KR101448118B1 - Histone4 promoter isolated from Capsicum annuum and uses thereof - Google Patents

Abstract

The present invention relates to a recombinant plant expression vector comprising the histone 4 promoter and terminator derived from pepper ( Capsicum annuum ), a recombinant plant expression vector comprising the promoter, a recombinant plant expression vector comprising the terminator, The present invention relates to a method for producing a transgenic plant expressing a gene, a transgenic plant expressing a foreign gene produced by the method, and a seed thereof, wherein the histone 4 promoter derived from pepper of the present invention can produce a useful gene in a large amount in plants So that it can be very useful industrially.

Description

Histone 4 promoter derived from pepper and its use {Histone4 promoter isolated from Capsicum annuum and uses thereof}

The present invention relates to a recombinant plant expression vector comprising the histone 4 promoter and terminator derived from pepper ( Capsicum annuum ), a recombinant plant expression vector comprising the promoter, a recombinant plant expression vector comprising the terminator, A method for producing a transgenic plant expressing a gene, a transgenic plant expressing a foreign gene produced by the method, and a seed thereof.

Recently, many studies have been conducted to produce useful substances using foreign gene introduction in plants using genetic engineering technology. In this process, when a foreign useful gene is expressed in a transgenic plant, a strong promoter involved in gene expression is required.

A promoter is a genomic region linked to the upper side of a structural gene, and plays a role of regulating the transcription of the structural gene linked thereto to mRNA. The promoter is activated by the binding of several general transcription factors, and it has a base sequence such as TATA box and CAT box that regulate gene expression in general. Since proteins required for basic metabolism in living organisms must maintain a constant concentration in cells, the promoter linked to these genes is always activated by the action of common transcription factors. On the other hand, proteins that normally do not have a role and require function only under specific circumstances are connected to an inducible promoter which induces the expression of the structural gene. Inducible promoters are activated by the binding of specific transcription factors activated by external stimuli in the development of organisms or from environmental factors from the environment.

Promoters derived from Cauliflower mosaic virus (CaMV35S) in which expression is induced in whole tissues of plants, such as promoters that are constantly and strongly expressed in the development of transgenic plants through gene transformation, Is widely used. However, since the expression level of the foreign gene is low, it is difficult to exceed 0.1% of the total expressed water-soluble protein.

In order to solve the above problems, a strong plant promoter capable of increasing expression of a foreign transgene in a transgenic plant was isolated from red pepper and the intensity of the promoter was confirmed.

Korean Patent No. 0784165 discloses a 'promoter of UDP-glucosyltransferase derived from a red pepper plant', Korean Patent Registration No. 0974820 discloses a root-specific expression promoter derived from 'red pepper aquaporin gene, Root specific expression vector 'has been disclosed, but the histone 4 promoter derived from pepper of the present invention and its use have not been described at all.

The present invention provides a novel promoter capable of overexpressing a foreign gene in a transgenic tobacco by transforming tobacco plants with a recombinant plant expression vector containing a histone 4 promoter derived from pepper The present invention has been completed.

In order to solve the above problems, the present invention provides a histone 4 promoter derived from pepper ( Capsicum annuum ).

The present invention also provides a histone 4 terminator derived from pepper ( Capsicum annuum ).

The present invention also provides a recombinant plant expression vector comprising the promoter.

The present invention also provides a recombinant plant expression vector comprising the terminator.

The present invention also provides a method for producing a transgenic plant expressing a foreign gene comprising the step of transforming a plant cell with the vector.

The present invention also provides a transgenic plant expressing a foreign gene produced by the above method.

The present invention also provides a seed of the transgenic plant.

In the present invention, a foreign gene was expressed in a transgenic plant using a histone 4 promoter derived from pepper. As a result, it was confirmed that the histone 4 promoter is a novel promoter capable of highly efficiently increasing the expression of a foreign gene introduced into the transgenic plant. Therefore, the histone 4 promoter derived from pepper of the present invention can be usefully used industrially because it can produce a useful gene in a large amount in plants.

FIG. 1 shows a recombinant vector map in which GFP is linked to the histone 4 promoter for measuring the intensity of the pepper-histone 4 promoter.
Figure 2 shows the effect of the agro-infiltration method on tobacco ( Nicotiana The expression of GFP was confirmed by RT-PCR after transient expression of the GFP gene in B. benthamiana . P represents pCAMBIA1302 (p35S: GFP vector), and H represents histone 4 (Histone4: GFP vector).

In order to accomplish the object of the present invention, the present invention provides a histone 4 promoter derived from a red pepper ( Capsicum annuum ) comprising the nucleotide sequence of SEQ ID NO: 1.

The histone 4 promoter of the present invention can significantly increase the expression of a target gene by promoting transcription initiation of a gene introduced from a plant, or increasing the stability of transcribed mRNA or the efficiency of gene decoding.

Also, homologues of the promoter sequences are included within the scope of the present invention. The homologue is a base sequence having a functional characteristic similar to that of SEQ ID NO: 1, although the base sequence is changed. Specifically, the promoter sequence has a nucleotide sequence that has at least 70% homology, more preferably at least 80% homology, more preferably at least 90% homology, and most preferably at least 95% homology with the nucleotide sequence of SEQ ID NO: 1 . "% Of sequence homology to polynucleotides" is ascertained by comparing the comparison region with two optimally aligned sequences, and a portion of the polynucleotide sequence in the comparison region is the reference sequence for the optimal alignment of the two sequences (I. E., A gap) relative to the < / RTI >

The present invention also provides a histone 4 terminator derived from a pepper ( Capsicum annuum ) comprising the nucleotide sequence of SEQ ID NO: 2.

Also, homologues of the terminator sequence are included within the scope of the present invention. The homologue is a base sequence having a functional characteristic similar to the base sequence of SEQ. Specifically, the promoter sequence has a nucleotide sequence having a sequence homology of at least 70%, more preferably at least 80%, even more preferably at least 90%, most preferably at least 95% with the nucleotide sequence of SEQ ID NO: 2 . "% Of sequence homology to polynucleotides" is ascertained by comparing the comparison region with two optimally aligned sequences, and a portion of the polynucleotide sequence in the comparison region is the reference sequence for the optimal alignment of the two sequences (I. E., A gap) relative to the < / RTI >

The present invention also provides a recombinant plant expression vector comprising the promoter.

The present invention also provides a recombinant plant expression vector comprising the terminator.

In a plant expression vector according to an embodiment of the present invention, the vector may be a recombinant plant expression vector prepared by operatively linking a foreign gene encoding a target protein downstream of the promoter, but is not limited thereto Do not. In the present invention, "operably linked" refers to a component of an expression cassette that functions as a unit for expressing a heterologous protein. For example, a promoter operably linked to a heterologous DNA encoding a protein promotes the production of a functional mRNA corresponding to a heterologous DNA.

The recombinant plant expression vector of the present invention can be used as a transient expression vector that can be transiently expressed in a plant into which a foreign gene is introduced and a plant expression vector that can be permanently expressed in a plant into which a foreign gene is introduced.

Binary vectors that can be used in the present invention include RB (right border) and LB (left border) of T-DNA, which can transform a plant when present together with the Ti plasmid of A. tumefaciens . (Cat #: 6018-1, Clontech, USA), pBIN19 (Genbank Accession No. U09365), pBI121, pCAMBIA vector, etc., which are frequently used in the art, may be used It is good.

The term "vector" is used to refer to a DNA fragment (s), nucleic acid molecule, which is transferred into a cell. The vector replicates the DNA and can be independently regenerated in the host cell. The term "carrier" is often used interchangeably with "vector ". The term "expression vector" means a recombinant DNA molecule comprising a desired coding sequence and a suitable nucleic acid sequence necessary for expressing a coding sequence operably linked in a particular host organism. Promoters, enhancers, termination signals and polyadenylation signals available in eukaryotic cells are known.

A preferred example of a plant expression vector is a Ti-plasmid vector that is capable of transferring a so-called T-region into a plant cell when it is present in a suitable host such as Agrobacterium tumefaciens. Other types of Ti-plasmid vectors (see EP 0 116 718 B1) are currently used to transfer hybrid DNA sequences to plant cells or protoplasts in which new plants capable of properly inserting hybrid DNA into the plant's genome can be produced have. A particularly preferred form of the Ti-plasmid vector is a so-called binary vector as claimed in EP 0 120 516 B1 and U.S. Patent No. 4,940,838. Other suitable vectors that can be used to introduce a gene according to the invention into a plant host include viral vectors such as those that can be derived from double-stranded plant viruses (e. G., CaMV) and single- For example, from non -complete plant virus vectors. The use of such vectors may be particularly advantageous when it is difficult to transform the plant host properly.

The expression vector preferably comprises one or more selectable markers. The marker is typically a nucleic acid sequence having a property that can be selected by a chemical method, and includes all genes capable of distinguishing a transformed cell from a non-transformed cell. Examples include antibiotic resistance genes such as herbicide resistance genes such as glyphosate or phosphinotricin, kanamycin, G418, Bleomycin, hygromycin, chloramphenicol, It is not.

In a plant expression vector according to an embodiment of the present invention, the terminator can be a conventional terminator, such as nopaline synthase (NOS), rice α-amylase RAmy1 A terminator, phaseoline terminator, Agro A terminator of the Octopine gene of Agrobacterium tumefaciens or a histone 4 terminator derived from pepper, and most preferably a pepper-derived histone 4 terminator consisting of the nucleotide sequence of SEQ ID NO: 2 , But is not limited thereto.

The present invention also provides a method for producing a plant cell, comprising the steps of: transforming a plant cell with said vector; And

And regenerating a transgenic plant from the transgenic plant cell. The present invention also provides a method of producing a transgenic plant expressing a foreign gene.

The method of the present invention comprises transforming a plant cell with a recombinant vector according to the invention, said transformation being mediated, for example, by Agrobacterium tumefiaciens . In addition, the method of the present invention comprises regenerating a transgenic plant from the transformed plant cell. Any of the methods known in the art can be used for regeneration of transgenic plants from transgenic plant cells.

Transformed plant cells must be regenerated into whole plants. Techniques for the regeneration of mature plants from callus or protoplast cultures are well known in the art for a number of different species (Handbook of Plant Cell Culture, Vol. 1-5, 1983-1989, Momillan, N. Y.).

The foreign gene may be any gene which desires mass expression in a plant, and may be located behind the promoter in the plant expression vector of the present invention and may be expressed by fusion with a reporter gene, if necessary. A method of transforming the recombinant plant expression vector into a plant can be carried out as follows.

Transformation of a plant means any method of transferring DNA to a plant. Such transformation methods do not necessarily have a regeneration and / or tissue culture period. Transformation of plant species is now common for plant species, including both terminal plants as well as dicotyledonous plants. In principle, any transformation method can be used to introduce the hybrid DNA according to the present invention into suitable progenitor cells. The method is based on the calcium / polyethylene glycol method for protoplasts (Krens, FA et al., 1982, Nature 296, 72-74; Negrutiu I. et al., June 1987, Plant Mol. Biol. 8, 363-373) (Shillito RD et al., 1985 Bio / Technol. 3, 1099-1102), microinjection into plant elements (Crossway A. et al., 1986, Mol. Gen. Genet. 202,179-185 (Klein et al., 1987, Nature 327, 70), the infiltration of plants or the transformation of mature pollen or vesicles into Agrobacterium tumefaciens Infection by viruses (non-integrative) in virus-mediated gene transfer (EP 0 301 316), and the like. A preferred method according to the present invention comprises Agrobacterium mediated DNA delivery. Particularly preferred is the use of so-called binary vector techniques as described in EP A 120 516 and U.S. Pat. No. 4,940,838.

"Plant cell" used for transformation of a plant may be any plant cell. The plant cell may be any of a cultured cell, a cultured tissue, a culture or whole plant, preferably a cultured cell, a cultured tissue or culture medium, and more preferably a cultured cell.

"Plant tissue" refers to cells of differentiated or undifferentiated plants, such as, but not limited to, various types of cells used in fruit, stem, leaf, pollen, seed, protoplasts, shoots and callus tissue. The plant tissue may be in planta or may be in an organ culture, tissue culture or cell culture.

The present invention also provides a transgenic plant expressing a foreign gene produced by the above method and a seed thereof.

The recombinant plant expression vector of the present invention can transform any plant regardless of dicotyledonous or monocotyledonous plants. According to one embodiment of the present invention, the plant is selected from the group consisting of Arabidopsis, potato, eggplant, cigarette, pepper, tomato, burdock, ciliaceae, lettuce, bellflower, spinach, modern sweet potato, celery, carrot, parsley, parsley, cabbage, It may be a dicotyledonous plant such as gibbousum, watermelon, melon, cucumber, amber, pak, strawberry, soybean, mung bean, kidney bean or pea or a terminal plant such as rice, barley, wheat, rye, corn, sugar cane, Preferably a dicotyledonous plant, more preferably a cigarette.

The present invention also provides a primer set consisting of SEQ ID NOS: 3 and 4 for amplifying the promoter.

Wherein the primer comprises at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, At least 29, at least 30, at least 31, at least 32, at least 33, at least 34, at least 35, at least 36, at least 37, at least 38, at least 39 consecutive nucleotides Lt; / RTI > oligonucleotides.

Wherein the primer comprises at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, Oligonucleotides consisting of fragments of at least 29, at least 30, at least 31, at least 32, at least 33, at least 34, at least 35, and at least 36 consecutive nucleotides.

In the present invention, a "primer" refers to a single strand oligonucleotide sequence complementary to a nucleic acid strand to be copied, and may serve as a starting point for synthesis of a primer extension product. The length and sequence of the primer should allow the synthesis of the extension product to begin. The specific length and sequence of the primer will depend on the primer usage conditions such as temperature and ionic strength, as well as the complexity of the desired DNA or RNA target.

In the present invention, the oligonucleotide used as a primer may also include a nucleotide analogue such as phosphorothioate, alkylphosphorothioate, or peptide nucleic acid, or And may include an intercalating agent.

Hereinafter, the present invention will be described in detail by way of examples. However, the following examples are illustrative of the present invention, and the present invention is not limited to the following examples.

Materials and methods

One) pepper HISTON 4  Promoters and Terminators Cloning

(5'-ATCCAAGCTCAAGCTGACGTCTCAGCTTAGAAATAGACAG-3 '; SEQ ID NO: 3) and ppro16 (5'-CATGGATCCATTTAAATTTTCACACACAGAAATTTGA-3'; SEQ ID NO: 4) were used as a gene-specific primer set for cloning Capsicum annuum histone 4 promoter Respectively. Genome-specific primer sets for cloning the pepper histone 4 terminator were prepared using ppro17 (5'-TGACTCGAGGGCGCGCCAGCAGTTGTTAGGTAGTAG-3 '; SEQ ID NO: 5) and ppro18 (5'-AGGCTAATTCTGGGGACCCTAGGATAAATCAATATCAAGGTAATC-3' DNA was amplified by polymerase chain reaction (PCR). The PCR reaction mixture consisted of 200 ng of DNA, 2X PrimeSTAR (Takara, Japan) and 10 pmole of each primer. The PCR conditions included initial denaturation at 95 ° C for 3 minutes; 30 cycles of denaturation at 94 DEG C for 15 seconds, annealing at 55 DEG C for 20 seconds, and polymerization at 72 DEG C for 2 minutes; Followed by an additional 5 minute polymerisation at 72 ° C using an Applied Biosystem GeneAmp PCR system 9700 amplifier.

In addition, ppro1 (5'-ATTTAAATGGATCCATGGTAGATCTGACTAGTAAAG-3 '; SEQ ID NO: 7) and ppro2 (5'-GGCGCGCCCTCGAGTCACACGTGGTGGTGGTGGTG-3'; SEQ ID NO: 8) were used as a gene specific primer set for cloning the GFP gene and pCAMBIA1302 plasmid DNA was amplified by polymerase chain reaction (PCR). The PCR reaction mixture consisted of 1 μg of plasmid DNA, 2 × PrimeSTAR (Takara, Japan) and 10 pmole of each primer. The PCR conditions included initial denaturation at 95 ° C for 3 minutes; 30 cycles of denaturation at 94 DEG C for 15 seconds, annealing at 55 DEG C for 20 seconds, and polymerization at 72 DEG C for 2 minutes; Followed by an additional 5 minute polymerization at 72 ° C using an Applied Biosystem GeneAmp PCR system 9700 amplifier.

The amplification product of the promoter, terminator and GFP produced by the PCR reaction was purified using a Gel Purification Kit (Gendocs, Korea), and overlapped PCR was performed to bind it again to one gene set . For the overlap PCR, ppro15 (SEQ ID NO: 3) and ppro18 (SEQ ID NO: 6) primers were used. The PCR reaction mixture consisted of 1 μl each of the PCR products of the promoter, terminator and GFP obtained through each PCR amplification, 2 × PrimeSTAR (Takara, Japan) and 10 pmole of each primer. The PCR conditions included initial denaturation at 95 ° C for 3 minutes; 30 cycles of denaturation at 94 DEG C for 15 seconds, annealing at 55 DEG C for 20 seconds, and polymerization at 72 DEG C for 2 minutes; Followed by an additional 5 minute polymerization at 72 ° C using an Applied Biosystem GeneAmp PCR system 9700 amplifier. Thereafter, the product bound with the promoter: GFP: terminator produced by the PCR reaction was purified using a gel purification kit.

2) pepper HISTON 4  Production of promoter expression vector

The reporter gene encoding the fluorescent marker protein, mGFP, and the DNA fusion construct conjugated with the pepper promoter were constructed by overlap PCR as described above, and the resulting PCR products were restricted using a cold fusion cloning kit (SBI, USA) It was inserted into the pPZP200 vector digested with the enzyme Bam HI, and H i nd ⅲ. The cold fusion cloning method is as follows. 1 μl of the promoter: GFP: terminator product obtained previously by PCR, 1 μl of pPZP200 vector digested with Bam HI / Hind III enzyme, and 5 × cold fusion cloning kit master mix were mixed and incubated for 5 minutes at room temperature and for 10 minutes at low temperature To give the final cloning product.

3) Tobacco transformation using Agrobacterium

Infiltration with Agrobacterium was performed to determine the expression level of the histone 4 promoter using transient expression, which is a method of obtaining a recombinant protein.

Six-week tobacco ( Nicotiana benthamiana) liquid culture of Agrobacterium two kinds (pCAMBIA1302 and Histone4 the leaves: after GFP) to 10mM MgCl _2, 10mM MES and 500μM acetonitrile ache Gon (acetosyringone) were suspended in solution using a syringe and were inoculated on the back of the tobacco leaf. Five days after the inoculation, the leaves inoculated with Agrobacterium were collected and RNA was extracted using an RNA extraction kit (Gendocs, Korea) and cDNA was synthesized after treatment with DNase (Promega, USA). 1 μl of total RNA, 10 μl of Oligo dT and 1 μl of 10 mM dNTPs were added to each well and incubated at 65 ° C. for 5 minutes. The reaction was carried out on ice for 5 minutes. 5 × PrimeScript buffer, 0.5 μl of RNase inhibitor and 40 μl of PrimeScript RTase / ㎕) (Takara, Japan) was added and reacted at 42 캜 for 60 minutes and then used for PCR reaction.

(5'-CGGCCTCGAGGTCGACCCCGGGTCTAGAGCCATGTGTAATCCCAGCAG-3 '; SEQ ID NO: 10) primer set was used to confirm the amount of GFP expression in the transgenic tobacco plants PCR was performed. As a control, tobacco transformed with the pCAMBIA1302 vector expressing the CaMV35S promoter was used. The PCR conditions included initial denaturation at 95 ° C for 3 minutes; 30 cycles of denaturation at 95 DEG C for 30 seconds, annealing at 55 DEG C for 30 seconds and polymerization at 72 DEG C for 1 minute; Followed by an additional 5 minute polymerization at 72 ° C using an Applied Biosystem GeneAmp PCR system 9700 amplifier and the amplified final product from the PCR reaction was confirmed by electrophoresis on a 1% agarose gel.

Example  1. Through tobacco transformation HISTON 4  Measurement of transient expression level of promoter

Using the agro-infiltration method, tobacco ( Nicotiana The expression of GFP was confirmed by RT-PCR after transient expression of the GFP gene in B. benthamiana .

As a result of RT-PCR, as shown in FIG. 2, it was confirmed that GFP gene showed high expression level in Histone 4: GFP inoculated tobacco leaves. As a result, it was confirmed that the histone 4 promoter of the present invention normally expressed gene . In addition, the amount of GFP expression in the leaf inoculated with the histone 4 promoter was significantly higher than that of the tobacco leaf inoculated with pCAMBIA1302 used as a control. Thus, it was confirmed that the pepper histone 4 promoter was a promoter having higher expression efficiency than the control CaMV35S promoter.

<110> GenDocs, Inc. <120> Histone4 promoter isolated from Capsicum annuum and uses thereof <130> PN12270 <160> 10 <170> Kopatentin 1.71 <210> 1 <211> 1500 <212> DNA <213> Capsicum annuum <400> 1 tcagcttaga aatagacagg atattcttct gtcaaactct aaacttcaca acgtgtgaaa 60 tcagaagaca acaagttccc tgcttttatg cataagactt gagcaaatac caattccatc 120 ctaagaaatt gacatgaatc agatagacga gttacacaga ggaaaataaa taagcactgc 180 aaaagatatg ggagaccgcc ccatatcttc agggtgagag cgaaaatcga aaaccattag 240 gttcacaatt ttcaaacgta tgttaagtag caaaacatca tctttcccaa tactaaagtt 300 cagtagcatt tagtgtcaag taaaatataa ggaaatagat cagtaatcta gtagtaactt 360 ttaactagtc gattatactc aaatgtttga cagatgactt gtatccagaa attaaaaaca 420 aactaagagc atatagagaa tggtagtact gcaatctcaa aacagaatac cacacaatta 480 tggaattagc attttgcgta agtatgaagt ctaattactc tattcaaaaa gtgtatatct 540 ttctgcattt tgatcagcat gaataggcta tcaatgtatg cattttcatc attagttggt 600 tcatattatc tatatcatca gccagttgat caacatcttc attcagttca ttggggtgcc 660 tccatcttga tcatgatttt gttccctttt ggaattggag cttgtttgct tgtggcgttt 720 caaagaaatt catgcgagct ttgatctacc gtaaataata tcatattatg ttaaaatatc 780 ttaaattatt ttagtccaac aattgatatg agtctatggt tgactaagga gatgggggag 840 tgatgatgta agtcccaact tagttccttt gcccgtttac ttgttgatgt tctattctta 900 tctatagttt taaagatgca tacataaaaa aagaaaagta tttatggttg tattagccac 960 aactactcgg tcgatttttt tttttttata taatactttt gtacatccta ttttatatga 1020 cacacttttc tttttaatcc gtccaaaaaa aatctgacct cactataatt agaaaaaaaa 1080 atatgtaact ttaaaatttt ccttaatgaa atgatttgca atcatataaa cattcaaaac 1140 taatttcaaa ctacatattt aaaaatcaca taaaatgaaa cggaggaagt aataattaaa 1200 tagcaacgag ctgcgagaaa gagaaaacaa atgcctaaat cttctttcac atggatcgta 1260 tccccgtcca atcatattca gtttccgcct ctcacatgga tcgacaattc tatccgtcgg 1320 attaacaagt aaatccccaa atatttaacg gctgcaattt accccccgcc acatttcgaa 1380 ccgcgaaaat tcacggcgcc caaatccccc aaatataaat atccaccacc atctacttta 1440 cttttcattc aaattcaaaa ccctagtatc tttttcttct tcaaatttct gtgtgtgaaa 1500                                                                         1500 <210> 2 <211> 500 <212> DNA <213> Capsicum annuum <400> 2 agcagttgtt aggtagtaga tgtagatgtg atggaaattt atgctgtttc tgtattttag 60 tttctatgaa gcgttgttct actttttaag ctttggatat tgtaatttct cattgcttat 120 tcaatgaaca atgttcagtt tctacaaatt tcaattgtta ttttccatct cactagtatt 180 gttgataatt ggttgaaatg acttttacta tctgtgcaat tcatatttat acatttgtga 240 aatgcatcaa ttttcattcc atatgagttc tttccatttc tttgttgttg agaaagctga 300 gttgtgggat aagcattaac tttgtggcca cagtatctaa gtttcttgct tgagagattc 360 aaactggaga cgatcatgtt ttttattttt taattattat tcacttgcct tttttgcaca 420 gtttctttgt taatttgaat ggctagactc tgtttaattg tagtttattt tagtagctga 480 ttaccttgat attgatttat 500 <210> 3 <211> 40 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 3 atccaagctc aagctgacgt ctcagcttag aaatagacag 40 <210> 4 <211> 37 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 4 catggatcca tttaaatttt cacacacaga aatttga 37 <210> 5 <211> 36 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 5 tgactcgagg gcgcgccagc agttgttagg tagtag 36 <210> 6 <211> 45 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 6 aggctaattc tggggaccct aggataaatc aatatcaagg taatc 45 <210> 7 <211> 36 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 7 atttaaatgg atccatggta gatctgacta gtaaag 36 <210> 8 <211> 35 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 8 ggcgcgccct cgagtcacac gtggtggtgg tggtg 35 <210> 9 <211> 49 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 9 cggcggatcc ccatgggata tcggtacctg gtgatgttaa tgggcacaa 49 <210> 10 <211> 48 <212> DNA <213> Artificial Sequence <220> <223> primer <400> 10 cggcctcgag gtcgaccccg ggtctagagc catgtgtaat cccagcag 48

Claims (9)

A histone 4 promoter derived from a pepper ( Capsicum annuum ) comprising the nucleotide sequence of SEQ ID NO: 1. A histone 4 terminator derived from the pepper ( Capsicum annuum ) consisting of the nucleotide sequence of SEQ ID NO: 2. A recombinant vector comprising the promoter of claim 1. A recombinant vector comprising the terminator of claim 2. 4. The recombinant vector according to claim 3, wherein the recombinant vector is produced by operatively linking a foreign gene encoding a target protein downstream of the promoter. 4. The recombinant vector according to claim 3, wherein the vector comprises a pepper-derived histone 4 terminator consisting of the nucleotide sequence of SEQ ID NO: 2. delete delete delete

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