LOCUS       sequence-306660-        6209 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 6209)
  AUTHORS   Gallegos LL, Kunkel MT, Newton AC
  TITLE     Targeting protein kinase C activity reporter to discrete 
            intracellular regions reveals spatiotemporal differences in 
            agonist-dependent signaling.
  JOURNAL   J Biol Chem. 2006 Oct 13. 281(41):30947-56.
  PUBMED    16901905
REFERENCE   2  (bases 1 to 6209)
  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: "J Biol 
            Chem. 2006 Oct 13. 281(41):30947-56."
COMMENT     Sequence Label: GolgiCFP
FEATURES             Location/Qualifiers
     source          1..6209
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(1030..1032,1033..1746)
                     /codon_start=1
                     /product="monomeric super enhanced variant of CFP (Matsuda 
                     et al., 2008)"
                     /label=mseCFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKA
                     HFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             join(1030..1032,1033..1746)
                     /codon_start=1
                     /product="cyan variant of GFP"
                     /label=CFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKA
                     HFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             join(1030..1032,1033..1746)
                     /codon_start=1
                     /product="enhanced CFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mECFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKA
                     HFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1072..1093)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1333..1352)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1680..1701
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(1762..1780)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1763..1780)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1800..1817)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1806..2030
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2076..2504
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2163..2182)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2373..2394
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2513..2533)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2518..2847
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2698..2833
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2760..2779
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2914..3708
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2968..2987)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3578..3597
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3882..4003
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3919..3938)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3973..3992
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4052..4068)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4052..4068)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4065..4087)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4076..4092
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4100..4106,4107..4124,4125..4130))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4145..4166
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4283..4300)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4454..5042)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4534..4553)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5213..6004,6005..6073))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5836..5855
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6074..6178)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttatgggc
      901 aacttgaaga gcgtggccca ggagcctggg ccaccctgcg gacttggact gggtctggga
      961 cttggattgt gcggcaagca gggcccagcc accccggcag gtggaggtgg tggaggaggt
     1021 ggaggtggtg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat cctggtcgag
     1081 ctggacggcg acgtaaacgg ccacaggttc agcgtgtccg gcgagggcga gggcgatgcc
     1141 acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg
     1201 cccaccctcg tgaccaccct gacctggggc gtgcagtgct tcagccgcta ccccgaccac
     1261 atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca ggagcgtacc
     1321 atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt cgagggcgac
     1381 accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg caacatcctg
     1441 ggacacaagc tggagtacaa ctacatcagc cacaacgtct atatcaccgc cgacaagcag
     1501 aagaacggca tcaaggccca cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag
     1561 ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct gctgcccgac
     1621 aaccactacc tgagcaccca gtccaagctg agcaaagacc ccaacgagaa gcgcgatcac
     1681 atggtcctgc tggagttcgt gaccgccgcc gggatcactc acggcatgga cgagctgtac
     1741 aagtaatcta gagggcccta ttctatagtg tcacctaaat gctagagctc gctgatcagc
     1801 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     1861 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     1921 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     1981 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
     2041 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     2101 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     2161 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     2221 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     2281 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     2341 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     2401 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     2461 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
     2521 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     2581 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     2641 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     2701 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     2761 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
     2821 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
     2881 tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     2941 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     3001 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     3061 aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg
     3121 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     3181 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     3241 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     3301 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     3361 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     3421 ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc
     3481 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     3541 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     3601 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     3661 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     3721 ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg
     3781 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     3841 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt
     3901 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     3961 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt
     4021 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     4081 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     4141 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     4201 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     4261 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     4321 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     4381 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     4441 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     4501 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     4561 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     4621 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4681 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4741 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4801 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4861 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     4921 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4981 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     5041 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     5101 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     5161 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     5221 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     5281 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     5341 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     5401 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     5461 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     5521 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     5581 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     5641 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     5701 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     5761 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5821 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5881 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     5941 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     6001 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     6061 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     6121 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     6181 catttccccg aaaagtgcca cctgacgtc
//
