu Glu Lys Phe Arg SerGly 20 Pro Arg Ile Gly Gly Gly Lys Tyr Arg Pro Val Val Ala His Asp Arg Ala Val Val ValGlu 40 Met Ser Ser Ile Asp Pro Gly Ser Ser Thr Leu Lys Asn Ile Lys Val Val Ala ProGly 60 Glu Val Asn Ala Ser Ala Arg Glu Gly Pro Arg Pro Glu Asp Gly Val Asn Gly HisGln 80 Lys Glu Ser Lys Leu Glu Leu Phe Gly Phe Asp Ser Leu Val Asn Ile Leu Gly LeuLys Ser Met Thr Gly Glu Gln Ile Pro Ala Pro Ser Ser Pro Arg Asp Gly Glu Asp IleSer 120 Ile Thr Gln Gly His Pro Gln Pro Ser Ala Leu Lys Met Gly Thr Met Met Gly ValPhe 140 Val Pro Cys Leu Gln Asn Ile Leu Gly Ile Ile Tyr Tyr Ile Arg Phe Thr Trp IleVal 160 Gly Met Ala Gly Val Gly Gln Gly Leu Val Leu Val Leu Leu Cys Gly Thr Cys ThrPhe 180 Leu Thr Thr Ile Ser Leu Ser Ala Ile Ala Thr Asn Gly Ala Met Lys Gly Gly GlyPro 200 Tyr Tyr Leu Ile Gly Arg Ala Leu Gly Pro Glu Val Gly Ile Ser Ile Gly Leu CysPhe 220 Phe Leu Gly Asn Ala Val Ala Gly Ala Leu Tyr Val Leu Gly Ala Val Glu Thr PheLeu 240 Lys Ala Phe Pro Ala Ala Gly Ile Phe Arg Glu Thr Ile Thr Lys Val Asn Gly ThrAla 260 Val Ala Glu Ser Ile Gln Ser Pro Asn Ser His Asp Leu Gln Ile Tyr Gly Ile ValVal 280 Thr Ile Leu Leu Cys Phe Ile Val Phe Gly Gly Val Lys Met Ile Asn Arg Val AlaPro 300 Ala Phe Leu Val Pro Val Leu Leu Ser Ile Phe Cys Ile Phe Ile Gly Ile Phe LeuAla 320 Lys Thr Asp Asp Pro Asp Thr Gly Ile Thr Gly Leu Arg Leu Lys Ser Phe Lys AspAsn 340 Trp Gly Ser Ala Tyr Gln Met Thr Asn His Ala Gly Ile Pro Asp Pro Thr Gly AspThr 360 Tyr Trp Ser Phe Asn Glu Leu Val Gly Leu Phe Phe Pro Ala Val Thr Gly Ile MetAla 380 Gly Ser Asn Arg Ser Ala Ser Leu Lys Asp Thr Gln Arg Ser Ile Pro Leu Gly ThrLeu 400 Ala Ala Thr Leu Thr Thr Thr Leu Leu Tyr Leu Ile Ser Val Leu Phe Phe Gly AlaVal 420 Ala Thr Arg Asn Lys Leu Leu Thr Asp Arg Leu Leu Thr Ala Thr Ile Ala Trp ProPhe 440 Pro Ala Ile Val His Val Gly Ile Ile Leu Ser Thr Leu Gly Ala Ala Leu Gln SerLeu 460 Thr Gly Ala Pro Arg Leu Leu Ala Ala Ile Ala Asn Asp Asp Ile Leu Pro Ile LeuAsn 480 Tyr Phe Lys Val Ala Asp Thr Ser Glu Pro His Ile Ala Thr Leu Phe Thr Ala LeuIle 500 Cys Ile Gly Cys Val Val Ile Gly Asn Leu Asp Leu Ile Thr Pro Thr Val Thr MetPhe 520 Tyr Leu Leu Cys Tyr Ala Gly Val Asn Leu Ser Cys Phe Leu Leu Asp Leu Leu AspAla 540 Pro Ser Trp Arg Pro Arg Trp Lys Tyr His His Trp Ser Leu Ser Phe Val Gly AlaSer 560 Leu Cys Ile Val Ile Met Phe Leu Ile Ser Trp Ser Phe Thr Val Val Ala Ile AlaLeu Ala Ser Leu Ile Tyr Lys Tyr Val Gly Leu Lys Gly Lys Ala Gly Asp Trp Gly AspGly 600 Phe Lys Server To Tyr Phe Gln Leu To Arg Ser Leu Arg Served To Gly To AsnGln 620 Val His Pro Lys Asn Trp Tyr Pro Ile Pro Leu Val Phe Cys Arg Pro Trp Gly GlnLeu 640 Pro Glu Asn Val Pro Cys His Pro Lys Leu Ala Asp Phe Ala Asn Cys Met Lys LysLys 660 Gly Arg Gly Met Ser Ile Phe Val Ser Ile Leu Asp Gly Asp Tyr Tyr Glu Cys AlaGlu 680 Glu Ala Lys Glu Ala Cys Lys Gln Leu Ala Thr Tyr Ile Glu Tyr Lys Arg Cys GluGly 700 Val Ala Glu Ile Val Val Ala Pro Asn Met Thr Glu Gly Phe Arg Gly Ile Ile GlnThr 720 Met Gly Leu Gly Asn Leu Lys Pro Asn Ile Val Val Met Arg Tyr Pro Glu Ile TrpArg 740 Arg Glu Asn Leu Thr Glu Ile Pro Ser Thr Phe Val Gly Ile Ile Asn Asp Cys IleThr 760 Ala Asn Lys Ala Val Val Ile Ile Lys Gly Leu Asp Glu Trp Pro Asn Glu Tyr GlnArg 780 Gln Tyr Gly Thr Ile Asp Leu Tyr Trp Ile Val Arg Asp Gly Gly Leu Met Leu LeuLeu 800 Ser Gln Leu Leu Leu Thr Lys Glu Ser Phe Glu Ser Cys Lys Ile Gln Leu Phe CysIle 820 Ala Glu Glu Asp Ser Asp Ala Glu Ala Leu Lys Ala Asp Val Lys Lys Phe Leu TyrAsp 840 Leu Arg Met Gln Ala Glu Val Ile Val Val Thr Met Lys Ser Trp Asp Ile Arg SerGlu 860 Gly Asn Ser Gln Glu Asp Ser Leu Glu Ala Phe Asp Ala Ala Gln Arg Arg Ile SerAsp 880 Tyr Leu Gly Glu Ile Lys Arg Gln Gly Ser Thr Pro Leu Leu Ala Asn Gly Lys ProMet 900 Val Val Asn Glu Gln Gln Val Glu Lys Phe Leu Tyr Thr Met Leu Lys Leu Asn SerThr 920 Ile Leu Ser Tyr Ser Arg Met Ala Ala Val Val Leu Val Ser Leu Pro Pro Pro ProLeu 940 Asn His Pro Ala Tyr Phe Tyr Met Glu Tyr Met Asp Leu Leu Val Glu Asn Val ProArg 960 Met Leu Ile Val Arg Gly Tyr His Arg Asp Val Val Thr Leu Phe Thr * 976 SEQ ID NO.3: CACAGCGACA CTAAGTATTT ATTTGCTTCT TTCTTATTTT GATGATTTAC AGCATCATCC 60 GTGGTTAGCG GAAACAGAAA GAGAGAATCT TTGCAGGTTG TTAGATTGTC AGAAACTCTC 120 TCTAGAAGCT TGCACACACG CTGCACAGAA CGAGAGATTG CCACTAAGAA TAATCGTCCA 180 AGTCCTCTTC TTTGAGCAGC TTCAGCTCAG GACCTCTGTA GCTGGATGCT TCCTGGTTTC 240 AGACAACCTA GATGGGGAAT CAAGACAGCT AAGAAGCGGA GGATTTGTAG GAGGATCAAC 300 CGAAGGAGGA GGTGGATGGG CAACCGCAGT GAGAGAGAAT CAAGTCCTGA AAGTTGGAAT 360 GGACAGTATG AGAATGCGAG TTTGTGAGTT AGAGAAAGAA TGTTCCAATA TGAGACAAGA 420 GATTGAGAAA CTCGGTAAGA CGACAAAAGG TGGTGGTTCT GGAAGCGGTG GAGGAGGTAG 480 CAAAACGTGG GAAAACGTTT CTAAGAAACT CGGGTTTGGT TTTAAGCTTA AGTCTCATCA 540 AATGTGCAGT GCTCAAGAAG GATCTGTGTC TAAGTCTAAC AATGAGAATG TGAAGATAGA 600 GAAGCTAAAA GATGTCAAAG AACGTCGTGA GAAGCATAAG AAAGCCTCGA GCATTAGTTC 660 TGACAGGTGA AAATGGTAAC TTTCCACTGT TAATTGTGCA GTTACAATGT TTGGGATTGT 720 TTAGCTAAAA AATGGAGCTT TCCGTTGTTA AATGTGTGCA TTGTAGTGAA ATGGTGCTCT 780 CATTTTTTTT TGTTTACAAT TGTTTATGTA TCTTTGACGA TCAACTGTGT GTTATTATTG 840 TTTTTAGTTT AACTTTTTTA AAACTATAAA AGAAAAAAAA AACAAATATT GCTGGTAACC 900 TATGGCGAAT CTATCAACTG TTTCTTAACA ACGCCAAATC TTAAGGATAA GAGAATTACA 960 AGATTGATAA CATTTTTTGT TAATTGAAAT GTGTTTTTGT TTTCATTGAG AAGCTCCTTT 1020 TTTCTCATTT CTCATTCATC TATCAGAGAA TTTTCAAATT TCTGAATATT CTTATTGGGA 1080 AGTTTCAAAT ATTATTCATG ATTTAATAAA GCCTTTAATT AATTTTTTTT CCTTCAGCCT 1140 TTAATTAACG CGAATCCAAA ATTTCTGTTT TTGCCCTAAA TAAAAGATAG TTTTCAATGG 1200 AAAAATAAAA ATGAATATTT GGTTATTCAG TTATTTTTTT CTATTATTTT TGGGTTTAAA 1260 AATTTTTTTC TCTCTAAAAA TAAGTTTTAT TTGGATTCAG TTAATACTTG ATTTCGGTTT 1320 GTTTCAGTTC AAGTTTTGTA TTATATTG GTTTTGTTTG ATTTTAAATA AAATAAAAA 1380 TATATATT TTTTTCCAAC ATTACTTA TTAATGTAAT AAGTCATTGT TTATAGT 1440 TCTTAAAAAA TTTGGACAAA HELP TTCCCAATCT ACATTTAGCT GATTTAGTT 1500 AGTATATGAG ATTTTATG TGTTAATAAG TTTTTTTATT TGTTTAATTA AAAATATTAG 1560 TTGTTTTAGA ATTGGTTA ATTTGGTGAA GTTTAAATTT GATTTGATTT GAAATCAGTT 1620 AGTTCAAATT AATTTTTA AAATTTTATT TGGTTTTA TTATAATTCG GTATAAAATA 1680 TAGTTTGGAA AATTTTAGCC GGATTTAGGT TTAAAAATTTT TAGTTTGGTT CAGTTCGGAT 1740 TCAGAAATTT TAGATTCCGG TTGAAATT TTAGTTTGCC CAACTCTAGA AGGAAATATG 1800 ASSOCIATETT CTGROUP ATTGTTCAAA TTGCTTCAAT ATTGCTGCTATGTA 1860 AAAGGTTAA TCTAATAT CTTCAAGATT CACTATT AAAGACTTATT ATGCACTATT 1920 ATTAAAGGAC ATTTCAGCCG TCAAGTTGAC CCAATTTAAA AGCCAAATT ACACTTTAAT 1980 TTTTCTCTTT TTCTTAATTT CTTCAGTAAT ACTTTGTCTG AAATCTGATT TCGTTTCTTT 2040 ATTTTTTTCTT ATGAATTTCT CTATTTTTTG CCGTTAATTC CTCGAGTCTC GATCTCAGTT 2100 CTCGTCAGAT TTCCATCTCT CCGATCTCTA ATTCCGTTGA GATATCCCTC TCATTGTTGG 2160 AAGCAGGATT AAACGGATCT GTCCAAATAC GGCGGGAGAA CC 2202 The technical solution provided by the present invention is introduced in detail above. Specific examples are used herein to illustrate the principles and implementation methods of the present invention. The description of the above embodiments is only used to help understand the method of the present invention and its core idea. It should be pointed out that for ordinary technicians in this technical field, without departing from the principles of the present invention, the present invention can also be improved and modified in a number of ways, and these improvements and modifications also fall within the scope of protection of the claims of the present invention.

Claims

1. A gene encoding a cation-Cl- cotransporter from Psoralea corylifolia, PcCCC1, characterized in that: The nucleotide sequence of the cation-Cl- cotransporter encoding gene PcCCC1 is shown in SEQ ID No. 1, and the amino acid sequence of the protein encoded by its full-length sequence is shown in SEQ ID No.

2.

2. The Psoralea coparsis cation-Cl- cotransporter encoding gene PcCCC1 according to claim 1, characterized in that: The sequence of the self-specific promoter of the cation-Cl- cotransporter encoding gene PcCCC1 is shown in SEQ ID No.

3.

3. Use of the Psoralea coparsis cation-Cl- cotransporter encoding gene PcCCC1 or the protein encoded by it or its own specific promoter as claimed in claim 1 or 2 in improving plant salt tolerance or in salt-tolerant plant breeding.

4. A plant expression vector, characterized in that The specific expression of the PcCCC1 gene is driven by its own specific promoter. The nucleotide sequence of the PcCCC1 gene is shown in SEQ ID No. 1, the amino acid sequence of the protein encoded by its full-length sequence is shown in SEQ ID No. 2, and the sequence of its own specific promoter is shown in SEQ ID No.

3.

5. Use of the plant expression vector according to claim 4 in improving plant salt tolerance or in breeding salt-tolerant plants.

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