CN1605589A - PC-1 gene promoter and its application - Google Patents

Abstract

The present invention discloses PC-1 gene promoter and its application. The PC-1 gene promoter of the present invention has one of the following nucleotide sequence: the DNA sequence of SEQ ID No. 1 in the sequence list and the DNA sequence with 70 % over homology with the DNA sequence limited by SEQ ID No. 1 in the sequence list and the same function. The promoter of the present invention has important function in the gene treatment of prostate cancer and breast cancer, especially high malignant prostate cancer, as well as important theoretic significance and practical significance.

Description

PC-1 gene promoter and application thereof

Technical field

The present invention relates to a kind of promotor and application thereof, particularly a kind of promotor of PC-1 gene and application thereof.

Background technology

Prostate cancer is the highest malignant tumour of American-European countries's sickness rate, calendar year 2001 U.S.'s number of being diagnosed as prostate cancer be 198,100 people, about 31,500 people's death.The U.S.'s cost of every year in prostate cancer therapy is 4,600,000,000 dollars.In addition, in the world without any a kind of tumour as prostate cancer, sickness rate increases significantly with the age and rises.According to statistics, to mid-21st Century, China more than 60 years old the aged will be increased to about 400,000,000.Prostate cancer also will become one of important diseases that influences men's health.Treatment prostate cancer medicine will face huge market.

Prostate gland is the androgen-dependent organ, and male sex hormone is the promoting factor of prostate cancer progress.The development of prostate cancer is a predictable process normally, initial stage is the male sex hormone sensitiveness normally, final cancer cells will change non-sensitive state over to by sensitiveness, promptly change the acme of deterioration over to, at present without any the methods of treatment at this stage.Prostate specific antigen (PSA) expression of gene rises with the prostate cancer progress, the inspection of blood-serum P SA concentration has been one of most widely used index in the prostate cancer clinical diagnosis, the PSA gene is only expressed in prostate organs, so the PSA promotor has the application prospect of particularly important.Be applied to exploring prostate cancer cell and change the mechanism of non-sensitive state over to, seek internal secretion opposing type prostate cancer therapy new way by the male sex hormone sensitiveness; Be applied to the prostate cancer gene therapy; Be applied to set up the animal model of prostate cancer.

The PC-1 gene is the new gene that the inventor at first is cloned into from prostate cancer cell, and full length cDNA sequence has been logined GenBank (AF202897).At aspects such as prostate organs tissue specific expression and PSA gene closely similar characteristic is arranged.Different is, PC-1 genetic expression can be induced the normal cell vicious transformation, and the PSA gene then can not.

Promotor is a necessary for gene expression, has determined the intensity etc. of space, time and the expression of exogenous gene expression, and genetic treatment of tumor is had vital role.

The innovation and creation content

The purpose of this invention is to provide a kind of PC-1 gene promoter.

PC-1 gene promoter provided by the present invention is one of following nucleotide sequences:

1) SEQ ID № in the sequence table: 1 dna sequence dna;

2) with sequence table in SEQ ID №: 1 dna sequence dna that limits has 70% above homology, and has the dna sequence dna of identical function.

SEQ ID № in the sequence table: 1 dna sequence dna is by 4398 based compositions; The enhancement sequences of transcribing that 1-1110 bit base of 5 ｀ of sequence 1 end is the PC-1 gene promoter in the sequence table.

Experiment showed, the also optional SEQ ID № in sequence table of described PC-1 gene promoter: 2 and SEQ ID №: 3.

The expression cassette, expression vector and the clone that contain described PC-1 gene promoter also belong to protection scope of the present invention.

Experimental results show that, promotor of the present invention is only expressed luciferase reporter gene in prostate cancer cell and breast cancer cell, express in the prostate cancer cell of specifically positive and estrogen receptor positive and the breast cancer cell at androgen receptor, in prostate cancer cell PC-3, the DU145 of androgen receptor feminine gender, do not express, in other tissue-derived tumour cells, do not express yet; And expression amount significantly increases with the prostate cancer progress; Point out promotor of the present invention to can be used as director element important in the gene therapy vector, be fit to the prostate cancer and the mastocarcinoma gene treatment of the androgen receptor positive and estrogen receptor positive, the gene therapy effect will significantly increase with tumour progression.With the PSA promotor relatively, the intensity of PC-1 gene promoter is identical among the prostate cancer cell LNCaP in the early stage, is higher than the PSA promotor in male sex hormone prostate cancer cell C4-2 non-sensitive, highly deterioration.Has stronger effect in the prostate cancer gene therapy that PC-1 gene promoter of the present invention carries therapeutic gene will be pernicious at height, male sex hormone is non-sensitive.Promotor of the present invention will play an important role in the gene therapy of prostate cancer and the particularly high malignant prostate cancer of mammary cancer, have important significance for theories and practical significance.

Description of drawings

Fig. 1 is the 5kb promoter sequence

Fig. 2 analyzes the cell specific expression histogram that the 5kb promotor instructs for luciferase reporter gene

Fig. 3 analyzes the cell specific expression photo that the 5kb promotor instructs for the green fluorescent protein reporter gene

The expression histogram of luciferase reporter gene in LNCaP and C42 cell that Fig. 4 instructs for the different lengths promotor

Fig. 5 is that the activity and the 5kb promoter activity of hybrid promoter compares histogram

Fig. 6 is that PC-1 promotor and PSA promoter activity compare

Embodiment

Material and analytical procedure

(1) bacterial strain, cell strain, plasmid: Human Prostate Cancer Cells strain C4-2, C4-2B, LNCaP, PC-3, DU145 and plasmid p61/pGL3 (the Yeung F that carries PSA promotor and luciferase reporter gene, LiX, EllettJ, Trapman J, Kao C, Chung LW.Regions of prostate-specific antigen (PSA) promoter conferandrogen-independent expression of PSA in prostate cancer cells.J Biol Chem.2000 Dec29; 275 (52): 40846-55.); The pGL3 series plasmid and the luciferase confidential reference items control plasmid pRL-TK of used promoter activity analysis are Promega company product.Cell strain 293, NIH3T3, HL7-21,, MCF-7, B16, HepG2, Hela cell be U.S. ATCC company product (www.atcc.org); Plasmid pEGFP-N1 is an American I NVITRIGEN company product (www.invitrigen.com).

(2) primer

Primers F: 5 '-CCGCTCGAGTCCAGACAAACGCAG-3 '

WJ750： 5’-CGACGCGTTGAGTGCATGTAACTGAAG-3’

WJ340： 5’-CGACGCGTGCCATATTGCAGAACCCT-3’

WJ1099： 5’-CGACGCG?TTAGATTAGAACACTGCCT-3’

WJ1337： 5’-CGACGCGTTAGAGTACAACATGAGAC-3’

WJ1579： 5’-CGACGCGTAGTTTGAGACCAGCCTG-3’

WJ1831： 5’-CGACGCGTAAGAAGCCAGACAATCC-3’

WJ3K： 5’-CGACGCGTTATGCCTGCAGTGCATC-3’

WJ4K： 5’-CGACGCGTAGATCTGAAGCACAGGC-3’

WJ5K： 5’-CGACGCGTACACTGACATATTTGAGTG-3’

E4K:5 '-CGACGCGTGCCTGTGCTTCAGATC-3 ' (complementary strand of primer WJ4K)

E:5 '-CACTCAAATATGTCAGTGTACGCGTCG-3 ' (complementary strand of primer WJ5K)

EGFPI： 5’-CATCCATGGATGGTGAGCAAGGGCGAG

EGFPII： 5’-TAATCTAGATTACTTGTACAGCTCGTC

(3) statistical analysis adopts the t check

The preparation of embodiment 1, PC-1 gene promoter and active detection the thereof

1, the structure of recombinant plasmid dna

Carry out following operation with reference to the molecular cloning experiment guide third edition:

(1) structure of pWJ340, pWJ750, pWJ1099, pWJ1337, pWJ1579, pWJ1831, pWJ3K, pWJ4K, pWJ5K

Be research PC-1 gene promoter activity, extract the genomic dna of C4-2 cell, with this genomic dna as template, with primer WJ5K and primers F, pcr amplification goes out the dna fragmentation of the preceding 4938bp of PC-1 gene translation initiation site, its sequence is as SEQ ID № in the sequence table: shown in 1, with its with restriction enzyme MluI and XhoI cutting after, fragment after reclaiming purifying directed cloning to containing luciferase reporter gene but do not contain on the plasmid pGL3-Basic of promotor.Identify positive colony through colony polymerase chain reaction (PCR) method and plasmid double digestion, called after pWJ5K.The plasmid pWJ5K DNA that extracts with Promega company plasmid extraction kit is a template, go out the promoter dna fragment of the different length of 5 ' flank by pcr amplification, and use with the quadrat method directed cloning to carrier pGL3-Basic plasmid with primer WJ340, WJ750, WJ1099, WJ1337, WJ1579, WJ1831, WJ3K, WJ4K respectively.Set up respectively contain PC-1 gene translation initiation site to upstream-340bp ,-750bp ,-1099bp ,-1337bp ,-1579bp ,-1831bp ,-3kb and-recombinant plasmid of 4kb dna fragmentation, called after pWJ340, pWJ750, pWJ1099, pWJ1337, pWJ1579 and pWJ1831, pWJ3K, pWJ4K respectively.

(2) structure of pWJ-KE+ and pWJ-KE-

Be the enhancement region of transcribing on the research promotor, with primer E and primer WJ4K is the fragment that masterplate amplifies 1.1Kb with pWJ5K, this fragment is the sequence between the 5kb-4kb before the PC-1 gene translation initiation site, cut rear clone to the plasmid PWJ1831 that cuts through same enzyme with the MluI enzyme, cut by enzyme and identify and sequencing result shows the front end that is inserted into PC-1 gene promoter area 1831bp that this sequence is forward and reverse respectively.Recombinant plasmid called after pWJ-KE+ and pWJ-KE-.

(3) structure of pWJ5KB-EGFP

The tissue specificity of transcribing for the long PC-1 promoter region of research 5kb, with plasmid pEGFP-N1 is masterplate, amplify the gene of the green fluorescent protein about 1700bp with primer EGFPI and EGFPII, cut, reclaim the big fragment of removing behind the luciferase with plasmid pGL3-Basic with same enzyme behind NcoI and the XbaI enzyme cutting and is connected back structure plasmid pWJBasic-EGFP.Primers F and primer E are template with pWJ5K, the PCR product that obtains is cut rear clone to the plasmid pWJ-EGFP that cuts through same enzyme through MluI and XhoI enzyme, has made up the recombinant plasmid called after pWJ5KB-EGFP that contains 1700bp left and right sides green fluorescence protein gene and the long PC-1 promotor of 5kb.

Above recombinant plasmid is all identified through sequential analysis.Residing position is as shown in Figure 1 in the PC-1 gene upstream sequence for the primer.

2, the expression characterization of 5kb promotor and activation analysis

(1) luciferase reporter gene is analyzed the cell specific expression that the 5kb promotor instructs

With 293, HepG2, NIH3T3, B16, HL7-21, MCF-7, HeLa, PC3, DU145, LNCaP and C4-2B tumor cell line cell be inoculated in respectively in 24 orifice plates, when treating that cell grows to the density that is paved with 60%-80%, use no RPMI-1640 nutrient solution phenol red and serum instead and cultivate 10h, add in the mixed solution of 20 μ lHBS and 10 μ l liposome DOTAP after plasmid that 0.5 μ g is to be detected and 0.1 μ g HBS (pH7.4) mixing as interior target pRL-TK plasmid and 20mmol/L, room temperature is placed 15min and is formed the DNA-DOTAP complex body.After the RPMI-1640 dilution with 500ul, add in 24 orifice plates.37 ℃ hatch 5h after, change the RPMI-1640 nutrient solution that normally contains 10% foetal calf serum into.Collecting cell behind the 48h, the activity of carrying out luciferase detects.The cell in each hole is transferred in the centrifuge tube of 1.5ml the 1min that vibrates, centrifugal then 3min after successively giving a baby a bath on the third day after its birth time with the PBS of 1ml behind the cell pyrolysis liquid cracking 15min with 500 μ l.After the Firely luciferase substrate of getting the last cleer and peaceful 100 μ l of 20 μ l mixes, measure the luminous value of luciferase with luminometer, the reaction terminating liquid that adds 100 μ l, mensuration is as the luminous value of interior target Renilla luciferase, and both ratio is the relative reactivity RLA (Relative Luciferase Activity) of luciferase.The numerical value of RLA is 3 repeated experiments results' mean+SD.

The result as shown in Figure 2, show that the 5kb promotor has the ability that makes luciferase reporter gene specificity overexpression in the androgen receptor positive, high virulent prostate cancer cell C4-2B, at elementary prostate cancer cell LNCaP with in elementary breast cancer cell MCF-7 a small amount of expression is arranged, in other tumour cells, do not express.

(2) the green fluorescent protein reporter gene is analyzed the cell specific expression that the 5kb promotor instructs

With length is the PC-1 gene promoter insertion green fluorescent protein reporter gene upstream of 5kb, transfection 293 respectively, HepG2, NIH3T3, B16, MCF-7 and C4-2B tumour cell, pass through fluorescence microscope, studied the tissue specificity of promotor, the result as shown in Figure 3, show in the C4-2B cell, to show strong green fluorescence, in elementary breast cancer cell MCF-7, faint green fluorescence is arranged, in other cells, do not express.

(3) expression of luciferase reporter gene in LNCaP and C42 cell of different lengths promotor guidance

The detection of plasmid transfection, two luciferase report system is with (1) in the step 2.

Studied length to be respectively-340bp ,-750bp ,-1099bp ,-1337bp ,-1579bp ,-1831bp ,-3kp ,-4kp and-expression of the luciferase reporter gene that the promoter sequence of 5kp instructs, the result as shown in Figure 4, show that the gene expression dose that the regulating and controlling sequence from translation initiation site to upstream-4kp instructs maintains a lower and relative constant level, after the length of regulating and controlling sequence increased to 5kb, the expression level of luciferase reporter gene significantly rises, and expression level increases about 3 times approximately.Infer that translation site upstream-4kb is transcribing enhancement sequences to-5kb region memory.For verifying this hypothesis, general-4kb to-5kb sequence with forward with oppositely after the insertion-1831bp sequence promotor, made up recombinant plasmid pWJ-KE+ and pWJ-KE-, the result shows that the activity of two kinds of hybrid promoters and complete 5kb promoter activity are suitable as shown in Figure 5.More than studies have shown that-4kb exists to-5kb the sequence and transcribes enhancement sequences, makes the 5kb promoter activity significantly increase.PC-1 gene 1 to-2940bp promoter sequence as SEQ ID № in the sequence table: 2 and SEQ ID №: shown in 3.

(4) with the comparison of PSA promoter activity

The research of PSA promotor is more deep, and application prospect has obtained abundant affirmation.For whether the intensity of determining the PC-1 gene promoter is enough to reach application need, particularly the effect in the non-sensitive prostate cancer gene therapy of male sex hormone has been carried out two kinds of promoter activities relatively.C4-2 is the higher prostate cancer cell of identical grade malignancy with LNCaP genetic background.For the intensity of research PC-1 gene promoter whether relevant with the grade malignancy of prostate cancer, PC-1 gene 5Kb promoter sequence is inserted in the luciferase reporter gene upstream respectively, with p61/pGL3 together according to the method transfection C4-2 and the LNCaP cell of (1) in the step 2.The physiological environment that male sex hormone is removed fully when simulating endocrine therapy has clinically been carried out expression analysis at serum-free condition.The result shows that among the prostate cancer cell LNCaP, the PSA promotor is identical with the 5Kb promotor intensity of PC-1 gene in the early stage.Along with grade malignancy increases, remarkable ascendant trend all appears in the activity of two kinds of promotors.The result as shown in Figure 6, show male sex hormone non-sensitive, androgen receptor is positive and the prostate cancer cell C4-2B of highly deterioration in, the intensity of PC-1 gene 5Kb promotor is significantly higher than the PSA promotor, and the expression level of luciferase reporter gene rises about 6 times.Prompting PC-1 promotor is carried therapeutic gene at more effective in the gene therapy that produces endocrine therapy opposing patient.

Sequence table

<160>3

<210>1

<211>4938

<212>DNA

＜213〉artificial sequence

<220>

<223>

<400>1

tacactgaca?tatttgagtg?atgagttaac?aggcagacag?aaacttaatt?ttctttatag 60

tcatttgtac?ctactaccat?ttccgtttgc?aatgacatga?gtcgtctccg?gttattaccc 120

tgcagtacat?acgcacttta?ctggctgcac?ctagttgact?tctgaactgc?ccttgggctc 180

cctctgctgg?catagtggtg?cgtgagcact?tttctggggt?catggaatgg?atttctcttt 240

ctgcccattt?tctttctttt?ggtttgtggt?tttctctctc?ataaattatt?acacaaacat 300

ggaccataca?atacaacctt?ctttcaaacc?tgatttgttt?gctaactgta?tcagtacctt 360

tccatataaa?attcataaat?atttttatat?aattatgtca?gtgtttccat?atttttctat 420

tacctgaatg?ttctttttag?catttaaata?atccattttt?agtaggcatt?tgtattgctt 480

ttttccattt?caggtagcat?ctaaagtgaa?tgtcccttgc?atgtatacta?ttcatatgtg 540

acttaataag?tagagcttgc?taaatgggta?ggaacttaca?ggaaaagggg?aaactctagt 600

ttgagaaaat?tgtctgggct?aagtttagga?tgtgttcaac?aaacaggaaa?gtcaagtttg 660

gctaaagcat?ggttgagtaa?tagtgggaaa?tatggtcaga?aagtaggtat?gttaggactc 720

tttctactga?aatgatgaaa?ttcaactcca?gccacaagtg?gagcagaaca?aaattgtgtt 780

tgtttaataa?ctgggaagcc?cagggtagct?ggaggcaggg?gcttagatgc?tgctgcgcca 840

tcccttacct?gtctgtttct?gtttctcctt?ctgtcccttc?ccagtctcag?cactgagtct 900

cttgcccatt?ggcctggtga?gggaaggagc?tgccagcccc?acccaacagc?tcaggttaca 960

gagagagtca?ctttcttcca?ttactcacag?agtaaacatc?aaggaaggcc?actgattgat 1020

tgacagtgtc?tgggtcagat?gtctatcctt?aggccagtcc?ctgtgaacaa?ggggatgggg 1080

tgtctgcgtg?gaccagatct?gaagcacagg?cccatgcctg?gggccagggg?gtgggaacta 1140

tggacctctc?tccccactga?gaaccccagg?gagcaggtga?ggtgaaattc?ctctagggga 1200

agaggggcaa?aattgacaag?atagcagatg?tctaccatac?tgctgtgggg?cctggtccct 1260

cccagaagga?aaaacatagt?aacaatagag?tgggtctcac?cctccacctg?ggtctcaagt 1320

agggtgtgga?tgaggacaat?ggaaatgaag?gaaaggttag?aaggcctgtg?gtaccggttg 1380

gtagatagct?cttcgtgctt?tctccatatg?gagtgagagt?gcttggatgt?gattccttca 1440

aagtcaggtc?taggagactc?aggatgccta?atctagaggt?aagaacattg?tgaggaaagc 1500

cagtgaattc?agtcttgtgc?atgctgactt?tgaagtactt?ttggaagagc?caagtggaat 1560

tatccacagg?acaggaccaa?atcttacctg?gttcttcccc?aggccgacta?gtccacaaca 1620

ggaaataaaa?agagttgccc?cgataccaag?gtgtactagt?ccattctcac?actgctatgg 1680

ggaaatacct?gagactgggt?aatttataaa?gggaaaaggt?ttaattgact?cacagttcta 1740

gatggctggg?gaggcttcag?gaaacttaca?atcatggcag?aaggcaccac?ttcacagggt 1800

ggcgggagag?agaatgagtg?cccagcgaag?ggagaagctc?cttataaaac?catctgttct 1860

ccttataaag?atctcttaat?aaaaccgtca?gagaactatc?tcattcacta?tcaggagaag 1920

agcatggggg?aaccgccccc?atgattcagt?ttactccacc?tggtcccgcc?cttgacatgt 1980

gggtgttatt?acaatttaag?gtgagatttg?ggtggggaca?cagagccaaa?ccatatcaca 2040

aggctttctc?ctccttgctg?ggattgtacc?catagcctct?ttctgagtcc?tctctctttt 2100

agcctcttta?tgcctgcagt?gcatccttat?accatttcta?gagtcatctt?tataaaactt 2160

atactctccg?tatgactcat?aaatcctgtt?tttttttttg?cacagtatat?taagtataaa 2220

atttgttaaa?gtctttaatg?gtctgccccc?aagctacatt?tccattttgt?atgtctttca 2280

gttcctttct?actttgtatt?tggctgttga?gttaactgaa?tttttgccat?tccattaacc 2340

catcccatgc?ttttcccact?tctagatttc?acttttcttg?taggctagaa?tgtcttgact 2400

gggatctgac?tggagataat?gagaacaaaa?actggttcaa?agagccagga?tgttgcataa 2460

aagtcctaag?attgtatcta?agcaggtaaa?ataaaaattt?taggcaatta?cttaaatttg 2520

aaatgctcac?atttattaat?aaggcatgta?acatctacat?gagccatcat?ttgctttttt 2580

aattccacat?tgattaggag?ccaaaccttc?agggcaggta?tccggtagag?cgccctggag 2640

aggccctgga?taggcacagg?cgcctgtcag?ggggctcttc?acatgctgtg?tgctgctgct 2700

gggagaagag?ggggccagag?actagggggc?ttctaagaag?aggtggcatt?tctgcctcag 2760

tgttgaagga?tgaataactt?tgacaggctg?gaaaaaggtg?acatttcagg?tagagcgtgt 2820

cacatggatg?taaataccaa?aggtcaagga?catgggcttg?agagatggtg?agaaggatgg 2880

aggtgactgt?ggcttgcatt?ctatccgtat?cactattaat?taccttctaa?tgcctttggc 2940

tctaggtggt?ggaacaagta?aagtaatgga?caaatacttt?ttctaccaat?atttagtgac 3000

caaatgcaga?gttatggaga?gggccaggga?cctcatgaac?catactcttt?ctagtctagg 3060

gacataactc?caatgccttt?cctgtcccag?taagaggcca?tggatttcaa?gaagccagac 3120

aatccattct?ttcagataat?gataaaaaag?aaaccattta?ttttatttct?aagtatagaa 3180

tgaaacattt?atagttgccc?aaattttggt?accttttagg?agaaaaatac?agattttttt 3240

gttgttaaaa?ataaacttaa?aaaaaaaaaa?aaaagactta?ccatagtccg?ggcacggtgg 3300

ctcacacctg?taatccgaga?actttgggag?gccgaggcag?gcggatcacc?tgaggtcagg 3360

agtttgagac?cagcctggcc?aacatggcga?aaccccatct?ctactaaaaa?tacaaaaatt 3420

agccgggcat?agtgggtggt?gggtgcctgt?aatcccagct?acttgggagg?ctgaggcagg 3480

agaatcactt?gaacccagaa?ggtggaggtt?gcagtgagct?gagattgtgc?cattgtactc 3540

cagcctggtc?acaagagcga?agctctgtat?cgaaaacaaa?acaaaaaaag?acctactgta 3600

aatagagtac?aacatgagac?taacaaaaat?aacaaataaa?atcatccagc?atatattctg 3660

tatttaaaaa?aaaaaaatca?caagatgaat?acagaccatc?ttggtgcata?tgtattttat 3720

acactagaca?catgctgatg?tttttaatgc?ttaaaatata?ttaggtttca?tacttgcttc 3780

caatccaagg?agagtatatc?tgagctcttt?ttttttagat?ttgaaaaccg?ccttttagat 3840

tagattagaa?cactgccttt?ttttttgtta?tgcagtaaca?gatcattgcc?cacctgaaag 3900

cgagtcgcct?ttatttttac?ctccttagta?gaaccagcag?gagctgcaat?tgctgttgct 3960

tgagccaagg?ccacagtcag?catagaaagg?ttctgctaag?agactgtgtt?aaaggaaata 4020

aatttggtga?cttagttttg?ttttacaagc?tagctgtggc?ccagtccttc?tgcagggtgg 4080

gtacgcaggt?ttattgcagc?tctgcagagc?catggcctta?gacaaacaaa?gaaggtgtgg 4140

gaatacccgg?gagctttcca?ttcaaagtag?attcttctcc?tttgagtgca?tgtaactgaa 4200

gcaccagctt?tctaaaaaaa?aaaaaaaaaa?aaaaaagaga?gagaaaagaa?aagtcagtct 4260

gaagtttttg?tgacactcag?ccagactcca?cattagataa?agtaatcatg?gaaatacagc 4320

tttattttta?taccaagttg?tgcttagaaa?gctgattgct?aaaccaggaa?gcttggcttt 4380

cagaatttta?ctgtaatcca?gggtggaaaa?aaagtatcac?tcagatggca?tttgtgggag 4440

attactgact?ggtaatccct?ccttgtccta?gaagattaag?tctgccctgg?tgtagtcatg 4500

gtcagctgat?ttttttattt?atttatttta?aataaattgt?agtttcttgt?tgtaagtcag 4560

ccagtgtctg?atgtttgcct?gaactgtttg?tacctctggg?ccatattgca?gaaccctgcc 4620

cttctttgtt?gactgaggaa?agctcgctcc?ctgcccaggt?ttttcattgt?tgatcgaaat 4680

taacaccagg?tggtgaatag?agcccctcct?aaggttgctc?aggataaatc?atttattaaa 4740

taggtctgct?tatcaggagg?ggcgtgaagg?ctcccaaaag?gaaatgctgg?cacctgggcc 4800

cagaagccag?ggcctctaac?tcctggggtt?gatttcttca?gtgaagttgc?accctacaaa 4860

gggaatatgg?ccaaagcggc?actcaactga?aggctgatat?caggcgatta?gacagccatg 4920

cattctgcgt?ttgtctgg 4938

<210>2

<211>2940

<212>DNA

＜213〉artificial sequence

<220>

<223>

<400>2

tacactgaca?tatttgagtg?atgagttaac?aggcagacag?aaacttaatt?ttctttatag 60

tcatttgtac?ctactaccat?ttccgtttgc?aatgacatga?gtcgtctccg?gttattaccc 120

tgcagtacat?acgcacttta?ctggctgcac?ctagttgact?tctgaactgc?ccttgggctc 180

cctctgctgg?catagtggtg?cgtgagcact?tttctggggt?catggaatgg?atttctcttt 240

ctgcccattt?tctttctttt?ggtttgtggt?tttctctctc?ataaattatt?acacaaacat 300

ggaccataca?atacaacctt?ctttcaaacc?tgatttgttt?gctaactgta?tcagtacctt 360

tccatataaa?attcataaat?atttttatat?aattatgtca?gtgtttccat?atttttctat 420

tacctgaatg?ttctttttag?catttaaata?atccattttt?agtaggcatt?tgtattgctt 480

ttttccattt?caggtagcat?ctaaagtgaa?tgtcccttgc?atgtatacta?ttcatatgtg 540

acttaataag?tagagcttgc?taaatgggta?ggaacttaca?ggaaaagggg?aaactctagt 600

ttgagaaaat?tgtctgggct?aagtttagga?tgtgttcaac?aaacaggaaa?gtcaagtttg 660

gctaaagcat?ggttgagtaa?tagtgggaaa?tatggtcaga?aagtaggtat?gttaggactc 720

tttctactga?aatgatgaaa?ttcaactcca?gccacaagtg?gagcagaaca?aaattgtgtt 780

tgtttaataa?ctgggaagcc?cagggtagct?ggaggcaggg?gcttagatgc?tgctgcgcca 840

tcccttacct?gtctgtttct?gtttctcctt?ctgtcccttc?ccagtctcag?cactgagtct 900

cttgcccatt?ggcctggtga?gggaaggagc?tgccagcccc?acccaacagc?tcaggttaca 960

gagagagtca?ctttcttcca?ttactcacag?agtaaacatc?aaggaaggcc?actgattgat 1020

tgacagtgtc?tgggtcagat?gtctatcctt?aggccagtcc?ctgtgaacaa?ggggatgggg 1080

tgtctgcgtg?gaccagatct?gaagcacagg?aagaagccag?acaatccatt?ctttcagata 1140

atgataaaaa?agaaaccatt?tattttattt?ctaagtatag?aatgaaacat?ttatagttgc 1200

ccaaattttg?gtacctttta?ggagaaaaat?acagattttt?ttgttgttaa?aaataaactt 1260

aaaaaaaaaa?aaaaaagact?taccatagtc?cgggcacggt?ggctcacacc?tgtaatccga 1320

gaactttggg?aggccgaggc?aggcggatca?cctgaggtca?ggagtttgag?accagcctgg 1380

ccaacatggc?gaaaccccat?ctctactaaa?aatacaaaaa?ttagccgggc?atagtgggtg 1440

gtgggtgcct?gtaatcccag?ctacttggga?ggctgaggca?ggagaatcac?ttgaacccag 1500

aaggtggagg?ttgcagtgag?ctgagattgt?gccattgtac?tccagcctgg?tcacaagagc 1560

gaagctctgt?atcgaaaaca?aaacaaaaaa?agacctactg?taaatagagt?acaacatgag 1620

actaacaaaa?ataacaaata?aaatcatcca?gcatatattc?tgtatttaaa?aaaaaaaaat 1680

cacaagatga?atacagacca?tcttggtgca?tatgtatttt?atacactaga?cacatgctga 1740

tgtttttaat?gcttaaaata?tattaggttt?catacttgct?tccaatccaa?ggagagtata 1800

tctgagctct?ttttttttag?atttgaaaac?cgccttttag?attagattag?aacactgcct 1860

ttttttttgt?tatgcagtaa?cagatcattg?cccacctgaa?agcgagtcgc?ctttattttt 1920

acctccttag?tagaaccagc?aggagctgca?attgctgttg?cttgagccaa?ggccacagtc 1980

agcatagaaa?ggttctgcta?agagactgtg?ttaaaggaaa?taaatttggt?gacttagttt 2040

tgttttacaa?gctagctgtg?gcccagtcct?tctgcagggt?gggtacgcag?gtttattgca 2100

gctctgcaga?gccatggcct?tagacaaaca?aagaaggtgt?gggaataccc?gggagctttc 2160

cattcaaagt?agattcttct?cctttgagtg?catgtaactg?aagcaccagc?tttctaaaaa 2220

aaaaaaaaaa?aaaaaaaaga?gagagaaaag?aaaagtcagt?ctgaagtttt?tgtgacactc 2280

agccagactc?cacattagat?aaagtaatca?tggaaataca?gctttatttt?tataccaagt 2340

tgtgcttaga?aagctgattg?ctaaaccagg?aagcttggct?ttcagaattt?tactgtaatc 2400

cagggtggaa?aaaaagtatc?actcagatgg?catttgtggg?agattactga?ctggtaatcc 2460

ctccttgtcc?tagaagatta?agtctgccct?ggtgtagtca?tggtcagctg?atttttttat 2520

ttatttattt?taaataaatt?gtagtttctt?gttgtaagtc?agccagtgtc?tgatgtttgc 2580

ctgaactgtt?tgtacctctg?ggccatattg?cagaaccctg?cccttctttg?ttgactgagg 2640

aaagctcgct?ccctgcccag?gtttttcatt?gttgatcgaa?attaacacca?ggtggtgaat 2700

agagcccctc?ctaaggttgc?tcaggataaa?tcatttatta?aataggtctg?cttatcagga 2760

ggggcgtgaa?ggctcccaaa?aggaaatgct?ggcacctggg?cccagaagcc?agggcctcta 2820

actcctgggg?ttgatttctt?cagtgaagtt?gcaccctaca?aagggaatat?ggccaaagcg 2880

gcactcaact?gaaggctgat?atcaggcgat?tagacagcca?tgcattctgc?gtttgtctgg 2940

<210>3

<211>2940

<212>DNA

＜213〉artificial sequence

<220>

<223>

<400>3

cctgtgcttc?agatctggtc?cacgcagaca?ccccatcccc?ttgttcacag?ggactggcct 60

aaggatagac?atctgaccca?gacactgtca?atcaatcagt?ggccttcctt?gatgtttact 120

ctgtgagtaa?tggaagaaag?tgactctctc?tgtaacctga?gctgttgggt?ggggctggca 180

gctccttccc?tcaccaggcc?aatgggcaag?agactcagtg?ctgagactgg?gaagggacag 240

aaggagaaac?agaaacagac?aggtaaggga?tggcgcagca?gcatctaagc?ccctgcctcc 300

agctaccctg?ggcttcccag?ttattaaaca?aacacaattt?tgttctgctc?cacttgtggc 360

tggagttgaa?tttcatcatt?tcagtagaaa?gagtcctaac?atacctactt?tctgaccata 420

tttcccacta?ttactcaacc?atgctttagc?caaacttgac?tttcctgttt?gttgaacaca 480

tcctaaactt?agcccagaca?attttctcaa?actagagttt?ccccttttcc?tgtaagttcc 540

tacccattta?gcaagctcta?cttattAAgt?cacatatgaa?tagtatacat?gcaagggaca 600

ttcactttag?atgctacctg?aaatggaaaa?aagcaataca?aatgcctact?aaaaatggat 660

tatttaaatg?ctaaaaagaa?cattcaggta?atagaaaaat?atggaaacac?tgacataatt 720

atataaaaat?atttatgaat?tttatatgga?aaggtactga?tacagttagc?aaacaaatca 780

ggtttgaaag?aaggttgtat?tgtatggtcc?atgtttgtgt?aataatttat?gagagagaaa 840

accacaaacc?aaaagaaaga?aaatgggcag?aaagagaaat?ccattccatg?accccagaaa 900

agtgctcacg?caccactatg?ccagcagagg?gagcccaagg?gcagttcaga?agtcaactag 960

gtgcagccag?taaagtgcgt?atgtactgca?gggtaataac?cggagacgac?tcatgtcatt 1020

gcaaacggaa?atggtagtag?gtacaaatga?ctataaagaa?aattaagttt?ctgtctgcct 1080

gttaactcat?cactcaaata?tgtcagtgta?aagaagccag?acaatccatt?ctttcagata 1140

atgataaaaa?agaaaccatt?tattttattt?ctaagtatag?aatgaaacat?ttatagttgc 1200

ccaaattttg?gtacctttta?ggagaaaaat?acagattttt?ttgttgttaa?aaataaactt 1260

aaaaaaaaaa?aaaaaagact?taccatagtc?cgggcacggt?ggctcacacc?tgtaatccga 1320

gaactttggg?aggccgaggc?aggcggatca?cctgaggtca?ggagtttgag?accagcctgg 1380

ccaacatggc?gaaaccccat?ctctactaaa?aatacaaaaa?ttagccgggc?atagtgggtg 1440

gtgggtgcct?gtaatcccag?ctacttggga?ggctgaggca?ggagaatcac?ttgaacccag 1500

aaggtggagg?ttgcagtgag?ctgagattgt?gccattgtac?tccagcctgg?tcacaagagc 1560

gaagctctgt?atcgaaaaca?aaacaaaaaa?agacctactg?taaatagagt?acaacatgag 1620

actaacaaaa?ataacaaata?aaatcatcca?gcatatattc?tgtatttaaa?aaaaaaaaat 1680

cacaagatga?atacagacca?tcttggtgca?tatgtatttt?atacactaga?cacatgctga 1740

tgtttttaat?gcttaaaata?tattaggttt?catacttgct?tccaatccaa?ggagagtata 1800

tctgagctct?ttttttttag?atttgaaaac?cgccttttag?attagattag?aacactgcct 1860

ttttttttgt?tatgcagtaa?cagatcattg?cccacctgaa?agcgagtcgc?ctttattttt 1920

acctccttag?tagaaccagc?aggagctgca?attgctgttg?cttgagccaa?ggccacagtc 1980

agcatagaaa?ggttctgcta?agagactgtg?ttaaaggaaa?taaatttggt?gacttagttt 2040

tgttttacaa?gctagctgtg?gcccagtcct?tctgcagggt?gggtacgcag?gtttattgca 2100

gctctgcaga?gccatggcct?tagacaaaca?aagaaggtgt?gggaataccc?gggagctttc 2160

cattcaaagt?agattcttct?cctttgagtg?catgtaactg?aagcaccagc?tttctaaaaa 2220

aaaaaaaaaa?aaaaaaaaga?gagagaaaag?aaaagtcagt?ctgaagtttt?tgtgacactc 2280

agccagactc?cacattagat?aaagtaatca?tggaaataca?gctttatttt?tataccaagt 2340

tgtgcttaga?aagctgattg?ctaaaccagg?aagcttggct?ttcagaattt?tactgtaatc 2400

cagggtggaa?aaaaagtatc?actcagatgg?catttgtggg?agattactga?ctggtaatcc 2460

ctccttgtcc?tagaagatta?agtctgccct?ggtgtagtca?tggtcagctg?atttttttat 2520

ttatttattt?taaataaatt?gtagtttctt?gttgtaagtc?agccagtgtc?tgatgtttgc 2580

ctgaactgtt?tgtacctctg?ggccatattg?cagaaccctg?cccttctttg?ttgactgagg 2640

aaagctcgct?ccctgcccag?gtttttcatt?gttgatcgaa?attaacacca?ggtggtgaat 2700

agagcccctc?ctaaggttgc?tcaggataaa?tcatttatta?aataggtctg?cttatcagga 2760

ggggcgtgaa?ggctcccaaa?aggaaatgct?ggcacctggg?cccagaagcc?agggcctcta 2820

actcctgggg?ttgatttctt?cagtgaagtt?gcaccctaca?aagggaatat?ggccaaagcg 2880

gcactcaact?gaaggctgat?atcaggcgat?tagacagcca?tgcattctgc?gtttgtctgg 2940

Claims (10)

1, a kind of PC-1 gene promoter is one of following nucleotide sequences:

1) SEQ ID № in the sequence table: 1 dna sequence dna;

2) with sequence table in SEQ ID №: 1 dna sequence dna that limits has 70% above homology, and has the dna sequence dna of identical function.

2, promotor according to claim 1 is characterized in that: described PC-1 gene promoter is SEQ ID № in the sequence table: 1.

3, promotor according to claim 2 is characterized in that: the enhancement sequences of transcribing that 1-1110 bit base of 5 ends of sequence 1 is the PC-1 gene promoter in the described sequence table.

4, promotor according to claim 1 is characterized in that: described PC-1 gene promoter is SEQ ID № in the sequence table: 2.

5, promotor according to claim 1 is characterized in that: described PC-1 gene promoter is SEQ ID № in the sequence table: 3.

6, the expression cassette that contains the described PC-1 gene promoter of claim 1.

7, the expression vector and the clone that contain the described PC-1 gene promoter of claim 1.

8, the application of the described PC-1 gene promoter of claim 1 in preparation treatment prostate cancer medicine.

9, application according to claim 8 is characterized in that: the application of described PC-1 gene promoter in the prostate cancer medicine of the preparation treatment androgen receptor positive and estrogen receptor positive.

10, the application of the described PC-1 gene promoter of claim 1 in preparation treatment breast cancer medicines.

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