LOCUS       sequence-254788-        4854 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 4854)
  AUTHORS   Gokirmak T, Campanale JP, Shipp LE, Moy GW, Tao H, Hamdoun A
  TITLE     Localization and Substrate Selectivity of Sea Urchin Multidrug (MDR)
            Efflux Transporters.
  JOURNAL   J Biol Chem. 2012 Nov 2.
  PUBMED    23124201
REFERENCE   2  (bases 1 to 4854)
  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. 2012 Nov 2."
COMMENT     Sequence Label: pCS2+8NeGFP
FEATURES             Location/Qualifiers
     source          1..4854
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        40..1017
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1052..1070
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1052..1069
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             1117..1830
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1147..1175)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1770..1790
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     polyA_signal    1923..2057
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1947..1966)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2001..2020
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2170..2190)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2170..2188)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2209..2225)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2209..2225)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2222..2244)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2233..2249
                     /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(2257..2263,2264..2281,2282..2287))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2302..2323
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2440..2457)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2611..3199)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2691..2710)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3370..4161,4162..4230))
                     /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     3993..4012
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4231..4335)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4361..4816)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4498..4519)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4710..4729
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 cgatcggtgc gggcctcttc gctattacgc cagtcgaccg ccaattcaat atggcgtata
       61 tggactcatg ccaattcaat atggtggatc tggacctgtg ccaattcaat atggcgtata
      121 tggactcgtg ccaattcaat atggtggatc tggaccccag ccaattcaat atggcggact
      181 tggcaccatg ccaattcaat atggcggacc tggcactgtg ccaactgggg aggggtctac
      241 ttggcacggt gccaagtttg aggaggggtc ttggccctgt gccaagtccg ccatattgaa
      301 ttggcatggt gccaataatg gcggccatat tggctatatg ccaggatcaa tatataggca
      361 atatccaata tggccctatg ccaatatggc tattggccag gttcaatact atgtattggc
      421 cctatgccat atagtattcc atatatgggt tttcctattg acgtagatag cccctcccaa
      481 tgggcggtcc catataccat atatggggct tcctaatacc gcccatagcc actcccccat
      541 tgacgtcaat ggtctctata tatggtcttt cctattgacg tcatatgggc ggtcctattg
      601 acgtatatgg cgcctccccc attgacgtca attacggtaa atggcccgcc tggctcaatg
      661 cccattgacg tcaataggac cacccaccat tgacgtcaat gggatggctc attgcccatt
      721 catatccgtt ctcacgcccc ctattgacgt caatgacggt aaatggccca cttggcagta
      781 catcaatatc tattaatagt aacttggcaa gtacattact attggaagta cgccagggta
      841 cattggcagt actcccattg acgtcaatgg cggtaaatgg cccgcgatgg ctgccaagta
      901 catccccatt gacgtcaatg gggaggggca atgacgcaaa tgggcgttcc attgacgtaa
      961 atgggcggta ggcgtgccta atgggaggtc tatataagca atgctcgttt agggaaccgc
     1021 cattctgcct ggggacgtcg gagcaagctt gatttaggtg acactataga atacaagcta
     1081 cttgttcttt ttgcaggatc cactagtcat cgattgatga gtaaaggaga agaacttttc
     1141 actggagttg tcccaattct tgttgaatta gatggtgatg ttaatgggca caaattttct
     1201 gtcagtggag agggtgaagg tgatgcaaca tacggaaaac ttacccttaa atttatttgc
     1261 actactggaa aactacctgt tccatggcca acacttgtca ctactctcac ttatggtgtt
     1321 caatgctttt caagataccc agatcatatg aaacggcatg actttttcaa gagtgccatg
     1381 cccgaaggtt atgtacagga aagaactata tttttcaaag atgacgggaa ctacaagaca
     1441 cgtgctgaag tcaagtttga aggtgatacc cttgttaata gaatcgagtt aaaaggtatt
     1501 gattttaaag aagatggaaa cattcttgga cacaaattgg aatacaacta taactcacac
     1561 aatgtataca tcatggcaga caaacaaaag aatggaatca aagttaactt caaaattaga
     1621 cacaacattg aagatggaag cgttcaacta gcagaccatt atcaacaaaa tactccaatt
     1681 ggcgatggcc ctgtcctttt accagacaac cattacctgt ccacacaatc tgccctttcg
     1741 aaagatccca acgaaaagag agaccacatg gtccttcttg agtttgtaac agctgctggg
     1801 attacacatg gcatggatga actatacaaa gaattcggag ccggcgcgcc attaattaaa
     1861 ggccggccag cgatcgccaa ggcctctcga gcctctagaa ctatagtgag tcgtattacg
     1921 tagatccaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
     1981 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
     2041 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
     2101 aggtgtggga ggttttttaa ttcgcggccg cggcgccaat gcattgggcc cggtacccag
     2161 cttttgttcc ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt
     2221 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg ggagcataaa
     2281 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     2341 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     2401 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     2461 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     2521 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     2581 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     2641 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     2701 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     2761 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     2821 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     2881 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     2941 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     3001 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     3061 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     3121 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     3181 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     3241 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     3301 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     3361 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     3421 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     3481 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     3541 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     3601 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     3661 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     3721 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     3781 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     3841 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     3901 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     3961 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     4021 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     4081 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     4141 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     4201 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     4261 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     4321 aataggggtt ccgcgcacat ttccccgaaa agtgccacct aaattgtaag cgttaatatt
     4381 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     4441 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     4501 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     4561 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     4621 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     4681 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     4741 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     4801 ccgctacagg gcgcgtccca ttcgccattc aggctgcgca actgttggga aggg
//
