LOCUS       sequence-391335-        4955 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a chimera of human transferrin receptor (hTfR) and GFP in 
            mammalian cells..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4955)
  AUTHORS   Roh H, Dorner BG, Ting AY
  TITLE     Cell-Type-Specific Intracellular Protein Delivery with Inactivated 
            Botulinum Neurotoxin.
  JOURNAL   J Am Chem Soc. 2023 May 10;145(18):10220-10226. doi: 
            10.1021/jacs.3c01145. Epub 2023 Apr 26.
  PUBMED    37098770
REFERENCE   2  (bases 1 to 4955)
  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"; doi: 
            "10.1021/jacs.3c01145"; journalName: "J Am Chem Soc"; date: 
            "2023-05-10- 10"; volume: "145"; issue: "18"; pages: "10220-10226"
COMMENT     Sequence Label: pAAV_CMV::hTfR-GFP
FEATURES             Location/Qualifiers
     source          1..4955
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   651..780
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        851..1154
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1155..1358
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1309..1329
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1403..1422
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1403..1421
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(1861..1863,1864..1866,1867..2577)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1861..1863,1864..1866,1867..2577)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1906..1927)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2167..2186)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2514..2535
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2626..2643)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2632..2839
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2868..3008
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2868..2997
                     /label=AAV2 ITR
     rep_origin      3083..3538
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3170..3189)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3380..3401
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3555..3574)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3674..3696
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3734..3752)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3820..3924
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3925..3993,3994..4785)
                     /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(4143..4162)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt cctgcaggca
      661 gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt ggtcgcccgg
      721 cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact aggggttcct
      781 gcggccgcac gcgttgtaca tagtaatcaa ttacggggtc attagttcat agcccatata
      841 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      901 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      961 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
     1021 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
     1081 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
     1141 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
     1201 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
     1261 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
     1321 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
     1381 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgccacc
     1441 atgatggatc aagctagatc agcattctct aacttgtttg gtggagaacc attgtcatat
     1501 acccggttca gcctggctcg gcaagtagat ggcgataaca gtcatgtgga gatgaaactt
     1561 gctgtagatg aagaagaaaa tgctgacaat aacacaaagg ccaatgtcac aaaaccaaaa
     1621 aggtgtagtg gaagtatctg ctatgggact attgctgtga tcgtcttttt cttgattgga
     1681 tttatgattg gctacttggg cggggagcag ccgccacagt accagtatcc cccacagtac
     1741 atgcagccac cgccgccgcc accgcagcag ccagccggag gacagtcatc gctggtggat
     1801 ggggcaggag caggagacgg agcaggagga tcggcaggag ctggcgatca cggcggcgtg
     1861 atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     1921 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
     1981 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     2041 ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag
     2101 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     2161 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     2221 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     2281 aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac
     2341 ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
     2401 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     2461 tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     2521 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagcac
     2581 cggtccacgt gataagaatt cctcgagcta gagctcgctg atcagcctcg actgtgcctt
     2641 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
     2701 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
     2761 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca
     2821 atagcaggca tgctggggag gtaaccacgt gcggaccgag cggccgcagg aacccctagt
     2881 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
     2941 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg
     3001 cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     3061 catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     3121 gtggttacgc gcagcgtgac cgctacactt gccagcgcct tagcgcccgc tcctttcgct
     3181 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     3241 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     3301 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     3361 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caactctatc
     3421 tcgggctatt cttttgattt ataagggatt ttgccgattt cggtctattg gttaaaaaat
     3481 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta
     3541 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     3601 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     3661 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     3721 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     3781 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     3841 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     3901 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     3961 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     4021 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     4081 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     4141 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     4201 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     4261 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     4321 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     4381 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     4441 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     4501 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     4561 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     4621 tctggagccg gtgagcgtgg aagccgcggt atcattgcag cactggggcc agatggtaag
     4681 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     4741 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     4801 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     4861 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     4921 gcgtcagacc ccgtagaaaa gatcaaagga tcttc
//
