LOCUS       sequence-224759-        6894 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6894)
  AUTHORS   Beals CR, Clipstone NA, Ho SN, Crabtree GR
  TITLE     Nuclear localization of NF-ATc by a calcineurin-dependent, 
            cyclosporin-sensitive intramolecular interaction.
  JOURNAL   Genes Dev. 1997 Apr 1. 11(7):824-34.
  PUBMED    9106655
REFERENCE   2  (bases 1 to 6894)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Genes Dev. 
            1997 Apr 1. 11(7):824-34."
COMMENT     Sequence Label: EGFPC1-huNFATc1EE-WT
FEATURES             Location/Qualifiers
     source          1..6894
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3499..3516
                     /codon_start=1
                     /product="epitope tag from middle T antigen of mouse 
                     polyoma virus"
                     /label=Glu-Glu tag
                     /translation="EYMPME"
     polyA_signal    3682..3803
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3719..3738)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3773..3792
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3810..4265)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3947..3968)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4159..4178
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4292..4396
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4398..4755
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4398..4589
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4421..4441)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4606..4741
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4790..5584
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4790..5584
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4844..4863)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5454..5473
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5772..5791)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5816..5863
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6192..6780
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6681..6700
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
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 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atccatgtca agcaccagct ttccagtccc ttccaagttt
     1381 ccacttggcc ctgcggctgc ggtcttcggg agaggagaaa ctttggggcc cgcgccgcgc
     1441 gccggcggca ccatgaagtc agcggaggaa gaacactatg gctatgcatc ctccaacgtc
     1501 agccccgccc tgccgctccc cacggcgcac tccaccctgc cggccccgtg ccacaacctt
     1561 cagacctcca caccgggcat catcccgccg gcggatcacc cctcggggta cggagcagct
     1621 ttggacggtg ggcccgcggg ctacttcctc tcctccggcc acaccaggcc tgatggggcc
     1681 cctgccctgg agagtcctcg catcgagata acctcgtgct tgggcctgta ccacaacaat
     1741 aaccagtttt tccacgatgt ggaggtggaa gacgtcctcc ctagctccaa acggtccccc
     1801 tccacggcca cgctgagtct gcccagcctg gaggcctaca gagacccctc gtgcctgagc
     1861 ccggccagca gcctgtcctc ccggagctgc aactcagagg cctcctccta cgagtccaac
     1921 tactcgtacc cgtacgcgtc cccccagacg tcgccatggc agtctccctg cgtgtctccc
     1981 aagaccacgg accccgagga gggctttccc cgcgggctgg gggcctgcac actgctgggt
     2041 tccccgcagc actccccctc cacctcgccc cgcgccagcg tcactgagga gagctggctg
     2101 ggtgcccgct cctccagacc cgcgtcccct tgcaacaaga ggaagtacag cctcaacggc
     2161 cggcagccgc cctactcacc ccaccactcg cccacgccgt ccccgcacgg ctccccgcgg
     2221 gtcagcgtga ccgacgactc gtggttgggc aacaccaccc agtacaccag ctcggccatc
     2281 gtggccgcca tcaacgcgct gaccaccgac agcagcctgg acctgggaga tggcgtccct
     2341 gtcaagtccc gcaagaccac cctggagcag ccgccctcag tggcgctcaa ggtggagccc
     2401 gtcggggagg acctgggcag ccccccgccc ccggccgact tcgcgcccga agactactcc
     2461 tctttccagc acatcaggaa gggcggcttc tgcgaccagt acctggcggt gccgcagcac
     2521 ccctaccagt gggcgaagcc caagcccctg tcccctacgt cctacatgag cccgaccctg
     2581 cccgccctgg actggcagct gccgtcccac tcaggcccgt atgagcttcg gattgaggtg
     2641 cagcccaagt cccaccaccg agcccactac gagacggagg gcagccgggg ggccgtgaag
     2701 gcgtcggccg gaggacaccc catcgtgcag ctgcatggct acttggagaa tgagccgctg
     2761 atgctgcagc ttttcattgg gacggcggac gaccgcctgc tgcgcccgca cgccttctac
     2821 caggtgcacc gcatcacagg gaagaccgtg tccaccacca gccacgaggc tatcctctcc
     2881 aacaccaaag tcctggagat cccactcctg ctggagaaca gcatgcgagc cgtcattgac
     2941 tgtgccggaa tcctgaaact cagaaactcc gacattgaac ttcggaaagg agagacggac
     3001 atcgggagga agaacacacg ggtacggctg gtgttccgcg ttcacgtccc gcaacccagc
     3061 ggccgcacgc tgtccctgca ggtggcctcc aaccccatcg aatgctccca gcgctcagct
     3121 caggagctgc ctctggtgga gaagcagagc acggacagct atccggtcgt gggcgggaag
     3181 aagatggtcc tgtctggcca caacttcctg caggactcca aggtcatttt cgtggagaaa
     3241 gccccagatg gccaccatgt ctgggagatg gaagcgaaaa ctgaccggga cctgtgcaag
     3301 ccgaattctc tggtggttga gatcccgcca tttcggaatc agaggataac cagccccgtt
     3361 cacgtcagtt tctacgtctg caacgggaag agaaagcgaa gccagtacca gcgtttcacc
     3421 taccttcccg ccaacggtaa cgccatcttt ctaaccgtaa gccgtgaaca tgagcgcgtg
     3481 gggtgctttt tcggcagcga atatatgcca atggaatgag cggccgcaaa aggaaaaaag
     3541 ggcgaattcg tttgatccac cggatctaga taactgatca taatcagcca taccacattt
     3601 gtagaggttt tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa
     3661 atgaatgcaa ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc
     3721 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     3781 tccaaactca tcaatgtatc ttaacgcgta aattgtaagc gttaatattt tgttaaaatt
     3841 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     3901 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     3961 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     4021 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     4081 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     4141 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     4201 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     4261 cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     4321 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     4381 ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg
     4441 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     4501 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     4561 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     4621 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     4681 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     4741 ctaggctttt gcaaagatcg atcaagagac aggatgagga tcgtttcgca tgattgaaca
     4801 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     4861 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     4921 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aagacgaggc
     4981 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     5041 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     5101 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     5161 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     5221 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     5281 gctcgcgcca gccgaactgt tcgccaggct caaggcgagc atgcccgacg gcgaggatct
     5341 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     5401 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     5461 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     5521 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     5581 ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga
     5641 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     5701 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccctagg
     5761 gggaggctaa ctgaaacacg gaaggagaca ataccggaag gaacccgcgc tatgacggca
     5821 ataaaaagac agaataaaac gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg
     5881 tcccagggct ggcactctgt cgatacccca ccgagacccc attggggcca atacgcccgc
     5941 gtttcttcct tttccccacc ccacccccca agttcgggtg aaggcccagg gctcgcagcc
     6001 aacgtcgggg cggcaggccc tgccatagcc tcaggttact catatatact ttagattgat
     6061 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     6121 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     6181 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     6241 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     6301 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     6361 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     6421 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     6481 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     6541 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     6601 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     6661 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     6721 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     6781 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     6841 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgccat gcat
//
