CN110129291B - Barley moisture-resistant regulatory gene HvACO1, protein and application thereof in breeding - Google Patents
Barley moisture-resistant regulatory gene HvACO1, protein and application thereof in breeding Download PDFInfo
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Abstract
The invention discloses a barley moisture-resistant regulatory gene HvACO1, protein and application thereof in breeding. Barley moisture-resistant geneHvACO1The CDS sequence of (1) is shown in SEQ ID NO. The invention clones the moisture-resistant gene from the moisture-resistant barley for the first timeHvACO1And proving that the gene is related to the moisture resistance of barley, adopting a genetic engineering method to performHvACO1The gene is over-expressed into arabidopsis, and compared with a control group, the moisture resistance of a transgenic plant is obviously enhanced, which shows thatHvACO1The over-expression of the gene improves the moisture resistance of the plant. In the inventionHvACO1The techniques of gene cloning and transformation, transgenosis and the like lay a foundation for researching barley moisture-resistant molecular mechanism and breeding application, and have wide breeding application prospect and certain economic value.
Description
Technical Field
The invention belongs to the technical field of genetic engineering, and particularly relates to a barley moisture-resistant regulatory geneHvACO1And breeding application thereof.
Background
The wet damage is the damage to the normal growth and development of crops when the soil is saturated or supersaturated with water. The damp damage occurs worldwide, especially in North America, Australia, China, India and other countries. According to data published by food and agriculture organizations in the united nations, about 10% of the planting area worldwide is affected by the moisture damage. The barley dry crops and the wet damage seriously affect the yield and the quality of the barley. The wet damage can generally reduce the yield of the barley by 20 to 30 percent, and even cause severe or no harvest.
The stress of wet injury hinders the free circulation of gas in plant rhizosphere tissues and influences the photosynthesis and respiration of plants. The plant can regulate and control programmed cell death and formation of new adventitious roots and ventilated tissues by synthesizing the biotin such as ethylene, and the gene related to ethylene synthesis has a regulating effect on improving the moisture resistance of the plant. Ethylene synthase (ACS) and ethylene oxidase (ACO) are two key enzymes in the ethylene synthesis pathway, and research has shown that cotton is processedGhACOThe gene is transferred into the arabidopsis thaliana,GhACOthe gene expression enhances the biosynthesis of ethylene in arabidopsis thaliana, and further enhances the tolerance of arabidopsis thaliana to abiotic stress. In recent years, many reports have been made about cloning and breeding utilization of stress-resistance related genes of crops, such as moisture resistance, salt resistance, drought resistance and the like, and a new technical approach is provided for breeding stress-resistance crop varieties. At present, the research on barley moisture-resistant related genes mainly focuses on QTL positioning level, and no research report on barley moisture-resistant gene cloning and breeding application is found.
Disclosure of Invention
The invention aims to provide a barley moisture-resistant regulatory geneHvACO1And its breeding application, cloning and constructionHvACO1The gene expression vector is introduced into arabidopsis thaliana by using a transgenic technology, and the transgenic arabidopsis thaliana is subjected to wet damage stress treatment and verifiedHvACO1The moisture-proof function of the barley provides gene resources and theoretical basis for improving moisture-proof varieties of barley.
In order to achieve the purpose of the invention, the technical scheme adopted by the invention is as follows: a barley moisture resistance regulation related protein is (a) or (b) as follows:
(a) a protein consisting of an amino acid sequence shown as SEQ ID NO. 2; the specific sequence is MEIPVIDLQGLDGDASQRSQTMARLHEACKDWGFFWVDSHGVDAALMEEVKRFVYAHYDEHLKDRFYASDLAKDLLLPAEESKAVSGEVDWETAYFIRHRPANNVADFPEIPPATREMLDVYIGQMVSLAERLAECMSLNLGLDGGRVKDTFAPPFVGTKFAMYPACPRPDLLWGLRAHTDAGGIILLLQDDVVGGLEFFRGDREWVPVGPTKGSRIFVNLGDQLEVMSGGAYRSVLHRVAAVAEGRRLSVATFYNPGAEAVVAPAPTARQPAAQVYPGPYRFGDYLDYYQGTKFADKAARLQAVKELFGSRILPD;
(b) a protein which is derived from the amino acid sequence of SEQ ID NO. 2 by substituting and/or deleting and/or adding one or more amino acid residues of the amino acid sequence of SEQ ID NO. 2 and is related to barley moisture resistance regulation.
The invention also provides a gene encoding the protein. The gene is a DNA molecule of any one of the following (a 1) - (a 3);
(a1) 1, DNA molecule shown in SEQ ID NO;
(a2) a DNA molecule which is hybridized with the DNA sequence limited by (a 1) under strict conditions and codes barley humidity-resistant regulation related protein;
(a3) a DNA molecule which has at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% homology with the DNA sequence defined in (a 1) and encodes a barley moisture-resistance regulation-related protein.
The invention also provides an expression cassette, a recombinant vector, a recombinant microorganism or a transgenic cell line containing the gene.
It is another object of the present invention to provide the use of (b 1) or (b 2) or (b 3):
(b1) the protein, or the gene, or an expression cassette, a recombinant vector, a recombinant microorganism or a transgenic cell line containing the gene is applied to the resistance to barley wet damage stress;
(b2) the protein, or the gene, or an expression cassette, a recombinant vector, a recombinant microorganism or a transgenic cell line containing the gene is applied to cultivating a new moisture-resistant variety of barley and arabidopsis thaliana;
(b3) the protein, or the gene, or an expression cassette, a recombinant vector, a recombinant microorganism or a transgenic cell line containing the gene is applied to the resistance to arabidopsis thaliana wet damage stress.
The invention also provides a method for cultivating the plant moisture-proof variety, which comprises the steps of transferring the gene into a target plant, or transferring the target plant into the expression cassette and the recombinant vector to obtain a transgenic plant; the transgenic plant has higher wet damage stress resistance than the target plant. The target plants are barley and arabidopsis thaliana. The transgenic plant expresses the barley moisture-resistance related protein.
HvACO1The DNA fragment of the gene is shown in a sequence table SEQ ID NO.1, or is basically equivalent to the DNA sequence shown in the SEQ ID NO.1, or is a partial fragment of the sequence shown in the SEQ ID NO.1 in function. The sequence analysis of the gene shows thatHvACO1The full length of the gene coding region is 952bp, 317 amino acids are coded, the molecular weight is 35.02kDa, the isoelectric point is 5.16, and the gene coding region contains conserved D10x-N and 20G-FelI-Oxy structural domains. The over-expression sequence shown in the sequence table SEQ ID NO.1 can enhance the tolerance of plants such as arabidopsis thaliana and barley to damp damage.
The gene can be applied to improving the moisture resistance of barley, and the specific operation is as follows:
(1) obtaining of genes: proteomics analysis on the root system of the barley moisture-proof material TF58 shows that moisture damage stress causes the root systemHvACO1The gene is obviously up-regulated and expressed, total RNA of TF58 root system is extracted and amplified by RT-PCRHvACO1And gene sequence, connecting the amplified product to pGEM-Teasy vector, and obtaining target gene clone through sequencing.
(2) Construction and genetic transformation of expression vectors: construction of barley Using Gateway technology from Invitrogen corporationHvACO1A gene overexpression vector. The target gene is linked to pDONR221 vector by BP clone enzyme to construct entry vector, and the entry vector is linked to pB2GW7 vector by LR clone enzyme to construct entry vectorHvACO1A gene overexpression vector. Freezing and thawing by liquid nitrogenHvACO1The gene over-expression vector is introduced into agrobacterium Gv 3101. By utilizing agrobacterium-mediated genetic transformation methodHvACO1The gene is introduced into Arabidopsis thaliana and expressed. Positive transgenic Arabidopsis plants are screened by antibiotic screening, RT-PCR and the like.
(3) Moisture resistance identification of transgenic plants andHvACO1and (3) gene function verification: transgenic and wild type lines that grew normally for 35 days were flooded. After 2 weeks of flooding, the moisture tolerance of the transgenic plants was observed to be significantly better than that of the wild type plants, thus demonstrating thatHvACO1The gene can obviously improve the moisture resistance of arabidopsis thaliana.
The invention provides a new method for improving the resistance of plants to the wet damage. The moisture-resistant plant is cultivated by a genetic engineering means, the defects of the traditional breeding are overcome, the breeding period is short, the operation is simple, and the high-resistance material is easy to obtain. The invention clones the barley moisture-resistant gene for the first timeHvACO1The gene is transferred into arabidopsis thaliana by an agrobacterium-mediated method, and the moisture damage identification and analysis prove that the moisture resistance of a transgenic plant is obviously improved compared with that of a wild plant, and the gene has wide application prospect in the aspect of improving the moisture resistance of barley.
Drawings
FIG. 1 shows the RT-PCR amplification product of barley HvACO1 gene sequence,
marker: DL2000DNA Marker (Dalianbao biology) composed of six DNA fragments of 2,000bp, 1,000bp, 750bp, 500bp, 250bp and 100 bp;
FIG. 2 shows a rotary drumHvACO1RT-PCR detection analysis of gene Arabidopsis thaliana strain and wild type strain;
FIG. 3 shows the variation of moisture resistance expression and partial growth indexes of transgenic Arabidopsis lines and wild type lines,
(A) phenotype of transgenic arabidopsis thaliana and wild type under normal growth conditions and after 2 weeks of flooding; (B) the plant height; (C) fresh weight of stem leaves; (D) dry weight of stem leaves; (E) root length; (F) survival rate.
Detailed Description
The present invention will be described in further detail with reference to specific examples.
Example 1HvACO1Cloning of genes
The barley strain TF58 is usedAnalysis of proteome results, designHvACO1Specific primer P for gene1Forward primers 5'-ATGGACATGGAGATCCCGGT-3' (SEQ ID NO. 3) and P25'-TCAATCGGGCAGAATCCGTG-3' (SEQ ID NO. 4), extracting the moisture-proof barley strain TF58 root system total RNA by CTAB method, reverse transcribing into cDNA, using cDNA as template, using primer P1And P2The CDS sequence of the barley moisture-resistant gene shown as SEQ ID NO.1 is amplified, and the geneHvACO1The full length of the CDS sequence of (2) 952bp (FIG. 1).
The method comprises the following specific steps:
(1) adding a CTAB (hexadecyl trimethyl ammonium bromide) extraction buffer solution [ 2% (W/V) CTAB, 1.4 mol/L of NaCl, 20mmol/L of EDTA (ethylene diamine tetraacetic acid), 100mmol/L of Tris-HCl, 2% (W/V) PVP ] and 10% beta-mercaptoethanol into a centrifugal tube, and preheating in a water bath kettle;
(2) cooling and grinding barley root system with liquid nitrogen, adding into the extract, mixing, and water bathing at 65 deg.C for 10 min;
(3) equal volume of chloroform was added: mixing isoamyl alcohol (volume ratio 24: 1), reversing, mixing, standing for 10min, and centrifuging at 4 ℃ at 12000g for 10 min;
(4) taking the supernatant, and repeating the step (3);
(5) taking the supernatant, adding LiCl with the final concentration of 2mol/L, carrying out ice bath for 10-12 hours, centrifuging for 15min at the temperature of 4 ℃ at the rpm of 11000rpm, discarding the supernatant, washing the precipitate twice by using 75% ethanol, and dissolving the precipitate in an appropriate amount of DEPC (diethyl pyrocarbonate) treatment water for later use;
(6) total RNA of root system extracted from the barley variety TF58 was used as a template and was reverse transcribed to synthesize the first strand cDNA using reverse transcriptase (purchased from Thermo Fisher Scientific Co.) under the following conditions: 5min at 65 ℃, 50min at 42 ℃ and 10min at 70 ℃;
(7) the barley ethylene oxidase gene was amplified from cDNA reverse-transcribed from RNA using the above primers P1 and P2HvACO1The CDS sequence of (a);
reaction conditions are as follows: pre-denaturation at 94 ℃ for 4 min; 30sec at 94 ℃, 30sec at 55 ℃, 1min at 72 ℃ and 33 cycles; extension at 72 ℃ for 10 min. The PCR product obtained by amplification was ligated with pMD18-T vector (purchased from Takara Bio Inc.), and the large intestine was transformedThe bacillus is infected with the competent cell, and positive clones are screened and sequenced to obtain the required full-length gene. Extracting the carrier from the positive cloneHvACO1Plasmid of gene CDS sequence.
Example 2HvACO1Construction and genetic transformation of gene overexpression vector
To better analyze the geneHvACO1The gene was overexpressed in Arabidopsis thaliana, and barley was constructed by Gateway technology of Invitrogen corporationHvACO1A gene overexpression vector. A target gene is connected with a pDONR221 vector by using a BP clone enzyme to construct an entry vector, and then the entry vector is connected with a pB2GW7 vector by using an LR clone enzyme to construct a super-expression vector containing the target gene. Freezing and thawing with liquid nitrogenHvACO1The overexpression vector of the gene is introduced into agrobacterium Gv 3101. By utilizing agrobacterium-mediated genetic transformation methodHvACO1The coding sequence is introduced into Arabidopsis thaliana and other plants for expression. Screening positive transgenic plants by antibiotic screening, RT-PCR and the like.
The practical genetic transformation method of the test is as follows:
(1) cultivation of Agrobacterium
Pre-culture of solid LB medium (10g/L peptone, 5g/L yeast extract, 10g/L sodium chloride, Kan 100mg/L agar 1.5g/L) with corresponding resistance selection carrying the gene of interestHvACO1The culture temperature of the agrobacterium tumefaciens is 28 ℃ for 48 hours; selecting a single colony of the pre-cultured agrobacterium, inoculating the single colony into a liquid LB culture medium (10g/L peptone, 5g/L yeast extract, 10g/L sodium chloride and Kan 100mg/L) of corresponding resistance selection, and culturing the single colony overnight in a shaking table at the temperature of 28 ℃ and the rpm of 200 until the OD600 value of the bacterial liquid concentration is about 0.8-1.0.
(2) Inflorescence method for infecting arabidopsis
Arabidopsis thaliana was transformed by modified inflorescence dip-staining. Selecting an agrobacterium tumefaciens single colony containing a target gene plasmid, transferring the agrobacterium tumefaciens single colony into 100mL of LB liquid culture medium, putting the 100mL of LB liquid culture medium into a shaking table at 28 ℃ for overnight culture at 200r/min, centrifuging the mixture for 10min at 4000r/min when the concentration of the agrobacterium tumefaciens bacterial liquid reaches OD600 of 0.8-1.0, discarding supernatant, and then re-suspending the agrobacterium tumefaciens bacterial liquid by 100mL of LB liquid culture medium containing 5g of cane sugar and Silwet L-7750 mu L. And (2) inversely buckling an arabidopsis flower pot in a beaker containing bacterial suspension, soaking the inflorescence of the arabidopsis into bacterial liquid for 10-20 s, culturing for 24h in the dark, placing the arabidopsis flower pot in an illumination incubator with the temperature of 25 ℃, the humidity of 60%, the illumination intensity of 3000-4000 lx and the 16h/8h photoperiod for culturing, watering and culturing plants every other day, and harvesting seeds after blooming. Harvested Arabidopsis seeds were sterilized in 8% NaClO alcohol solution (prepared in 95% alcohol) for 5min, and then screened on a selection medium (MS +50mg/L kanamycin +7g/L agar +30g/L sucrose).
Example 3HvACO1RT-PCR detection of gene transgenic T3 generation seedling
To verify whether the transgenic Arabidopsis thaliana T3 lines have altered ability to resist moistureHvACO1Gene correlation, using RT-PCR method to do partial transgenic arabidopsis plantHvACO1The gene expression is detected, the result is shown in figure 2, and the change of the moisture resistance of transgenic arabidopsis T3 strain and the transferred transgenic arabidopsis T3 strain can be knownHvACO1Overexpression of the gene is involved.
The method comprises the following specific steps:
total RNA of plants was extracted from 3T 3-generation strains of transgenic Arabidopsis thaliana using TRIZOL reagent (available from Byobo bioengineering, Dalian, Ltd.) (the extraction method was described in reference to TRIZOL reagent instructions), and reverse-transcribed to synthesize cDNA first strand using reverse transcriptase (available from Thermo Fisher Scientific Co., Ltd.) under conditions of 5min at 65 ℃, 50min at 42 ℃ and 10min at 70 ℃. Firstly, the cDNA obtained by reverse transcription is detected and the concentration is adjusted by using the reported internal reference gene Actin, and PCR detection is carried out, so that the internal reference genes can be amplified in the plants of contrast wild type arabidopsis thaliana and transgenic arabidopsis thaliana. Then, RT-PCR detection was performed using primers P1 and P2 based on the sequence of HvACO1 gene under the reaction conditions: pre-denaturation at 94 ℃ for 4 min; 30sec at 94 ℃, 30sec at 55 ℃, 1.5 min at 72 ℃ and 33 cycles; extension at 72 ℃ for 10 min. The agarose gel electrophoresis results of the amplified products show (FIG. 2), and the expression of HvACO1 gene was detected in all 3 transgenic lines.
Example 4 transferHvACO1Determination of moisture resistance of Arabidopsis plants
Obtained from screeningHvACO1Transgenic Arabidopsis thaliana T3 generation positiveRandomly selecting seeds of each 3 transgenic lines, culturing on 1/2MS culture medium to 4-leaf stage with wild type Arabidopsis thaliana seed control (WT), transplanting the seedlings into pots filled with nutrient soil, and culturing in artificial climate chamber for 35 days. Putting the transgenic plant line and the wild type into a water tank, performing flooding treatment, observing the difference of moisture resistance of the transgenic material and the wild type material after 2 weeks of flooding, and measuring the characters of different materials, such as seedling height, chlorophyll content, fresh weight, dry weight, root length, survival rate and the like. The wild plant height is reduced by 49.09%, and the height of 3 transgenic lines is reduced by 11.70%, 11.40% and 10.28% respectively; the wild chlorophyll content is reduced by 61.56 percent, and the content of 3 transgenic lines is reduced by 20.45 percent, 31.82 percent and 34.23 percent respectively; the fresh weight of wild stems and leaves is reduced by 65.8 percent, and the fresh weight of 3 transgenic lines is reduced by 36.11 percent, 42.25 percent and 44.03 percent respectively; the dry weight of wild stems and leaves is reduced by 51.01 percent, and the dry weight of 3 transgenic lines is reduced by 17.98 percent, 36.48 percent and 31.10 percent respectively; the wild root length is reduced by 74.87 percent, and the 3 transgenic lines are respectively reduced by 49.62 percent, 40.59 percent and 38.39 percent; the survival rate of the wild-type material was only 27.61%, while the survival rate of the transgenic material was significantly higher than that of the non-transgenic material, 68.85%, 68.79% and 71.82%, respectively. The results show that overexpression is performed in ArabidopsisHvACO1The gene obviously enhances the resistance of the transgenic material to the damp damage.
The foregoing shows and describes the general principles, essential features, and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, which are described in the specification and illustrated only to illustrate the principle of the present invention, but that various changes and modifications may be made therein without departing from the spirit and scope of the present invention, which fall within the scope of the invention as claimed. The scope of the invention is defined by the appended claims and equivalents thereof.
Sequence listing
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<120> barley moisture-resistant regulatory gene HvACO1, protein and application thereof in breeding
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Claims (7)
1. The barley humidity resistance regulation related protein is characterized in that the protein is a protein consisting of an amino acid sequence shown in SEQ ID NO. 2.
2. A gene encoding the protein of claim 1.
3. The gene according to claim 2, characterized in that: the gene is a DNA molecule shown as SEQ ID NO. 1.
4. An expression cassette, a recombinant vector, a recombinant microorganism comprising the gene of claim 2 or 3.
Use of (b 1) or (b 2) or (b 3):
(b1) use of the protein of claim 1, or the gene of claim 2 or 3, or an expression cassette, recombinant vector, recombinant microorganism or transgenic cell line comprising the gene of claim 2 or 3 for the resistance to barley wet injury stress;
(b2) use of the protein of claim 1, or the gene of claim 2 or 3, or an expression cassette, a recombinant vector, a recombinant microorganism or a transgenic cell line comprising the gene of claim 2 or 3 for breeding a new moisture-resistant variety of barley or arabidopsis;
(b3) use of the protein of claim 1, or the gene of claim 2 or 3, or an expression cassette, recombinant vector, recombinant microorganism or transgenic cell line comprising the gene of claim 2 or 3 for the resistance to Arabidopsis thaliana wet injury stress.
6. A method for cultivating a plant moisture-resistant variety, which comprises transferring the gene of claim 2 or 3 into a target plant, or transforming the target plant with the expression cassette or recombinant vector of claim 4 to obtain a transgenic plant; the transgenic plant has higher wet damage stress resistance than the target plant; the plant is barley or arabidopsis thaliana.
7. The method for producing a moisture-resistant plant variety according to claim 6, wherein the transgenic plant expresses the barley moisture-resistance regulation-related protein according to claim 1.
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| CN112521475B (en) * | 2020-12-25 | 2022-11-18 | 山东农业大学 | Wheat TaLAX1-A gene and application thereof in improving wheat immature embryo regeneration efficiency |
| CN114107333B (en) * | 2021-10-27 | 2023-05-30 | 上海市农业科学院 | Application of barley receptor kinase HvSERK1 in root hair growth |
| CN114874302B (en) * | 2022-05-27 | 2023-08-01 | 盐城师范学院 | Barley moisture-resistant gene HvMADS1, protein and application thereof in moisture damage stress resistance |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1571899A2 (en) * | 2002-12-06 | 2005-09-14 | Del Monte Fresh Produce Company | Organogenic transformation and regeneration |
| CN102604985A (en) * | 2011-01-25 | 2012-07-25 | 华中农业大学 | Method for culturing transgenic dianthus through agrobacterium-mediated embryogenic callus transformation |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6448474B1 (en) * | 1995-06-07 | 2002-09-10 | University Of Hawaii | Purified proteins, recombinant DNA sequences and processes for controlling the ripening of coffee plants |
| CN102835297A (en) * | 2012-09-14 | 2012-12-26 | 扬州大学 | Identification method of salt tolerance of barley |
| CN105660335B (en) * | 2016-02-16 | 2019-11-12 | 扬州大学 | A kind of method and system for identifying moisture-proof barley variety |
| CN109536509B (en) * | 2018-11-23 | 2021-11-23 | 中国农业科学院油料作物研究所 | Sesame drought-resistant, moisture-resistant and salt-tolerant gene SiNAC56, protein coded by same and application of sesame drought-resistant, moisture-resistant and salt-tolerant gene SiNAC56 |
-
2019
- 2019-04-16 CN CN201910302541.2A patent/CN110129291B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1571899A2 (en) * | 2002-12-06 | 2005-09-14 | Del Monte Fresh Produce Company | Organogenic transformation and regeneration |
| CN102604985A (en) * | 2011-01-25 | 2012-07-25 | 华中农业大学 | Method for culturing transgenic dianthus through agrobacterium-mediated embryogenic callus transformation |
Non-Patent Citations (3)
| Title |
|---|
| Elucidating the hypoxic stress response in barley (Hordeum vulgare L.) during waterlogging: A proteomics approach;Haiye Luan等;《Sci Rep.》;20180625;第8卷(第1期);9655 * |
| Plant Growth Promotion Under Water: Decrease of Waterlogging-Induced ACC and Ethylene Levels by ACC Deaminase-Prodcing Bacteria;Sajid Ali等;《Front. Microbiol.》;20180525;全文 * |
| 乙烯调控植物耐涝机制的研究进展;陈发棣等;《南京农业大学学报》;20180119;第41卷(第2期);203-208 * |
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