CN109354611B - Potassium transport protein KUP3 from tobacco, and coding gene and application thereof - Google Patents
Potassium transport protein KUP3 from tobacco, and coding gene and application thereof Download PDFInfo
- Publication number
- CN109354611B CN109354611B CN201811339731.3A CN201811339731A CN109354611B CN 109354611 B CN109354611 B CN 109354611B CN 201811339731 A CN201811339731 A CN 201811339731A CN 109354611 B CN109354611 B CN 109354611B
- Authority
- CN
- China
- Prior art keywords
- kup3
- leu
- gene
- val
- tobacco
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/415—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
- C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
- C12N15/8242—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits
- C12N15/8243—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Genetics & Genomics (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Biotechnology (AREA)
- Molecular Biology (AREA)
- Biochemistry (AREA)
- Zoology (AREA)
- Biophysics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biomedical Technology (AREA)
- Wood Science & Technology (AREA)
- Physics & Mathematics (AREA)
- Microbiology (AREA)
- Cell Biology (AREA)
- Plant Pathology (AREA)
- Nutrition Science (AREA)
- Botany (AREA)
- Gastroenterology & Hepatology (AREA)
- Medicinal Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention relates to the field of genetic engineering, and particularly discloses a potassium transport protein KUP3 from tobacco, and a coding gene and application thereof. The invention provides a KUP3 gene separated from tobacco for the first time, the full length of the KUP3 gene is 2364bp, and after functional verification, after the KUP3 gene provided by the invention is transferred into a potassium absorption defective yeast mutant strain R5421, a recombinant yeast expressing the KUP3 gene has the functions of potassium ion absorption and transfer again. Therefore, the KUP3 gene provided by the invention has the function of promoting potassium ion absorption and transportation.
Description
Technical Field
The invention relates to the field of genetic engineering, in particular to a potassium transport protein KUP3 from tobacco, and a coding gene and application thereof.
Background
The potassium transporter is a carrier egg capable of transporting potassium ions into cells when the concentration of external potassium ions is extremely lowWhite. Potassium transporters are generally transmembrane proteins, and transport of potassium ions across the membrane is accomplished by conformational changes, an active transport process requiring energy from ATP, usually K+/H+Or K+/Na+And (5) carrying out coordinated transportation.
The research on potassium transporter genes in the prior art is relatively extensive in model plant Arabidopsis, for example, the research shows that the survival rate of an Arabidopsis mutant KUP6 is reduced compared with that of a wild plant under drought stress, and 35S shows that the drought tolerance of KUP6 overexpression transgenic plants is obviously enhanced (Shabala et al, 2007); and mutant seedlings of KUP7 gene show a low-potassium sensitive phenotype with etiolated leaves under low potassium stress, the content of potassium ions at roots of the mutant seedlings is obviously reduced, and the loss of the function of KUP7 gene reduces the potassium absorption capacity of arabidopsis seedlings under low potassium conditions and the concentration of potassium ions in xylem sap (Min et al, 2016).
Tobacco is a crop with large potassium consumption, the potassium content of tobacco leaves is an important index for measuring the quality of the tobacco leaves, at present, researches on potassium transporter genes in the tobacco are less, and the function of KUP of the tobacco is unknown.
Disclosure of Invention
In order to solve the problems in the prior art, the invention aims to provide a potassium transporter KUP3 from tobacco as well as a coding gene and application thereof.
In order to realize the purpose of the invention, the technical scheme of the invention is as follows:
the invention firstly provides a potassium transport protein which is obtained from tobacco and is named as KUP3 protein and is (a) or (b)
(a) The amino acid sequence is shown as SEQ ID NO. 1;
(b) and (b) a protein which is derived from the protein (a) and related to plant potassium ion absorption and transport regulation and control through substitution and/or deletion and/or addition of one or more amino acid residues in the sequence shown in SEQ ID NO. 1.
The KUP3 protein in (b) may be synthesized by man, or may be obtained by synthesizing the coding gene and then performing biological expression. The gene encoding KUP3 protein in (b) above can be obtained by deleting one or more amino acid residues from the DNA sequence shown in SEQ ID NO.2, and/or by performing missense mutation of one or more base pairs.
The gene (KUP3 gene) for coding the KUP3 protein also belongs to the protection scope of the invention.
The gene can be specifically the following DNA molecules of 1) or 2) or 3):
1) the coding region is a DNA molecule shown as SEQ ID NO. 2;
2) a DNA molecule which is hybridized with the DNA sequence defined in 1) under strict conditions and codes a protein related to the regulation and control of plant potassium ion absorption and transport;
3) DNA molecules which have more than 90 percent of homology with the DNA sequences limited by 1) or 2) and encode proteins related to plant potassium ion absorption and transport regulation.
The stringent conditions can be hybridization and washing with 0.1 XSSPE (or 0.1 XSSC), 0.1% SDS solution at 65 ℃ in DNA or RNA hybridization experiments.
The research shows that the KUP3 gene plays a significant role in promoting potassium ion absorption and transport.
Further, the KUP3 gene of the invention is prepared by the following steps:
(1) designing PCR amplification primers, wherein the PCR amplification primers comprise a forward primer and a reverse primer:
a forward primer: 5'-ATGGATATCGACTACGGGAA-3' the flow of the air in the air conditioner,
reverse primer: 5'-TTAAACAATGTAGACCATCC-3', respectively;
(2) extracting total RNA of the tobacco cells, synthesizing cDNA of the tobacco cells, carrying out PCR amplification of KUP3 gene by taking the cDNA of the tobacco cells as a template to obtain a target segment, and sequencing.
Preferably, the PCR amplification system is a 20 μ L system, including Premix ExTaq 10 μ L, forward primer 0.5 μ L of 10 μ M, reverse primer 0.5 μ L of 10 μ M, tobacco cell cDNA 1 μ L, ddH2O 8μL。
Preferably, the reaction procedure of the PCR amplification is as follows: pre-denaturation at 95 ℃ for 5 min; denaturation at 95 ℃ for 30 s; annealing at 55 ℃ for 30 s; extending for 2min at 72 ℃; 35 cycles.
Preferably, the target fragment is introduced into escherichia coli DH5 alpha competent cells for colony PCR verification before sequencing, and sequencing is performed after positive clones are verified.
Preferably, the nucleotide sequence of the forward primer used for colony PCR verification is as follows: 5'-ATGGATATCGACTACGGGAA-3', the nucleotide sequence of the reverse primer used for colony PCR verification is: 5'-TTAAACAATGTAGACCATCC-3' are provided.
Preferably, the colony PCR verification system is 10 μ L, including 5 μ L of Premix ExTaq, 0.5 μ L of 10 μ M forward primer, 0.5 μ L of 10 μ M reverse primer, ddH2O 4μL。
Furthermore, biological materials such as recombinant expression vectors, expression cassettes, transgenic cell lines or recombinant bacteria containing the KUP3 gene belong to the protection scope of the invention.
The recombinant expression vector can be constructed by using the existing plant expression vector. The recombinant expression vector can be, for example, a binary Agrobacterium vector, a vector useful for microprojectile bombardment of plants, and the like. When the KUP3 gene is used for constructing a recombinant expression vector, any one of enhanced, constitutive, tissue-specific or inducible promoters can be added in front of the transcription initiation nucleotide, and can be used alone or combined with other plant promoters; in addition, when constructing a recombinant expression vector using KUP3 gene, enhancers, including translational or transcriptional enhancers, may be used, and these enhancer regions may be ATG initiation codon or initiation codon of adjacent regions, etc., but must be in the same reading frame as the coding sequence to ensure proper translation of the entire sequence. The translational control signals and initiation codons are widely derived, either naturally or synthetically. The translation initiation region may be derived from a transcription initiation region or a structural gene.
The recombinant expression vector carrying the KUP3 gene can be transformed into plant cells or tissues by conventional biological methods such as Ti plasmid, Ri plasmid, plant virus vector, direct DNA transformation, microinjection, conductance, Agrobacterium mediation and the like.
Furthermore, the invention also protects the application of the KUP3 protein, the KUP3 gene and the biological material containing the gene in promoting the absorption and the transportation of potassium ions of plants or microorganisms.
The plant comprises tobacco and arabidopsis, and the microorganism comprises yeast.
For example, in a specific embodiment of the present invention, after the KUP3 gene is transformed into a potassium uptake-deficient yeast mutant strain R5421 (described in Maathuis F J M and Sanders D1996 mechanics of potassium uptake by high plant roots. Physiol. plant.96, 158-168.), the recombinant yeast expressing the KUP3 gene has again potassium uptake and transport functions; after the KUP3 gene in the tobacco plant is subjected to overexpression, the content of potassium ions in tobacco leaves of the tobacco plant can be obviously improved.
The application can be selected as that the KUP3 gene of the tobacco is transferred into a tobacco plant, and the KUP3 gene is overexpressed to improve the content of potassium ions in tobacco leaves of the tobacco plant.
Furthermore, the invention also protects the application of the KUP3 protein, the KUP3 gene and the biological material containing the gene in preparing transgenic plants.
Furthermore, the invention also protects the application of the KUP3 protein, the KUP3 gene and the biological material containing the gene in plant breeding.
The breeding aim is to promote the absorption and the transportation of plant potassium ions, and preferably to improve the content of the potassium ions in tobacco leaves of tobacco plants.
The plant is a monocotyledon or a dicotyledon. Such as tobacco or Arabidopsis thaliana KUP3 mutants.
The invention has the beneficial effects that: the invention provides a KUP3 gene separated from tobacco for the first time, the full length of the KUP3 gene is 2364bp, and after functional verification, after the KUP3 gene provided by the invention is transferred into a potassium absorption defective yeast mutant strain R5421, a recombinant yeast expressing the KUP3 gene has the functions of potassium ion absorption and transfer again. Therefore, the KUP3 gene provided by the invention has the function of promoting potassium ion absorption and transportation.
Drawings
FIG. 1 shows the results of the yeast function complementation test in example 2 of the present invention. Wherein A is the growth condition on a culture medium with the potassium ion concentration of 20uM, and B is the growth condition on a culture medium with the potassium ion concentration of 2 mM; in the figure, 1 is a negative control (transferred into an empty vector), 2 is a recombinant yeast transferred into KUP3 gene, and 3 is a positive control yeast transferred into Arabidopsis thaliana KUP gene; the growth results of the stock solution, the 10-time diluent, the 100-time diluent and the 1000-time diluent on the culture medium are sequentially shown from left to right.
Detailed Description
Preferred embodiments of the present invention will be described in detail with reference to the following examples. It is to be understood that the following examples are given for illustrative purposes only and are not intended to limit the scope of the present invention. Various modifications and alterations of this invention will become apparent to those skilled in the art without departing from the spirit and scope of this invention.
The experimental procedures used in the following examples are all conventional procedures unless otherwise specified. Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
The potassium uptake-deficient yeast mutant strain R5421 of the invention is described in Maathuis F J M and Sanders D1996 mechanics of potassium uptake by high promoter plants, Physiol.plant.96, 158-168. The gene is characterized by comprising a P416 yeast episomal shuttle expression vector, a TEF constitutive promoter, a CYC1 terminator, a CEN6 ARSH4 replication origin, a selection marker in yeast of URA3 and a selection marker in Escherichia coli of Amp. Described in Functional Expression of a ω -3 Fatty Acid Desaturase Gene from Glycine max in Saccharomyces cerevisiae.
Example 1 acquisition of KUP3 Gene
Taking 0.5g of fresh tobacco leaves, extracting total RNA of tobacco cells by a Trizol method, synthesizing cDNA by a cDNA synthesis kit of TaKaRa company, further designing by Primer5.0 software and obtaining primers through artificial optimization, wherein the primers comprise a forward primer and a reverse primer, and the nucleotide sequence of the forward primer is as follows: 5'-ATGGATATCGACTACGGGAA-3', respectively; the nucleotide sequence of the reverse primer is 5'-TTAAACAATGTAGACCATCC-3', and the synthesized cDNA is used as a template for PCR amplification.
The PCR amplification system is a 20 mu L system and comprises: premix ExTaq 10. mu.L, forward primer 0.5. mu.L at 10. mu.M, reverse primer 0.5. mu.L at 10. mu.M, tobacco cell cDNA 1. mu.L, ddH2O 8μL。
The PCR amplification reaction program is as follows: pre-denaturation at 95 ℃ for 5 min; denaturation at 95 ℃ for 30s, annealing at 55 ℃ for 30s, extension at 72 ℃ for 2min, 35 cycles.
After PCR amplification is completed, a DNA purification kit is used for purifying a target fragment, the purified target fragment is connected with a pMD19-T vector for 12 hours at 16 ℃ to obtain a connection product, the obtained connection product is transformed into escherichia coli DH5 alpha competent cells to obtain transformed escherichia coli DH5 alpha, and the transformed escherichia coli DH5 alpha is inoculated on an LB plate coated with ampicillin to be screened and cultured to obtain a positive clone. After obtaining positive clones, verifying the positive clones by adopting a colony PCR method, wherein forward primers of the colony PCR are as follows: 5'-ATGGATATCGACTACGGGAA-3', the reverse primer is: 5'-TTAAACAATGTAGACCATCC-3', respectively; the colony PCR system is 10 μ L, including 5 μ L of Premix ExTaq, 0.5 μ L of 10 μ M forward primer, 0.5 μ L of 10 μ M reverse primer, ddH2O4. mu.L. Then randomly selecting 3 independent positive clones from the verified positive clones, sending the positive clones to a biotechnology company for sequencing, and obtaining the sequence of the KUP3 gene after sequencing, wherein the sequence is shown as SEQ ID NO. 2.
Example 2 Effect of KUP3 Gene on promoting Potassium ion absorption and transport
The T-vector connected with the KUP3 gene in example 1 and the expression vector P416 are subjected to double enzyme digestion (enzyme digestion sites are SmaI and Xho I), the target gene and the expression vector P416 are recovered and then connected by ligase, the connected recombinant yeast expression vector is transferred into competent cells of escherichia coli DH5 alpha, PCR amplification and enzyme digestion are carried out on a single colony of the transformed escherichia coli to verify whether the construction is successful, and the successfully constructed recombinant yeast expression vector is transferred into R5421.
The method comprises the following specific steps: taking the preserved R5421 yeast by an inoculating ring, streaking on a solid culture medium YPDA, and culturing at 28 ℃ for 12 h; picking a single colony of the R5421 yeast in an Ep tube, adding 1mL of YPDA culture solution, and vortexing; transferring all the bacterial liquid into a triangular flask filled with YPDA culture solution, and shaking at 30 ℃ and 250rpm until the OD600 is 1.2; switching over according to the ratio of 1:10, and shaking until OD600 is 1.0-1.2; centrifuging at 28 deg.C and 1000rpm for 5min, and resuspending with 1/2 volume of sterilized ultrapure water; centrifuging at 28 deg.C and 1000rpm for 5min for collecting bacteria, and sucking off supernatant; the following ingredients (per 5mL of original bacterial liquid) were added in sequence:
vortex for 1min to make the transformation system completely mixed; incubating in 30 deg.C water bath for 30 min; placing in 42 deg.C water bath, thermally shocking for 28min, and cooling on ice for 10 min; centrifuging at 7000rpm for 15s, and discarding the supernatant; gently resuspend the pellet with 1mL of sterile water; spreading 200. mu.L of the transformation mixture on an auxotrophic plate; cultured at 30 ℃ for 3 days. And extracting yeast plasmids and identifying the result.
Selecting identified yeast single colony, streaking on auxotrophic plate, and culturing at 30 deg.C for 3 days; dipping a small amount of thallus on the auxotrophic flat plate by using a toothpick, and culturing in 2mL of auxotrophic solution for 12 h; centrifuging at 8000rpm for 1min, and collecting thallus; discarding the supernatant, suspending the thallus with 1mL of double distilled water, and centrifuging at 8000rpm for 1 min;
discard the supernatant, resuspend with 1mL double distilled water, adjust OD600Is 0.8; the undiluted bacterial solution and the 10-fold, 100-fold, and 1000-fold diluted bacterial solutions were cultured in 5uL of 2mM potassium ion medium at 30 ℃ for 3 days, and the results were observed.
As shown in FIG. 1, the yeast of the negative control group (transformed into P416 empty vector) hardly grew and both the recombinant yeast of the tobacco KUP3 gene and the recombinant yeast of the positive control group (transformed into Arabidopsis thaliana KUP gene) could grow on a 2mM medium (AP medium (1L): 546. mu.L phosphate, 1.742g L-arginine, 1mL 1000 Xvitamin solution, 1mL 1000 Xmicroelement solution, 0.77g uracil, 10mL 100 XUra, 20g glucose, 15g agar powder) with potassium ion concentration 20 uM. With the increase of dilution factor, the recombinant yeast transferred into the KUP3 gene of tobacco and the recombinant yeast of the positive control group can still grow. The results prove that the KUP3 gene of the tobacco has potassium absorption and transport functions.
Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.
Sequence listing
<110> Guizhou province tobacco science research institute
<120> potassium transporter KUP3 from tobacco, and coding gene and application thereof
<130> KHP171117841.1
<160> 4
<170> SIPOSequenceListing 1.0
<210> 1
<211> 787
<212> PRT
<213> tobacco (Nicotiana tabacum)
<400> 1
Met Asp Ile Asp Tyr Gly Lys Cys Trp Asp Thr Ser Lys Ser Trp Lys
1 5 10 15
Ser Thr Leu Ile Leu Ala Tyr Gln Ser Leu Gly Val Val Tyr Gly Asp
20 25 30
Leu Ser Ile Ser Pro Leu Tyr Val Tyr Lys Ser Thr Phe Ala Glu Asp
35 40 45
Ile His His Ser Glu Thr Asn Glu Glu Ile Phe Gly Val Leu Ser Phe
50 55 60
Val Phe Trp Thr Leu Thr Leu Val Pro Leu Phe Lys Tyr Val Phe Ile
65 70 75 80
Val Leu Arg Ala Asp Asp Asn Gly Glu Gly Gly Thr Phe Ala Leu Tyr
85 90 95
Ser Leu Ile Cys Arg His Ala Lys Val Ser Leu Leu Pro Asn Arg Gln
100 105 110
Val Ala Asp Glu Ala Leu Ser Thr Tyr Lys Leu Glu His Pro Pro Glu
115 120 125
Thr Lys Asn Ser Ser Arg Val Lys Leu Leu Leu Glu Lys His Lys Phe
130 135 140
Leu His Thr Ala Leu Leu Ile Leu Val Leu Leu Gly Thr Cys Met Val
145 150 155 160
Ile Gly Asp Gly Leu Leu Thr Pro Ala Ile Ser Val Phe Ser Ala Val
165 170 175
Ser Gly Leu Glu Leu Ser Met Ser Arg Asp His His Gln Tyr Ala Val
180 185 190
Ile Pro Ile Thr Cys Phe Ile Leu Val Cys Leu Phe Ala Leu Gln His
195 200 205
Tyr Gly Thr His Arg Val Gly Phe Cys Phe Ala Pro Ile Val Leu Ile
210 215 220
Trp Leu Leu Cys Ile Ser Ala Leu Gly Leu Tyr Asn Ile Ile His Trp
225 230 235 240
Asn Pro Leu Val Tyr Lys Ala Leu Ser Pro Tyr Tyr Met Val Lys Phe
245 250 255
Leu Lys Lys Thr Arg Lys Gly Gly Trp Met Ser Leu Gly Gly Ile Leu
260 265 270
Leu Cys Ile Thr Gly Ser Glu Ala Met Phe Ala Asp Leu Gly His Phe
275 280 285
Ser Tyr Ser Ala Ile Gln Ile Ala Phe Thr Phe Leu Val Tyr Pro Ala
290 295 300
Leu Ile Leu Ala Tyr Met Gly Gln Ala Ala Phe Leu Ser Lys His His
305 310 315 320
His Thr Ile His Lys Ile Gly Phe Tyr Val Ser Val Pro Asp Cys Val
325 330 335
Arg Trp Pro Val Leu Val Ile Ala Ile Leu Ala Ser Val Val Gly Ser
340 345 350
Gln Ala Ile Ile Ser Gly Thr Phe Ser Ile Ile Asn Gln Ser Gln Ser
355 360 365
Leu Gly Cys Phe Pro Arg Val Lys Val Val His Thr Ser Ala Lys Ile
370 375 380
His Gly Gln Ile Tyr Ile Pro Glu Ile Asn Trp Ile Leu Met Ile Leu
385 390 395 400
Cys Val Ala Val Thr Ile Gly Phe Arg Asp Thr Lys His Met Gly Asn
405 410 415
Ala Ser Gly Leu Ala Val Met Ala Val Met Leu Val Thr Thr Cys Leu
420 425 430
Thr Ser Leu Val Ile Ile Leu Cys Trp Asn Lys Pro Pro Ile Leu Ala
435 440 445
Leu Gly Phe Leu Leu Leu Phe Gly Ser Ile Glu Leu Leu Tyr Phe Ser
450 455 460
Ala Ser Val Ile Lys Phe Leu Glu Gly Ala Trp Leu Pro Ile Leu Leu
465 470 475 480
Ala Leu Phe Leu Val Thr Val Met Phe Val Trp His Tyr Ala Thr Val
485 490 495
Lys Lys Tyr Glu Tyr Asp Leu His Asn Lys Val Ser Leu Glu Trp Leu
500 505 510
Leu Ala Leu Gly Pro Ser Leu Gly Ile Thr Arg Val Pro Gly Ile Gly
515 520 525
Leu Val Phe Thr Asp Leu Thr Ser Gly Ile Pro Ala Asn Phe Ser Arg
530 535 540
Phe Val Thr Asn Leu Pro Ala Tyr His Arg Ile Leu Val Phe Val Cys
545 550 555 560
Val Lys Ser Val Pro Val Pro Phe Val Pro Pro Ala Glu Arg Tyr Leu
565 570 575
Val Gly Arg Val Gly Pro Ala Ala His Arg Ser Tyr Arg Cys Ile Val
580 585 590
Arg Tyr Gly Tyr Arg Asp Val His Gln Asp Val Asp Ser Phe Glu Ser
595 600 605
Glu Leu Val Ser Lys Leu Ala Asp Phe Ile Arg Tyr Asp Trp Tyr Lys
610 615 620
Thr His Gly Ile Ile Asp Asn Glu Asp Asp Gly Ser Arg Ser Gly Ala
625 630 635 640
Ser Ser Gly Glu Cys Arg Leu Thr Val Ile Gly Thr Leu Ala Phe Ser
645 650 655
Gly Thr Pro Ala Phe Glu Leu Glu Asp Asn Val Gln Leu Ala Ser Val
660 665 670
Ser Val Gly Phe Pro Thr Ala Glu Ser Val Thr Asp Val Ile Glu Met
675 680 685
Arg Pro Val Glu Arg Arg Val Arg Phe Ala Ile Asp Asn Glu Ser Glu
690 695 700
Val Asp Ser Arg Glu Glu Met Asp Thr Gln Leu Gln Glu Glu Leu Glu
705 710 715 720
Asp Leu Tyr Ala Ala Gln Gln Ala Gly Thr Ala Phe Val Leu Gly His
725 730 735
Ser His Val Lys Ala Lys Gln Gly Ser Ser Met Leu Lys Arg Leu Ala
740 745 750
Ile Asn Tyr Gly Tyr Asn Phe Leu Arg Arg Asn Cys Arg Gly Ala Asp
755 760 765
Val Ser Leu Lys Val Pro Pro Ala Ser Leu Leu Glu Val Gly Met Val
770 775 780
Tyr Ile Val
785
<210> 2
<211> 2364
<212> DNA
<213> tobacco (Nicotiana tabacum)
<400> 2
atggatatcg actacgggaa gtgttgggac acttcaaagt cttggaagag tacactgatt 60
ctggcatacc aaagtcttgg ggtagtgtat ggtgatctca gtatttcccc tctctatgtc 120
tacaagagca catttgcaga agacatccat cattccgaga ccaacgaaga gattttcggc 180
gttctctctt tcgttttttg gactttaacc ttagttcctt tgttcaaata tgtctttatc 240
gtacttcgag ctgatgataa tggagagggt ggtacttttg ctctctattc cttaatatgc 300
aggcatgcca aagtaagcct tcttccgaac agacaagttg ccgatgaagc tctttctacc 360
tacaaacttg agcaccctcc cgagacgaag aacagctcaa gggtgaagtt gcttcttgag 420
aaacacaaat tcttgcacac tgctttgcta atcttggtgc ttcttggcac ttgtatggtc 480
attggagatg gactacttac tccagccata tctgtattct ctgcagtttc tggccttgag 540
ttatcaatgt ctcgggatca ccatcaatat gcagtgattc caataacttg cttcatatta 600
gtctgtctgt tcgcattaca acattatggc acacatcgag ttggtttttg ttttgcacca 660
attgtgttga tctggttact ctgcatcagt gctcttggat tatacaacat tatccactgg 720
aatccacttg tttataaagc tctttccccc tattacatgg taaagttcct gaagaaaaca 780
agaaaaggag gatggatgtc tttgggtgga attctcctct gcataactgg ttctgaagca 840
atgtttgctg atcttggaca cttctcatat tctgcaattc aaattgcatt cacctttctg 900
gtttatccag cgttaatttt ggcatatatg ggtcaagcag ctttcttatc aaagcatcac 960
cacaccattc acaagattgg tttctatgta tcagttccag attgcgtgag atggccagtg 1020
ctagtaatag caattctggc ttccgttgtg ggaagtcagg caatcatcag tggaacattc 1080
tcaattatca accagagtca atcccttggt tgcttcccaa gagtcaaagt tgttcacacc 1140
tctgccaaaa tacatggcca gatatacatt cctgagatca attggatact catgattctt 1200
tgtgttgctg tgactattgg attcagagat acaaagcaca tgggaaatgc gtcaggacta 1260
gcagtgatgg cggtgatgct ggtgaccact tgccttactt cattagttat catcctctgc 1320
tggaacaagc ctccaatact ggccctcggg tttctccttc tatttggatc tattgagtta 1380
ctctacttct cagcctctgt cattaagttt cttgagggcg cttggcttcc aatcctgctt 1440
gcactctttt tggttactgt catgtttgtt tggcactatg ccacagttaa aaagtatgaa 1500
tatgatctgc acaacaaggt ttcattagaa tggctgctgg cactaggtcc aagcttgggt 1560
attactcgag tccctggcat cggcctcgta ttcactgatt tgacttctgg gattcctgcc 1620
aacttctcac gttttgttac caacctccct gcctaccatc gcatacttgt ttttgtgtgt 1680
gtgaaatccg tgcctgtccc ttttgtgccc ccggctgaga gataccttgt aggccgcgtt 1740
ggtcctgcag ctcatcgttc ctatagatgc attgtccgtt atggttaccg agacgttcac 1800
caagatgtcg actcctttga atccgaactt gtcagtaagc tggctgattt catccggtat 1860
gattggtaca agacgcacgg aatcattgac aacgaggatg acggctcacg ttctggtgca 1920
tcgtcgggag aatgcagact gaccgttata ggaaccctgg ctttctcagg tacaccagct 1980
ttcgagctcg aggacaatgt gcaactggca agtgtgtccg tcgggttccc tacagctgaa 2040
agtgtgacag atgtcatcga gatgcgacca gtggaaagaa gagtgagatt tgccatagat 2100
aatgagtcgg aagtcgattc acgggaagaa atggatactc agctgcagga ggagctagaa 2160
gatttgtatg cagcacaaca agctgggaca gcattcgtat taggacattc acatgtaaaa 2220
gcaaaacaag gatcatctat gttgaagagg ttggctatta attatggtta taatttcctt 2280
aggaggaatt gtaggggtgc agatgtatcc ctcaaggtgc ctccagcttc ccttcttgaa 2340
gttgggatgg tctacattgt ttaa 2364
<210> 3
<211> 20
<212> DNA
<213> Artificial primer (Artificial Sequence)
<400> 3
atggatatcg actacgggaa 20
<210> 4
<211> 20
<212> DNA
<213> Artificial primer (Artificial Sequence)
<400> 4
ttaaacaatg tagaccatcc 20
Claims (3)
1. An application of protein KUP3 from tobacco in promoting absorption and transport of yeast potassium ions is disclosed, wherein the amino acid sequence of the protein KUP3 is shown in SEQ ID NO. 1.
2. An application of a gene for coding protein KUP3 from tobacco in promoting absorption and transportation of yeast potassium ions, wherein the nucleotide sequence of the gene is shown in SEQ ID NO. 2.
3. The application of the biological material containing the gene of protein KUP3 from tobacco in promoting the absorption and the transportation of yeast potassium ions, wherein the nucleotide sequence of the gene is shown as SEQ ID NO.2, and the biological material is a recombinant expression vector, an expression cassette or a recombinant bacterium.
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201811339731.3A CN109354611B (en) | 2018-11-12 | 2018-11-12 | Potassium transport protein KUP3 from tobacco, and coding gene and application thereof |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201811339731.3A CN109354611B (en) | 2018-11-12 | 2018-11-12 | Potassium transport protein KUP3 from tobacco, and coding gene and application thereof |
Publications (2)
Publication Number | Publication Date |
---|---|
CN109354611A CN109354611A (en) | 2019-02-19 |
CN109354611B true CN109354611B (en) | 2021-08-31 |
Family
ID=65344839
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN201811339731.3A Active CN109354611B (en) | 2018-11-12 | 2018-11-12 | Potassium transport protein KUP3 from tobacco, and coding gene and application thereof |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN109354611B (en) |
Citations (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN105524157A (en) * | 2016-01-27 | 2016-04-27 | 中国农业大学 | Potassium ion channel protein KC1-D as well as encoding gene and application thereof |
-
2018
- 2018-11-12 CN CN201811339731.3A patent/CN109354611B/en active Active
Patent Citations (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN105524157A (en) * | 2016-01-27 | 2016-04-27 | 中国农业大学 | Potassium ion channel protein KC1-D as well as encoding gene and application thereof |
Non-Patent Citations (3)
Title |
---|
《PREDICTED: Nicotiana tabacum potassium transporter 2-like (LOC107791389), transcript variant X4, mRNA》;NCBI;《GenBank Database》;20160503;FEATURES中CDS部分的translation部分,ORIGIN部分 * |
《高等植物钾转运蛋白》;刘贯山;《生物技术通报》;20061231(第5期);第15页右栏第2.3节 * |
NCBI.《PREDICTED: Nicotiana tabacum potassium transporter 2-like (LOC107791389), transcript variant X4, mRNA》.《GenBank Database》.2016,XM_016613450.1. * |
Also Published As
Publication number | Publication date |
---|---|
CN109354611A (en) | 2019-02-19 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN107090461B (en) | Tobacco HKT1 gene and preparation method and application thereof | |
CN112831478A (en) | Protein OsCAT8 for regulating rice chalkiness and coding gene and application thereof | |
CN109371037B (en) | Tobacco AKT1 gene and application thereof | |
CN109553666B (en) | Potassium transport protein KUP9 from tobacco, and coding gene and application thereof | |
WO2023227137A1 (en) | Use of tapdil4-1b gene in fusarium head blight resistance of plant and method for constructing tapdil4-1b transgenic plant | |
CN109438563B (en) | Tobacco KUP7 protein and coding gene and application thereof | |
CN109369786B (en) | Tobacco KUP8 protein and coding gene and application thereof | |
CN109354613B (en) | Potassium transport protein TPK1 from tobacco as well as encoding gene and application thereof | |
CN109354611B (en) | Potassium transport protein KUP3 from tobacco, and coding gene and application thereof | |
CN109354612B (en) | Tobacco AKT2/3 gene and application thereof | |
CN109369787B (en) | Potassium transport protein KUP11 from tobacco, and coding gene and application thereof | |
CN109336958B (en) | Potassium transport protein KUP10 from tobacco, and coding gene and application thereof | |
CN113481210B (en) | Application of cotton GhDof1.7 gene in promotion of salt tolerance of plants | |
CN109438564B (en) | Tobacco KUP6 protein and coding gene and application thereof | |
CN109354610B (en) | Tobacco KUP5 protein and coding gene and application thereof | |
CN109485709B (en) | Tobacco KUP4 protein and coding gene and application thereof | |
CN109369788B (en) | Potassium transport protein TPK1-1 from tobacco as well as encoding gene and application thereof | |
CN114456242A (en) | PRP protein and coding gene and application thereof | |
CN110484542B (en) | Arabidopsis thaliana disease-resistant related gene EIJ1 and application thereof | |
CN109553667B (en) | Tobacco KUP2 gene and application thereof | |
CN109553665B (en) | Tobacco KC1 gene and application thereof | |
CN109553668B (en) | Tobacco KUP1 gene and application thereof | |
CN114525298A (en) | Application of soybean protein GmFVE in plant salt tolerance regulation | |
CN107699580B (en) | Application of arabidopsis U1A gene in improving salt tolerance of plants | |
CN114716522A (en) | Application of KIN10 protein and related biological materials thereof in saline-alkali tolerance of plants |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PB01 | Publication | ||
PB01 | Publication | ||
SE01 | Entry into force of request for substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
GR01 | Patent grant | ||
GR01 | Patent grant |