LOCUS       sequence-59865-e        6988 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 6988)
  TITLE     Gene transfer to polarized airway epithelia
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6988)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pEIAV-SIN6.1 CBGFPW
FEATURES             Location/Qualifiers
     source          1..6988
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        146..525
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        526..729
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     680..700
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      2498..2507
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2504..2506,2507..2509,2510..3223)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2504..2506,2507..2509,2510..3220)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2549..2570)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2810..2829)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3157..3178
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3425..4013
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3478..3498)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3896..3907)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     rep_origin      4410..4865
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4497..4516)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4707..4728
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4934..4953)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5053..5075
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5113..5131)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5199..5303
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5304..5372,5373..6164)
                     /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     complement(5522..5541)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6335..6923
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6824..6843
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 acagatcttc aatattggcc attagccata ttattcattg gttatatagc ataaatcaat
       61 attggctatt ggccattgca tacgttgtat ctatatcata atatgtacat ttatattggc
      121 tcatgtccaa tatgaccgcc atgttggcat tgattattga ctagttatta atagtaatca
      181 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
      241 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
      301 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
      361 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtccgcc ccctattgac
      421 gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt acgggacttt
      481 cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg
      541 cagtacacca atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc
      601 attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt
      661 aataaccccg ccccgttgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata
      721 agcagagctc gtttagtgaa ccgggcactc agattctgcg gtctgagtcc cttctctgct
      781 gggctgaaaa ggcctttgta ataaatataa ttctctactc agtccctgtc tctagtttgt
      841 ctgttcgaga tcctacagtt ggcgcccgaa cagggacctg agaggggcgc agaccctacc
      901 tgttgaacct ggctgatcgt aggatccccg ggacagcaga ggagaactta cagaagtctt
      961 ctggaggtgt tcctggccag aacacaggag gacaggtaag tagggagacc ctttgacatg
     1021 gagcaaggcg ctcaagaagt tagagaaggt gacggtacaa gggtctcaga aattaactac
     1081 tggtaactgt aattgggcgc taagtctagt agacttattt catgatacca actttgtaaa
     1141 agaaaaggac tggcagctga gggatgtcat tccattgctg gaagatgtaa ctcagacgct
     1201 ggaattcggg tattgtagaa agggcaaata ggaccttgaa agagaagatt caaagtcata
     1261 gagacaacac tcaaacactg gaggcagctt tacaacttgc tctcattact tgtaacaaag
     1321 ggagggaaag tatgggagga cagacaccat gggaagtatt tatcactaat caagcacaag
     1381 taatacatga gaaactttta ctacagcaag cacaatcctc caaaaaattt tgtttttaca
     1441 aaatccctgg tgaacatgat tggaagggac ctactagggt gctgtggaag ggtgatggtg
     1501 cagtagtagt taatgatgaa ggaaagggaa taattgctgt accattaacc aggactaagt
     1561 tactaataaa gccaaattga gtattgttgc atgcatttga tatatgggat tatttggtaa
     1621 aggggtaaca tggtcagcat cgcattctat ggggggatcc cagggggaat ctcaacccct
     1681 attacccaac agtcagaaaa atctaagtgt gaggagaaca caatgtttca accttattgt
     1741 tataataatg acagtaagaa cagcatggca gaatcgaagg aagcaagaga ccaagaaatg
     1801 aacctgaaag aagaatctaa agaagaaaaa agaagaaatg actggtggaa aaaagctgca
     1861 gatagctagc ggtacctctg gtcgttacat aacttacggt aaatggcccg cctggctgac
     1921 cgcccaacga ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
     1981 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
     2041 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
     2101 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
     2161 ctcgaggcca cgttctgctt cactctcccc atctcccccc cctccccacc cccaattttg
     2221 tatttattta ttttttaatt attttgtgca gcgatggggg cggggggggg gggggggcgc
     2281 gcgccaggcg gggcggggcg gggcgagggg cggggcgggg cgaggcggag aggtgcggcg
     2341 gcagccaatc agagcggcgc gctccgaaag tttcctttta tggcgaggcg gcggcggcgg
     2401 cggccctata aaaagcgaag cgcgcggcgg gcgggagcgg gatcagccac cgcggtggcg
     2461 gcctagagtc gacggtaccg cgggcccggg atccatcgcc accatggtga gcaagggcga
     2521 ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca
     2581 caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa
     2641 gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac
     2701 ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa
     2761 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
     2821 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
     2881 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta
     2941 caacagccac aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt
     3001 caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa
     3061 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc
     3121 cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
     3181 cgccgccggg atcactctcg gcatggacga gctgtacaag taaagcggcc gcgagcatct
     3241 taccgccatt tattcccata tttgttctgt ttttcttgat ttgggtatac atttgaatgt
     3301 caataaaaca aaatggtggg gcaatcatct acatttcatg ggatatgtga ttactagttc
     3361 aggtgtattg ccacaagaca aacatgttaa gaaaatttcc cgttatttgc actctgttcc
     3421 tgttaatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     3481 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     3541 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
     3601 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
     3661 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
     3721 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
     3781 gctgttgggc actgacaatt ccgtggtgtt gtcggggaag ctgacgtcct ttccatggct
     3841 gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc
     3901 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg
     3961 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcctgtttc
     4021 gcctcggcgt ccggtccgtg ttgcttggtc ttcacctgtg cagacttgcg aaccatggat
     4081 tccaccgtga actttgtctc ctggcatgca aatcgtcaac ttggcatgcc aaatcgatta
     4141 gaaaaacaag gggggaactg tggggttttt atgaggggtt ttatacaatt gggcactcag
     4201 attctgcggt ctgagtccct tctctgctgg gctgaaaagg cctttgtaat aaatataatt
     4261 ctctactcag tccctgtctc tagtttgtct gttcgagatc ctacagagct catgccttgg
     4321 ttaattaagg agatccgggc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc
     4381 aacagttgcg cagcctgaat ggcgaatgga cgcgccctgt agcggcgcat taagcgcggc
     4441 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc
     4501 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     4561 tcgggggctc cctttagggt tccgatttag agctttacgg cacctcgacc gcaaaaaact
     4621 tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     4681 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     4741 ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt
     4801 aaaaaatgag ctgatttaac aaatatttaa cgcgaatttt aacaaaatat taacgtttac
     4861 aatttcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatat
     4921 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc
     4981 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     5041 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     5101 cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg
     5161 tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     5221 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     5281 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     5341 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     5401 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     5461 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     5521 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     5581 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     5641 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     5701 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     5761 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     5821 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     5881 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     5941 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     6001 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     6061 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     6121 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     6181 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     6241 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     6301 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     6361 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     6421 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     6481 tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     6541 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     6601 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     6661 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     6721 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     6781 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     6841 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     6901 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     6961 tttgctggcc ttttgctcac atggctcg
//
