LOCUS       sequence-324986-        5496 bp    DNA     circular SYN 15-APR-2025
DEFINITION  expression of luciferase in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5496)
  TITLE     Improved system for intracellular delivery of biologic therapeutics 
            by nanocage-containing extracellular vesicles.
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5496)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: NanoLuc-Jun pCMV
FEATURES             Location/Qualifiers
     source          1..5496
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    1..625
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(647..667)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        652..982
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      833..968
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          1021..1117
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     primer_bind     complement(1193..1212)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1194..1212)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1227..1243)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1227..1243)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1240..1262)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1251..1267
                     /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(1275..1281,1282..1299,1300..1305))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1320..1341
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1458..1475)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1628..2216)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1708..1727)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2387..3178,3179..3247))
                     /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     3010..3029
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3248..3352)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      3379..3807
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3466..3485)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3676..3697
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3973..3995
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3987..4004
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3988..4004
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        4165..4544
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4545..4748
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4699..4719
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             4825..5337
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
ORIGIN
        1 gggtggcatc cctgtgaccc ctccccagtg cctctcctgg ccttggaagt tgccactcca
       61 gtgcccacca gccttgtcct aataaaatta agttgcatca ttttgtctga ctaggtgtcc
      121 tctataatat tatggggtgg aggggggtgg tatggagcaa ggggcccaag ttgggaagac
      181 aacctgtagg gcctgcgggg tctattcggg aaccaagctg gagtgcagtg gcacaatctt
      241 ggctcactgc aatctccgcc tcctgggttc aagcgattct cctgcctcag cctcccgagt
      301 tgttgggatt ccaggcatgc atgaccaggc tcagctaatt tttgtttttt tggtagagac
      361 ggggtttcac catattggcc aggctggtct ccaactccta atctcaggtg atctacccac
      421 cttggcctcc caaattgctg ggattacagg cgtgaaccac tgctcccttc cctgtccttc
      481 tgattttaaa ataactatac cagcaggagg acgtccagac acagcatagg ctacctgcca
      541 tggcccaacc ggtgggacat ttgagttgct tgcttggcac tgtcctctca tgcgttgggt
      601 ccactcagta gatgcctgtt gaattgggta cgcggccagc ttctgtggaa tgtgtgtcag
      661 ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
      721 aattagtcag caaccaggtg tggaaaagtc cccaggctcc ccagcaggca gaagtatgca
      781 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
      841 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
      901 tgcagaggcc gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt
      961 tggaggccta ggcttttgca aaaagctcct cgaggaactg aaaaaccaga aagttaactg
     1021 gtaagtttag tctttttgtc ttttatttca ggtcccggat ccggtggtgg tgcaaatcaa
     1081 agaactgctc ctcagtggat gttgccttta cttctaggcc tgtacggaag tgttacttct
     1141 gctctaaaag ctgctgcagg tcgactctag ggccgcgtac cgagctcgaa ttccctatag
     1201 tgagtcgtat taaattcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
     1261 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
     1321 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
     1381 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
     1441 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
     1501 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     1561 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaag gccgcgttgc
     1621 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     1681 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     1741 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     1801 cgggaagcgt ggcgctttct caatgctcac gctgtaggta tctcagttcg gtgtaggtcg
     1861 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     1921 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     1981 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     2041 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     2101 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     2161 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     2221 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     2281 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     2341 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     2401 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     2461 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     2521 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     2581 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     2641 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     2701 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     2761 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     2821 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     2881 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     2941 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     3001 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     3061 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     3121 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     3181 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     3241 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     3301 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     3361 cccgaaaagt gccacctgac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     3421 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     3481 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cggggcatcc
     3541 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     3601 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     3661 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     3721 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     3781 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacaaaatat taacgtttac
     3841 aatttcccca ttcgccattc aggctgcgca actgttggga agggcgatcg gtgcgggcct
     3901 cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa
     3961 cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgccaag ctgatctata
     4021 cattgaatca atattggcaa ttagccatat tagtcattgg ttatatagca taaatcaata
     4081 ttggctattg gccattgcat acgttgtatc tatatcataa tatgtacatt tatattggct
     4141 catgtccaat atgaccgcca tgttgacatt gattattgac tagttattaa tagtaatcaa
     4201 ttacggggtc attagttcat agcccatata tggagttccg cgttacataa cttacggtaa
     4261 atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg
     4321 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
     4381 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtccgccc cctattgacg
     4441 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta cgggactttc
     4501 ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc
     4561 agtacaccaa tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca
     4621 ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta
     4681 ataaccccgc cccgttgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa
     4741 gcagagctcg tttagtgaac cgtcagaatt gtttttattt ttaattttct ttcaaatact
     4801 tccatcgaat tcagatctgg taccatggtc ttcacactcg aagatttcgt tggggactgg
     4861 cgacagacag ccggctacaa cctggaccaa gtccttgaac agggaggtgt gtccagtttg
     4921 tttcagaatc tcggggtgtc cgtaactccg atccaaagga ttgtcctgag cggtgaaaat
     4981 gggctgaaga tcgacatcca tgtcatcatc ccgtatgaag gtctgagcgg cgaccaaatg
     5041 ggccagatcg aaaaaatttt taaggtggtg taccctgtgg atgatcatca ctttaaggtg
     5101 atcctgcact atggcacact ggtaatcgac ggggttacgc cgaacatgat cgactatttc
     5161 ggacggccgt atgaaggcat cgccgtgttc gacggcaaaa agatcactgt aacagggacc
     5221 ctgtggaacg gcaacaaaat tatcgacgag cgcctgatca accccgacgg ctccctgctg
     5281 ttccgagtaa ccatcaacgg agtgaccggc tggcggctgt gcgaacgcat tctggcgggt
     5341 ggtggtggtg gcaagcttag aatcgcccgg ctggaggaaa aagtgaaaac cttgaaagct
     5401 cagaactcgg agctggcgtc cacggccaac atgctcaggg aacaggtggc acagcttaaa
     5461 cagaaagtca tgaaccacta gatctagagg atcccc
//
