LOCUS       sequence-43253-e        4732 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 4732)
  AUTHORS   Rizzo MA, Springer GH, Granada B, Piston DW
  TITLE     An improved cyan fluorescent protein variant useful for FRET.
  JOURNAL   Nat Biotechnol. 2004 Apr . 22(4):445-9.
  PUBMED    14990965
REFERENCE   2  (bases 1 to 4732)
  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: "Nat 
            Biotechnol. 2004 Apr . 22(4):445-9."
COMMENT     Sequence Label: Cerulean
FEATURES             Location/Qualifiers
     source          1..4732
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(51..407)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      65..199
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        216..407
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     364..384
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(409..513)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      540..995
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(627..646)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     837..858
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    1002..1124
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1013..1032)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1068..1087
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     misc_feature    1248..1313
                     /label=MCS
                     /note="multiple cloning site"
     CDS             complement(join(1314..2024,2025..2027,2028..2030))
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1356..1377)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1705..1724
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1964..1985
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2027..2036
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(2075..2278)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2104..2124)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2279..2582
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(2757..3345)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2837..2856)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3675..3722
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3747..3766
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(join(3954..4732,1..16))
                     /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             complement(join(3954..4732,1..16))
                     /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(4065..4084)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4675..4694
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 catcttgttc aatcatgcga aacgatcctc atcctgtctc ttgatcgatc tttgcaaaag
       61 cctaggcctc caaaaaagcc tcctcactac ttctggaata gctcagaggc cgaggcggcc
      121 tcggcctctg cataaataaa aaaattagtc agccatgggg cggagaatgg gcggaactgg
      181 gcggagttag gggcgggatg ggcggagtta ggggcgggac tatggttgct gactaattga
      241 gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca cacctggttg
      301 ctgactaatt gagatgcatg ctttgcatac ttctgcctgc tggggagcct ggggactttc
      361 cacaccctaa ctgacacaca ttccacagct ggttctttcc gcctcaggac tcttcctttt
      421 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
      481 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
      541 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
      601 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
      661 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
      721 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
      781 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
      841 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
      901 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
      961 cgcgaatttt aacaaaatat taacgcttac aatttacgcc ttaagataca ttgatgagtt
     1021 tggacaaacc acaactagaa tgcagtgaaa aaaatagctt tatttgtgaa atttgtgatg
     1081 ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca
     1141 ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc
     1201 tctacaaatg tggtatggct gattatgatc agttatctag atccggtgga tcccgggccc
     1261 gcggtaccgt cgactgcaga attcgaagct tgagctcgag atctgagtcc ggacttgtac
     1321 agctcgtcca tgccgagagt gatcccggcg gcggtcacga actccagcag gaccatgtga
     1381 tcgcgcttct cgttggggtc tttgctcagc ttggactggg tgctcaggta gtggttgtcg
     1441 ggcagcagca cggggccgtc gccgatgggg gtgttctgct ggtagtggtc ggcgagctgc
     1501 acgctgccgt cctcgatgtt gtggcggatc ttgaagttgg ccttgatgcc gttcttctgc
     1561 ttgtcggcgg tgatatagac gttgtcgctg atggcgttgt actccagctt gtgccccagg
     1621 atgttgccgt cctccttgaa gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg
     1681 ccctcgaact tcacctcggc gcgggtcttg tagttgccgt cgtccttgaa gaagatggtg
     1741 cgctcctgga cgtagccttc gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg
     1801 tcggggtagc gggcgaagca ctgcacgccc caggtcaggg tggtcacgag ggtgggccag
     1861 ggcacgggca gcttgccggt ggtgcagatg aacttcaggg tcagcttgcc gtaggtggca
     1921 tcgccctcgc cctcgccgga cacgctgaac ttgtggccgt ttacgtcgcc gtccagctcg
     1981 accaggatgg gcaccacccc ggtgaacagc tcctcgccct tgctcaccat ggtggcgacc
     2041 ggtagcgcta gcggatctga cggttcacta aaccagctct gcttatatag acctcccacc
     2101 gtacacgcct accgcccatt tgcgtcaatg gggcggagtt gttacgacat tttggaaagt
     2161 cccgttgatt ttggtgccaa aacaaactcc cattgacgtc aatggggtgg agacttggaa
     2221 atccccgtga gtcaaaccgc tatccacgcc cattgatgta ctgccaaaac cgcatcacca
     2281 tggtaatagc gatgactaat acgtagatgt actgccaagt aggaaagtcc cataaggtca
     2341 tgtactgggc ataatgccag gcgggccatt taccgtcatt gacgtcaata gggggcgtac
     2401 ttggcatatg atacacttga tgtactgcca agtgggcagt ttaccgtaaa tactccaccc
     2461 attgacgtca atggaaagtc cctattggcg ttactatggg aacatacgtc attattgacg
     2521 tcaatgggcg ggggtcgttg ggcggtcagc caggcgggcc atttaccgta agttatgtaa
     2581 cgcggaactc catatatggg ctatgaacta atgaccccgt aattgattac tattaataac
     2641 taatgcatgg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     2701 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     2761 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     2821 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     2881 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     2941 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     3001 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     3061 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     3121 aggattagca gagcgaggta tgtaggcggt gatacagagt tcttgaagtg gtggcctaac
     3181 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     3241 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     3301 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     3361 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat ttttggtcat
     3421 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     3481 aatctaaagt atatatgagt aacctgaggc tatggcaggg cctgccgccc cgacgttggc
     3541 tgcgagccct gggccttcac ccgaacttgg ggggtggggt ggggaaaagg aagaaacgcg
     3601 ggcgtattgg ccccaatggg gtctcggtgg ggtatcgaca gagtgccagc cctgggaccg
     3661 aaccccgcgt ttatgaacaa acgacccaac accgtgcgtt ttattctgtc tttttattgc
     3721 cgtcatagcg cgggttcctt ccggtattgt ctccttccgt gtttcagtta gcctccccct
     3781 agggtgggcg aagaactcca gcatgagatc cccgcgctgg aggatcatcc agccggcgtc
     3841 ccggaaaacg attccgaagc ccaacctttc atagaaggcg gcggtggaat cgaaatctcg
     3901 tgatggcagg ttgggcgtcg cttggtcggt catttcgaac cccagagtcc cgctcagaag
     3961 aactcgtcaa gaaggcgata gaaggcgatg cgctgcgaat cgggagcggc gataccgtaa
     4021 agcacgagga agcggtcagc ccattcgccg ccaagctctt cagcaatatc acgggtagcc
     4081 aacgctatgt cctgatagcg gtccgccaca cccagccggc cacagtcgat gaatccagaa
     4141 aagcggccat tttccaccat gatattcggc aagcaggcat cgccatgggt cacgacgaga
     4201 tcctcgccgt cgggcatgct cgccttgagc ctggcgaaca gttcggctgg cgcgagcccc
     4261 tgatgctctt cgtccagatc atcctgatcg acaagaccgg cttccatccg agtacgtgct
     4321 cgctcgatgc gatgtttcgc ttggtggtcg aatgggcagg tagccggatc aagcgtatgc
     4381 agccgccgca ttgcatcagc catgatggat actttctcgg caggagcaag gtgagatgac
     4441 aggagatcct gccccggcac ttcgcccaat agcagccagt cccttcccgc ttcagtgaca
     4501 acgtcgagca cagctgcgca aggaacgccc gtcgtggcca gccacgatag ccgcgctgcc
     4561 tcgtcttgca gttcattcag ggcaccggac aggtcggtct tgacaaaaag aaccgggcgc
     4621 ccctgcgctg acagccggaa cacggcggca tcagagcagc cgattgtctg ttgtgcccag
     4681 tcatagccga atagcctctc cacccaagcg gccggagaac ctgcgtgcaa tc
//
