出品公司: | Clontech |
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载体名称: | pAcGFP1-N3 |
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: | 4722 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-N3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: | 632484 |
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-N3载体质粒基本信息
pAcGFP1-N3质粒图谱载体图谱和pAcGFP1-N3载体序列质粒序列多克隆位点信息
pAcGFP1-N3质粒载体简介
pAcGFP1-N3载体描述
pAcGFP1-N3 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-N3 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-N3 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-N3 such that it is in frame with the AcGFP1 coding sequences, with no intervening inframe 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-N3 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-N3质粒序列载体序列
LOCUS pAcGFP1-N3 4722 bp DNA SYN DEFINITION pAcGFP1-N3 ACCESSION KEYWORDS SOURCE ORGANISM other sequences; artificial sequences; vectors. FEATURES Location/Qualifiers source 1..4722 /organism="pAcGFP1-N3" /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 669..1388 /label="ORF frame 3" /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH MIYFGFVTAAAITHGMDELYK*" gene 672..1385 /label="AcGFP1" /gene="AcGFP1" /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH MIYFGFVTAAAITHGMDELYK*" misc_feature 1601..1620 /label="EBV_rev_primer" /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH MIYFGFVTAAAITHGMDELYK*" rep_origin complement(1770..2076) /label="f1_origin" /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH MIYFGFVTAAAITHGMDELYK*" promoter 2155..2183 /label="AmpR_promoter" /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH MIYFGFVTAAAITHGMDELYK*" misc_feature complement(2249..2269) /label="pBABE_3_primer" /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH MIYFGFVTAAAITHGMDELYK*" misc_feature complement(2255..2470) /label="SV40_enhancer" /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH MIYFGFVTAAAITHGMDELYK*" promoter 2267..2535 /label="SV40_promoter" /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH MIYFGFVTAAAITHGMDELYK*" rep_origin 2434..2511 /label="SV40_origin" /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH MIYFGFVTAAAITHGMDELYK*" misc_feature 2496..2515 /label="SV40pro_F_primer" /translation="MVSKGAELFTGIVPILIELNGDVNGHKFSVSGEGEGDATYGKLT LKFICTTGKLPVPWPTLVTTLSYGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFE DDGNYKSRAEVKFEGDTLVNRIELTGTDFKEDGNILGNKMEYNYNAHNVYIMTDKAKN GIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDH MIYFGFVTAAAITHGMDELYK*" CDS 2618..3412 /label="ORF frame 2" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGR PVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE EHQGLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRY QDIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*" gene 2621..3409 /label="NeoR/KanR" /gene="NeoR/KanR" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGR PVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE EHQGLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRY QDIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF*" CDS complement(2927..3463) /label="ORF frame 3" /translation="MAGWASLGRSFRTPESRSEELVKKAIEGDALRIGSGDTVKHEEA VSPFAAKLFSNITGSQRYVLIAVRHTQPATVDESRKAAIFHHDIRQAGIAMGHDEILA VGHARLEPGEQFGWREPLMLFVQIILIDKTGFHPSTCSLDAMFRLVVEWAGSRIKRMQ PPHCISHDGYFLGRSKVR*" terminator 3587..3856 /label="TK_PA_terminator" /translation="MAGWASLGRSFRTPESRSEELVKKAIEGDALRIGSGDTVKHEEA VSPFAAKLFSNITGSQRYVLIAVRHTQPATVDESRKAAIFHHDIRQAGIAMGHDEILA VGHARLEPGEQFGWREPLMLFVQIILIDKTGFHPSTCSLDAMFRLVVEWAGSRIKRMQ PPHCISHDGYFLGRSKVR*" rep_origin 4004..4623 /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 GATCCATCAT GGTGAGCAAG GGCGCCGAGC TGTTCACCGG CATCGTGCCC ATCCTGATCG 721 AGCTGAATGG CGATGTGAAT GGCCACAAGT TCAGCGTGAG CGGCGAGGGC GAGGGCGATG 781 CCACCTACGG CAAGCTGACC CTGAAGTTCA TCTGCACCAC CGGCAAGCTG CCTGTGCCCT 841 GGCCCACCCT GGTGACCACC CTGAGCTACG GCGTGCAGTG CTTCTCACGC TACCCCGATC 901 ACATGAAGCA GCACGACTTC TTCAAGAGCG CCATGCCTGA GGGCTACATC CAGGAGCGCA 961 CCATCTTCTT CGAGGATGAC GGCAACTACA AGTCGCGCGC CGAGGTGAAG TTCGAGGGCG 1021 ATACCCTGGT GAATCGCATC GAGCTGACCG GCACCGATTT CAAGGAGGAT GGCAACATCC 1081 TGGGCAATAA GATGGAGTAC AACTACAACG CCCACAATGT GTACATCATG ACCGACAAGG 1141 CCAAGAATGG CATCAAGGTG AACTTCAAGA TCCGCCACAA CATCGAGGAT GGCAGCGTGC 1201 AGCTGGCCGA CCACTACCAG CAGAATACCC CCATCGGCGA TGGCCCTGTG CTGCTGCCCG 1261 ATAACCACTA CCTGTCCACC CAGAGCGCCC TGTCCAAGGA CCCCAACGAG AAGCGCGATC 1321 ACATGATCTA CTTCGGCTTC GTGACCGCCG CCGCCATCAC CCACGGCATG GATGAGCTGT 1381 ACAAGTGAGC GGCCGCGACT CTAGATCATA ATCAGCCATA CCACATTTGT AGAGGTTTTA 1441 CTTGCTTTAA AAAACCTCCC ACACCTCCCC CTGAACCTGA AACATAAAAT GAATGCAATT 1501 GTTGTTGTTA ACTTGTTTAT TGCAGCTTAT AATGGTTACA AATAAAGCAA TAGCATCACA 1561 AATTTCACAA ATAAAGCATT TTTTTCACTG CATTCTAGTT GTGGTTTGTC CAAACTCATC 1621 AATGTATCTT AAGGCGTAAA TTGTAAGCGT TAATATTTTG TTAAAATTCG CGTTAAATTT 1681 TTGTTAAATC AGCTCATTTT TTAACCAATA GGCCGAAATC GGCAAAATCC CTTATAAATC 1741 AAAAGAATAG ACCGAGATAG GGTTGAGTGT TGTTCCAGTT TGGAACAAGA GTCCACTATT 1801 AAAGAACGTG GACTCCAACG TCAAAGGGCG AAAAACCGTC TATCAGGGCG ATGGCCCACT 1861 ACGTGAACCA TCACCCTAAT CAAGTTTTTT GGGGTCGAGG TGCCGTAAAG CACTAAATCG 1921 GAACCCTAAA GGGAGCCCCC GATTTAGAGC TTGACGGGGA AAGCCGGCGA ACGTGGCGAG 1981 AAAGGAAGGG AAGAAAGCGA AAGGAGCGGG CGCTAGGGCG CTGGCAAGTG TAGCGGTCAC 2041 GCTGCGCGTA ACCACCACAC CCGCCGCGCT TAATGCGCCG CTACAGGGCG CGTCAGGTGG 2101 CACTTTTCGG GGAAATGTGC GCGGAACCCC TATTTGTTTA TTTTTCTAAA TACATTCAAA 2161 TATGTATCCG CTCATGAGAC AATAACCCTG ATAAATGCTT CAATAATATT GAAAAAGGAA 2221 GAGTCCTGAG GCGGAAAGAA CCAGCTGTGG AATGTGTGTC AGTTAGGGTG TGGAAAGTCC 2281 CCAGGCTCCC CAGCAGGCAG AAGTATGCAA AGCATGCATC TCAATTAGTC AGCAACCAGG 2341 TGTGGAAAGT CCCCAGGCTC CCCAGCAGGC AGAAGTATGC AAAGCATGCA TCTCAATTAG 2401 TCAGCAACCA TAGTCCCGCC CCTAACTCCG CCCATCCCGC CCCTAACTCC GCCCAGTTCC 2461 GCCCATTCTC CGCCCCATGG CTGACTAATT TTTTTTATTT ATGCAGAGGC CGAGGCCGCC 2521 TCGGCCTCTG AGCTATTCCA GAAGTAGTGA GGAGGCTTTT TTGGAGGCCT AGGCTTTTGC 2581 AAAGATCGAT CAAGAGACAG GATGAGGATC GTTTCGCATG ATTGAACAAG ATGGATTGCA 2641 CGCAGGTTCT CCGGCCGCTT GGGTGGAGAG GCTATTCGGC TATGACTGGG CACAACAGAC 2701 AATCGGCTGC TCTGATGCCG CCGTGTTCCG GCTGTCAGCG CAGGGGCGCC CGGTTCTTTT 2761 TGTCAAGACC GACCTGTCCG GTGCCCTGAA TGAACTGCAA GACGAGGCAG CGCGGCTATC 2821 GTGGCTGGCC ACGACGGGCG TTCCTTGCGC AGCTGTGCTC GACGTTGTCA CTGAAGCGGG 2881 AAGGGACTGG CTGCTATTGG GCGAAGTGCC GGGGCAGGAT CTCCTGTCAT CTCACCTTGC 2941 TCCTGCCGAG AAAGTATCCA TCATGGCTGA TGCAATGCGG CGGCTGCATA CGCTTGATCC 3001 GGCTACCTGC CCATTCGACC ACCAAGCGAA ACATCGCATC GAGCGAGCAC GTACTCGGAT 3061 GGAAGCCGGT CTTGTCGATC AGGATGATCT GGACGAAGAG CATCAGGGGC TCGCGCCAGC 3121 CGAACTGTTC GCCAGGCTCA AGGCGAGCAT GCCCGACGGC GAGGATCTCG TCGTGACCCA 3181 TGGCGATGCC TGCTTGCCGA ATATCATGGT GGAAAATGGC CGCTTTTCTG GATTCATCGA 3241 CTGTGGCCGG CTGGGTGTGG CGGACCGCTA TCAGGACATA GCGTTGGCTA CCCGTGATAT 3301 TGCTGAAGAG CTTGGCGGCG AATGGGCTGA CCGCTTCCTC GTGCTTTACG GTATCGCCGC 3361 TCCCGATTCG CAGCGCATCG CCTTCTATCG CCTTCTTGAC GAGTTCTTCT GAGCGGGACT 3421 CTGGGGTTCG AAATGACCGA CCAAGCGACG CCCAACCTGC CATCACGAGA TTTCGATTCC 3481 ACCGCCGCCT TCTATGAAAG GTTGGGCTTC GGAATCGTTT TCCGGGACGC CGGCTGGATG 3541 ATCCTCCAGC GCGGGGATCT CATGCTGGAG TTCTTCGCCC ACCCTAGGGG GAGGCTAACT 3601 GAAACACGGA AGGAGACAAT ACCGGAAGGA ACCCGCGCTA TGACGGCAAT AAAAAGACAG 3661 AATAAAACGC ACGGTGTTGG GTCGTTTGTT CATAAACGCG GGGTTCGGTC CCAGGGCTGG 3721 CACTCTGTCG ATACCCCACC GAGACCCCAT TGGGGCCAAT ACGCCCGCGT TTCTTCCTTT 3781 TCCCCACCCC ACCCCCCAAG TTCGGGTGAA GGCCCAGGGC TCGCAGCCAA CGTCGGGGCG 3841 GCAGGCCCTG CCATAGCCTC AGGTTACTCA TATATACTTT AGATTGATTT AAAACTTCAT 3901 TTTTAATTTA AAAGGATCTA GGTGAAGATC CTTTTTGATA ATCTCATGAC CAAAATCCCT 3961 TAACGTGAGT TTTCGTTCCA CTGAGCGTCA GACCCCGTAG AAAAGATCAA AGGATCTTCT 4021 TGAGATCCTT TTTTTCTGCG CGTAATCTGC TGCTTGCAAA CAAAAAAACC ACCGCTACCA 4081 GCGGTGGTTT GTTTGCCGGA TCAAGAGCTA CCAACTCTTT TTCCGAAGGT AACTGGCTTC 4141 AGCAGAGCGC AGATACCAAA TACTGTTCTT CTAGTGTAGC CGTAGTTAGG CCACCACTTC 4201 AAGAACTCTG TAGCACCGCC TACATACCTC GCTCTGCTAA TCCTGTTACC AGTGGCTGCT 4261 GCCAGTGGCG ATAAGTCGTG TCTTACCGGG TTGGACTCAA GACGATAGTT ACCGGATAAG 4321 GCGCAGCGGT CGGGCTGAAC GGGGGGTTCG TGCACACAGC CCAGCTTGGA GCGAACGACC 4381 TACACCGAAC TGAGATACCT ACAGCGTGAG CTATGAGAAA GCGCCACGCT TCCCGAAGGG 4441 AGAAAGGCGG ACAGGTATCC GGTAAGCGGC AGGGTCGGAA CAGGAGAGCG CACGAGGGAG 4501 CTTCCAGGGG GAAACGCCTG GTATCTTTAT AGTCCTGTCG GGTTTCGCCA CCTCTGACTT 4561 GAGCGTCGAT TTTTGTGATG CTCGTCAGGG GGGCGGAGCC TATGGAAAAA CGCCAGCAAC 4621 GCGGCCTTTT TACGGTTCCT GGCCTTTTGC TGGCCTTTTG CTCACATGTT CTTTCCTGCG 4681 TTATCCCCTG ATTCTGTGGA TAACCGTATT ACCGCCATGC AT //
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