| 出品公司: | Clontech |
|---|---|
| 载体名称: | pAcGFP1-N2 |
| 质粒类型: | 哺乳动物细胞表达载体；荧光报告载体 |
| 高拷贝/低拷贝: | 高拷贝 |
| 克隆方法: | 限制性内切酶，多克隆位点 |
| 启动子: | CMV IE |
| 载体大小: | 4730 bp |
| 5' 测序引物及序列: | -- |
| 3' 测序引物及序列: | -- |
| 载体标签: | AcGFP1 (C-端） |
| 载体抗性: | 卡那霉素 |
| 筛选标记: | 新霉素（Neomycin） |
| 克隆菌株: | DH5α, HB101 |
| 宿主细胞（系）: | 常规细胞系（293、CV-1、CHO等） |
| 备注: |
哺乳动物载体pAcGFP1-N2组成型表达C端AcGFP融合蛋白； AcGFP1与GFP相比，密码子经过了优化，更适合在哺乳动物细胞中高水平表达，同时也增强了亮度。 |
| 产品目录号: | 632483 |
| 稳定性: | 稳表达 或 瞬表达 |
| 组成型/诱导型: | 组成型 |
| 病毒/非病毒: | 非病毒 |
买家导航
pAcGFP1-N2载体质粒基本信息
pAcGFP1-N2质粒图谱载体图谱和pAcGFP1-N2载体序列质粒序列多克隆位点信息
pAcGFP1-N2质粒载体简介
pAcGFP1-N2载体描述
pAcGFP1-N2 encodes a Green Fluorescent Protein (GFP) from Aequorea coerulescens. (Excitation maximum = 475 nm; emission maximum = 505 nm.) The coding sequence of the AcGFP1 gene contains silent base changes, which correspond to human codon-usage preferences (1). The MCS in pAcGFP1-N2 is between the immediate early promoter of CMV (PCMV IE) and the AcGFP1 coding sequences. Genes cloned into the MCS will be expressed as fusions to the N-terminus of AcGFP1 if they are in the same reading frame as AcGFP1 and there are no intervening stop codons. SV40 polyadenylation signals downstream of the AcGFP1 gene direct proper processing of the 3' end of the AcGFP1 mRNA. The vector backbone also contains an SV40 origin for replication in mammalian cells expressing the SV40 T antigen. A neomycin-resistance cassette (Neor)—consisting of the SV40 early promoter, the neomycin/ kanamycin resistance gene of Tn5, and polyadenylation signals from the Herpes simplex virus thymidine kinase (HSV TK) gene—allows stably transfected eukaryotic cells to be selected using G418. A bacterial promoter upstream of the gene expresses kanamycin resistance in E. coli. The pAcGFP1-N2 backbone also provides a pUC origin of replication for propagation in E. coli and an f1 origin for single-stranded DNA production.
Fusions to the N-terminus of AcGFP1 retain the fluorescent properties of the native protein allowing the localization of the fusion protein in vivo . The target gene should be cloned into pAcGFP1-N2 such that it is in frame with the AcGFP1 coding sequences and contains no intervening in-frame stop codons. The inserted gene should include the initiating ATG codon. The recombinant AcGFP1 vector can be transfected into mammalian cells using any standard transfection method. If required, stable transformants can be selected using G418 (2). pAcGFP1-N2 can also be used simply to express AcGFP1 in a cell line of interest (e.g., as a transfection marker).
Propagation in E. coli:
Suitable host strains: DH5α, HB101, and other general purpose strains. Single-stranded DNA production requires
a host containing an F plasmid such as JM109 or XL1-Blue.
Selectable marker: plasmid confers resistance to kanamycin (50 μg/ml) in E. coli hosts.
E. coli replication origin: pUC
Copy number: ≈500
Plasmid incompatibility group: pMB1/ColE1
pAcGFP1-N2质粒序列载体序列
LOCUS pAcGFP1-N2 4730 bp DNA SYN
DEFINITION pAcGFP1-N2
ACCESSION
KEYWORDS
SOURCE
ORGANISM other sequences; artificial sequences; vectors.
FEATURES Location/Qualifiers
source 1..4730
/organism="pAcGFP1-N2"
/mol_type="other DNA"
promoter 10..562
/label="CMV_immearly_promoter"
misc_feature 65..352
/label="CAG_enhancer"
misc_feature 519..539
/label="CMV_fwd_primer"
promoter 520..589
/label="CMV_promoter"
CDS 677..1396
/label="ORF frame 2"
/translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MIYFGFVTAAAITHGMDELYK*"
gene 680..1393
/label="AcGFP1"
/gene="AcGFP1"
/translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MIYFGFVTAAAITHGMDELYK*"
misc_feature 1609..1628
/label="EBV_rev_primer"
/translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MIYFGFVTAAAITHGMDELYK*"
rep_origin complement(1778..2084)
/label="f1_origin"
/translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MIYFGFVTAAAITHGMDELYK*"
promoter 2163..2191
/label="AmpR_promoter"
/translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MIYFGFVTAAAITHGMDELYK*"
misc_feature complement(2257..2277)
/label="pBABE_3_primer"
/translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MIYFGFVTAAAITHGMDELYK*"
misc_feature complement(2263..2478)
/label="SV40_enhancer"
/translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MIYFGFVTAAAITHGMDELYK*"
promoter 2275..2543
/label="SV40_promoter"
/translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MIYFGFVTAAAITHGMDELYK*"
rep_origin 2442..2519
/label="SV40_origin"
/translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MIYFGFVTAAAITHGMDELYK*"
misc_feature 2504..2523
/label="SV40pro_F_primer"
/translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT
LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE
DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN
GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH
MIYFGFVTAAAITHGMDELYK*"
CDS 2626..3420
/label="ORF frame 1"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGR
PVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
EHQGLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRY
QDIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*"
gene 2629..3417
/label="NeoR/KanR"
/gene="NeoR/KanR"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGR
PVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
EHQGLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRY
QDIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*"
CDS complement(2935..3471)
/label="ORF frame 3"
/translation="MAGWASLGRSFRTPESRSEELVKKAIEGDALRIGSGDTVKHEEA
VSPFAAKLFSNITGSQRYVLIAVRHTQPATVDESRKAAIFHHDIRQAGIAMGHDEILA
VGHARLEPGEQFGWREPLMLFVQIILIDKTGFHPSTCSLDAMFRLVVEWAGSRIKRMQ
PPHCISHDGYFLGRSKVR*"
terminator 3595..3864
/label="TK_PA_terminator"
/translation="MAGWASLGRSFRTPESRSEELVKKAIEGDALRIGSGDTVKHEEA
VSPFAAKLFSNITGSQRYVLIAVRHTQPATVDESRKAAIFHHDIRQAGIAMGHDEILA
VGHARLEPGEQFGWREPLMLFVQIILIDKTGFHPSTCSLDAMFRLVVEWAGSRIKRMQ
PPHCISHDGYFLGRSKVR*"
rep_origin 4012..4631
/label="pBR322_origin"
/translation="MAGWASLGRSFRTPESRSEELVKKAIEGDALRIGSGDTVKHEEA
VSPFAAKLFSNITGSQRYVLIAVRHTQPATVDESRKAAIFHHDIRQAGIAMGHDEILA
VGHARLEPGEQFGWREPLMLFVQIILIDKTGFHPSTCSLDAMFRLVVEWAGSRIKRMQ
PPHCISHDGYFLGRSKVR*"
ORIGIN
1 TAGTTATTAA TAGTAATCAA TTACGGGGTC ATTAGTTCAT AGCCCATATA TGGAGTTCCG
61 CGTTACATAA CTTACGGTAA ATGGCCCGCC TGGCTGACCG CCCAACGACC CCCGCCCATT
121 GACGTCAATA ATGACGTATG TTCCCATAGT AACGCCAATA GGGACTTTCC ATTGACGTCA
181 ATGGGTGGAG TATTTACGGT AAACTGCCCA CTTGGCAGTA CATCAAGTGT ATCATATGCC
241 AAGTACGCCC CCTATTGACG TCAATGACGG TAAATGGCCC GCCTGGCATT ATGCCCAGTA
301 CATGACCTTA TGGGACTTTC CTACTTGGCA GTACATCTAC GTATTAGTCA TCGCTATTAC
361 CATGGTGATG CGGTTTTGGC AGTACATCAA TGGGCGTGGA TAGCGGTTTG ACTCACGGGG
421 ATTTCCAAGT CTCCACCCCA TTGACGTCAA TGGGAGTTTG TTTTGGCACC AAAATCAACG
481 GGACTTTCCA AAATGTCGTA ACAACTCCGC CCCATTGACG CAAATGGGCG GTAGGCGTGT
541 ACGGTGGGAG GTCTATATAA GCAGAGCTGG TTTAGTGAAC CGTCAGATCC GCTAGCGCTA
601 CCGGACTCAG ATCTCGAGCT CAAGCTTCGA ATTCTGCAGT CGACGGTACC GCGGGCCCGG
661 GATCCACCGG CCGGTCATGG TGAGCAAGGG CGCCGAGCTG TTCACCGGCA TCGTGCCCAT
721 CCTGATCGAG CTGAATGGCG ATGTGAATGG CCACAAGTTC AGCGTGAGCG GCGAGGGCGA
781 GGGCGATGCC ACCTACGGCA AGCTGACCCT GAAGTTCATC TGCACCACCG GCAAGCTGCC
841 TGTGCCCTGG CCCACCCTGG TGACCACCCT GAGCTACGGC GTGCAGTGCT TCTCACGCTA
901 CCCCGATCAC ATGAAGCAGC ACGACTTCTT CAAGAGCGCC ATGCCTGAGG GCTACATCCA
961 GGAGCGCACC ATCTTCTTCG AGGATGACGG CAACTACAAG TCGCGCGCCG AGGTGAAGTT
1021 CGAGGGCGAT ACCCTGGTGA ATCGCATCGA GCTGACCGGC ACCGATTTCA AGGAGGATGG
1081 CAACATCCTG GGCAATAAGA TGGAGTACAA CTACAACGCC CACAATGTGT ACATCATGAC
1141 CGACAAGGCC AAGAATGGCA TCAAGGTGAA CTTCAAGATC CGCCACAACA TCGAGGATGG
1201 CAGCGTGCAG CTGGCCGACC ACTACCAGCA GAATACCCCC ATCGGCGATG GCCCTGTGCT
1261 GCTGCCCGAT AACCACTACC TGTCCACCCA GAGCGCCCTG TCCAAGGACC CCAACGAGAA
1321 GCGCGATCAC ATGATCTACT TCGGCTTCGT GACCGCCGCC GCCATCACCC ACGGCATGGA
1381 TGAGCTGTAC AAGTGAGCGG CCGCGACTCT AGATCATAAT CAGCCATACC ACATTTGTAG
1441 AGGTTTTACT TGCTTTAAAA AACCTCCCAC ACCTCCCCCT GAACCTGAAA CATAAAATGA
1501 ATGCAATTGT TGTTGTTAAC TTGTTTATTG CAGCTTATAA TGGTTACAAA TAAAGCAATA
1561 GCATCACAAA TTTCACAAAT AAAGCATTTT TTTCACTGCA TTCTAGTTGT GGTTTGTCCA
1621 AACTCATCAA TGTATCTTAA GGCGTAAATT GTAAGCGTTA ATATTTTGTT AAAATTCGCG
1681 TTAAATTTTT GTTAAATCAG CTCATTTTTT AACCAATAGG CCGAAATCGG CAAAATCCCT
1741 TATAAATCAA AAGAATAGAC CGAGATAGGG TTGAGTGTTG TTCCAGTTTG GAACAAGAGT
1801 CCACTATTAA AGAACGTGGA CTCCAACGTC AAAGGGCGAA AAACCGTCTA TCAGGGCGAT
1861 GGCCCACTAC GTGAACCATC ACCCTAATCA AGTTTTTTGG GGTCGAGGTG CCGTAAAGCA
1921 CTAAATCGGA ACCCTAAAGG GAGCCCCCGA TTTAGAGCTT GACGGGGAAA GCCGGCGAAC
1981 GTGGCGAGAA AGGAAGGGAA GAAAGCGAAA GGAGCGGGCG CTAGGGCGCT GGCAAGTGTA
2041 GCGGTCACGC TGCGCGTAAC CACCACACCC GCCGCGCTTA ATGCGCCGCT ACAGGGCGCG
2101 TCAGGTGGCA CTTTTCGGGG AAATGTGCGC GGAACCCCTA TTTGTTTATT TTTCTAAATA
2161 CATTCAAATA TGTATCCGCT CATGAGACAA TAACCCTGAT AAATGCTTCA ATAATATTGA
2221 AAAAGGAAGA GTCCTGAGGC GGAAAGAACC AGCTGTGGAA TGTGTGTCAG TTAGGGTGTG
2281 GAAAGTCCCC AGGCTCCCCA GCAGGCAGAA GTATGCAAAG CATGCATCTC AATTAGTCAG
2341 CAACCAGGTG TGGAAAGTCC CCAGGCTCCC CAGCAGGCAG AAGTATGCAA AGCATGCATC
2401 TCAATTAGTC AGCAACCATA GTCCCGCCCC TAACTCCGCC CATCCCGCCC CTAACTCCGC
2461 CCAGTTCCGC CCATTCTCCG CCCCATGGCT GACTAATTTT TTTTATTTAT GCAGAGGCCG
2521 AGGCCGCCTC GGCCTCTGAG CTATTCCAGA AGTAGTGAGG AGGCTTTTTT GGAGGCCTAG
2581 GCTTTTGCAA AGATCGATCA AGAGACAGGA TGAGGATCGT TTCGCATGAT TGAACAAGAT
2641 GGATTGCACG CAGGTTCTCC GGCCGCTTGG GTGGAGAGGC TATTCGGCTA TGACTGGGCA
2701 CAACAGACAA TCGGCTGCTC TGATGCCGCC GTGTTCCGGC TGTCAGCGCA GGGGCGCCCG
2761 GTTCTTTTTG TCAAGACCGA CCTGTCCGGT GCCCTGAATG AACTGCAAGA CGAGGCAGCG
2821 CGGCTATCGT GGCTGGCCAC GACGGGCGTT CCTTGCGCAG CTGTGCTCGA CGTTGTCACT
2881 GAAGCGGGAA GGGACTGGCT GCTATTGGGC GAAGTGCCGG GGCAGGATCT CCTGTCATCT
2941 CACCTTGCTC CTGCCGAGAA AGTATCCATC ATGGCTGATG CAATGCGGCG GCTGCATACG
3001 CTTGATCCGG CTACCTGCCC ATTCGACCAC CAAGCGAAAC ATCGCATCGA GCGAGCACGT
3061 ACTCGGATGG AAGCCGGTCT TGTCGATCAG GATGATCTGG ACGAAGAGCA TCAGGGGCTC
3121 GCGCCAGCCG AACTGTTCGC CAGGCTCAAG GCGAGCATGC CCGACGGCGA GGATCTCGTC
3181 GTGACCCATG GCGATGCCTG CTTGCCGAAT ATCATGGTGG AAAATGGCCG CTTTTCTGGA
3241 TTCATCGACT GTGGCCGGCT GGGTGTGGCG GACCGCTATC AGGACATAGC GTTGGCTACC
3301 CGTGATATTG CTGAAGAGCT TGGCGGCGAA TGGGCTGACC GCTTCCTCGT GCTTTACGGT
3361 ATCGCCGCTC CCGATTCGCA GCGCATCGCC TTCTATCGCC TTCTTGACGA GTTCTTCTGA
3421 GCGGGACTCT GGGGTTCGAA ATGACCGACC AAGCGACGCC CAACCTGCCA TCACGAGATT
3481 TCGATTCCAC CGCCGCCTTC TATGAAAGGT TGGGCTTCGG AATCGTTTTC CGGGACGCCG
3541 GCTGGATGAT CCTCCAGCGC GGGGATCTCA TGCTGGAGTT CTTCGCCCAC CCTAGGGGGA
3601 GGCTAACTGA AACACGGAAG GAGACAATAC CGGAAGGAAC CCGCGCTATG ACGGCAATAA
3661 AAAGACAGAA TAAAACGCAC GGTGTTGGGT CGTTTGTTCA TAAACGCGGG GTTCGGTCCC
3721 AGGGCTGGCA CTCTGTCGAT ACCCCACCGA GACCCCATTG GGGCCAATAC GCCCGCGTTT
3781 CTTCCTTTTC CCCACCCCAC CCCCCAAGTT CGGGTGAAGG CCCAGGGCTC GCAGCCAACG
3841 TCGGGGCGGC AGGCCCTGCC ATAGCCTCAG GTTACTCATA TATACTTTAG ATTGATTTAA
3901 AACTTCATTT TTAATTTAAA AGGATCTAGG TGAAGATCCT TTTTGATAAT CTCATGACCA
3961 AAATCCCTTA ACGTGAGTTT TCGTTCCACT GAGCGTCAGA CCCCGTAGAA AAGATCAAAG
4021 GATCTTCTTG AGATCCTTTT TTTCTGCGCG TAATCTGCTG CTTGCAAACA AAAAAACCAC
4081 CGCTACCAGC GGTGGTTTGT TTGCCGGATC AAGAGCTACC AACTCTTTTT CCGAAGGTAA
4141 CTGGCTTCAG CAGAGCGCAG ATACCAAATA CTGTTCTTCT AGTGTAGCCG TAGTTAGGCC
4201 ACCACTTCAA GAACTCTGTA GCACCGCCTA CATACCTCGC TCTGCTAATC CTGTTACCAG
4261 TGGCTGCTGC CAGTGGCGAT AAGTCGTGTC TTACCGGGTT GGACTCAAGA CGATAGTTAC
4321 CGGATAAGGC GCAGCGGTCG GGCTGAACGG GGGGTTCGTG CACACAGCCC AGCTTGGAGC
4381 GAACGACCTA CACCGAACTG AGATACCTAC AGCGTGAGCT ATGAGAAAGC GCCACGCTTC
4441 CCGAAGGGAG AAAGGCGGAC AGGTATCCGG TAAGCGGCAG GGTCGGAACA GGAGAGCGCA
4501 CGAGGGAGCT TCCAGGGGGA AACGCCTGGT ATCTTTATAG TCCTGTCGGG TTTCGCCACC
4561 TCTGACTTGA GCGTCGATTT TTGTGATGCT CGTCAGGGGG GCGGAGCCTA TGGAAAAACG
4621 CCAGCAACGC GGCCTTTTTA CGGTTCCTGG CCTTTTGCTG GCCTTTTGCT CACATGTTCT
4681 TTCCTGCGTT ATCCCCTGAT TCTGTGGATA ACCGTATTAC CGCCATGCAT
//
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