LOCUS       sequence-422147-        7522 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of cardiac troponin T.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7522)
  AUTHORS   Fetterman KA, Blancard M, Lyra-Leite DM, Vanoye CG, Fonoudi H, Jouni
            M, DeKeyser JL, Lenny B, Sapkota Y, George AL Jr, Burridge PW
  TITLE     Independent compartmentalization of functional, metabolic, and 
            transcriptional maturation of hiPSC-derived cardiomyocytes.
  JOURNAL   Cell Rep. 2024 May 28;43(5):114160. doi: 
            10.1016/j.celrep.2024.114160. Epub 2024 Apr 27.
  PUBMED    38678564
REFERENCE   2  (bases 1 to 7522)
  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: "Cell Rep.";
            date: "2024-05-28"; volume: "43(5):114160. doi"; pages: " 
            10.1016/j.celrep.2024.114160. Epub 2024 Apr 27"
COMMENT     Sequence Label: pAAVS1-Puro-TNNT2-Neo
FEATURES             Location/Qualifiers
     source          1..7522
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    796..821
                     /label=SA
                     /note="splice acceptor site"
     CDS             845..898
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             908..1507
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(908..927)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     1404..1424
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(1539..1556)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1545..1769
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     CDS             2422..3216
                     /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(2476..2495)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3086..3105
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3314..3333)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3379..3434
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3433..3452)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     misc_feature    3990..4519
                     /label=HA-R
                     /note="right homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     primer_bind     complement(4665..4684)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4666..4684)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4691..4708)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4691..4707)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4700..4722)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4849..5304
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4936..4955)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5146..5167
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5330..5434
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5435..5503,5504..6295)
                     /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(5653..5672)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6466..7054
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6955..6974
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7208..7225
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7342..7363
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7378..7383,7384..7401,7402..7408)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7416..7432
                     /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)."
     primer_bind     7421..7443
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7440..7456
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7440..7456
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        7477..7495
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    join(7508..7522,1..789)
                     /label=HA-L
                     /note="left homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
ORIGIN
        1 gcattctctc ccctgggcct gtgccgcttt ctgtctgcag cttgtggcct gggtcacctc
       61 tacggctggc ccagatcctt ccctgccgcc tccttcaggt tccgtcttcc tccactccct
      121 cttccccttg ctctctgctg tgttgctgcc caaggatgct ctttccggag cacttccttc
      181 tcggcgctgc accacgtgat gtcctctgag cggatcctcc ccgtgtctgg gtcctctccg
      241 ggcatctctc ctccctcacc caaccccatg ccgtcttcac tcgctgggtt cccttttcct
      301 tctccttctg gggcctgtgc catctctcgt ttcttaggat ggccttctcc gacggatgtc
      361 tcccttgcgt cccgcctccc cttcttgtag gcctgcatca tcaccgtttt tctggacaac
      421 cccaaagtac cccgtctccc tggctttagc cacctctcca tcctcttgct ttctttgcct
      481 ggacaccccg ttctcctgtg gattcgggtc acctctcact cctttcattt gggcagctcc
      541 cctacccccc ttacctctct agtctgtgct agctcttcca gccccctgtc atggcatctt
      601 ccaggggtcc gagagctcag ctagtcttct tcctccaacc cgggccccta tgtccacttc
      661 aggacagcat gtttgctgcc tccagggatc ctgtgtcccc gagctgggac caccttatat
      721 tcccagggcc ggttaatgtg gctctggttc tgggtacttt tatctgtccc ctccacccca
      781 cagtggggca agcttctgac ctcttctctt cctcccacag ggcctcgaga gatctggcag
      841 cggagagggc agaggaagtc ttctaacatg cggtgacgtg gaggagaatc ccggccctag
      901 gctcgagatg accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag
      961 ggccgtacgc accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga
     1021 tccggaccgc cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg
     1081 gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac
     1141 gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt
     1201 gagcggttcc cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc
     1261 caaggagccc gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg
     1321 tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc
     1381 cttcctggag acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt
     1441 caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg
     1501 tgcctgatct agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg
     1561 ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc
     1621 cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc
     1681 tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag
     1741 gcatgctggg gatgcggtgg gctctatgga ctagcggtgg cggcctcgac attgattatt
     1801 gactagtgtc atggagaaga cccaccttgc agatgtcctc actggggctg gcagagccgg
     1861 caacctgccc aaggctgctc agtccattag gagccagtag cctggaagat gtctttaccc
     1921 ccagcatcag ttcaagtgga gcagcacata actcttgccc tctgccttcc aagattctgg
     1981 tgctgagact tatggagtgt cttggaggtt gccttctgcc ccccaaccct gctcccagct
     2041 ggccctccca ggcctgggtt gctggcctct gctttatcag gattctcaag agggacagct
     2101 ggtttatgtt gcatgactgt tccctgcata tctgctctgg ttttaaatag cttatctgag
     2161 cagctggagg accacatggg cttatatggc gtggggtaca tgttcctgta gccttgtccc
     2221 tggcacctgc caaaatagca gccaacaccc cccaccccca ccgccatccc cctgccccac
     2281 ccgtcccctg tcgcacattc ctccctccgc agggctggct caccaggccc cagcccacat
     2341 gcctgcttaa agccctctcc atcctctgcc tcacccagtc cccgctgaga ctgagcagac
     2401 gcctccagga tcgcggccgc catgattgaa caagatggat tgcacgcagg ttctccggcc
     2461 gcttgggtgg agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat
     2521 gccgccgtgt tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg
     2581 tccggtgccc tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg
     2641 ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta
     2701 ttgggcgaag tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta
     2761 tccatcatgg ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc
     2821 gaccaccaag cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc
     2881 gatcaggatg atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg
     2941 ctcaaggcgc gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg
     3001 ccgaatatca tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt
     3061 gtggcggacc gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc
     3121 ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc
     3181 atcgccttct atcgccttct tgacgagttc ttctgacctc gaggaagggc gaattcactc
     3241 ctcaggtgca ggctgcctat cagaaggtgg tggctggtgt ggccaatgcc ctggctcaca
     3301 aataccactg agatcttttt ccctctgcca aaaattatgg ggacatcatg aagccccttg
     3361 agcatctgac ttctggctaa taaaggaaat ttattttcat tgcaatagtg tgttggaatt
     3421 ttttgtgtct ctcactcgga aggacatatg ggagggcaaa tcatttaaaa catcagaatg
     3481 agtatttggt ttagagtttg gcaacatatg cccatatgct ggctgccatg aacaaaggtt
     3541 ggctataaag aggtcatcag tatatgaaac agccccctgc tgtccattcc ttattccata
     3601 gaaaagcctt gacttgaggt tagatttttt ttatattttg ttttgtgtta tttttttctt
     3661 taacatccct aaaattttcc ttacatgttt tactagccag atttttcctc ctctcctgac
     3721 tactcccagt catagctgtc cctcttctct tatggagatc cctcgacctg cagcccaagc
     3781 ttggatccct cgagttaatt aaatctagaa gtcgacagta ctaagctttg acagaaaagc
     3841 cccatcctta ggcctcctcc ttcctagtct cctgatattg ggtctaaccc ccacctcctg
     3901 ttaggcagat tccttatctg gtgacacacc cccatttcct ggagccatct ctctccttgc
     3961 cagaacctct aaggtttgct tacgatggag ccagagagga tcctgggagg gagagcttgg
     4021 cagggggtgg gagggaaggg ggggatgcgt gacctgcccg gttctcagtg gccaccctgc
     4081 gctaccctct cccagaacct gagctgctct gacgcggctg tctggtgcgt ttcactgatc
     4141 ctggtgctgc agcttcctta cacttcccaa gaggagaagc agtttggaaa aacaaaatca
     4201 gaataagttg gtcctgagtt ctaactttgg ctcttcacct ttctagtccc caatttatat
     4261 tgttcctccg tgcgtcagtt ttacctgtga gataaggcca gtagccagcc ccgtcctggc
     4321 agggctgtgg tgaggagggg ggtgtccgtg tggaaaactc cctttgtgag aatggtgcgt
     4381 cctaggtgtt caccaggtcg tggccgcctc tactcccttt ctctttctcc atccttcttt
     4441 ccttaaagag tccccagtgc tatctgggac atattcctcc gcccagagca gggtcccgct
     4501 tccctaaggc cctgctctgg gcttctgggt ttgagtcctt ggcaagccca ggagaggcgc
     4561 tcaggcttcc ctgtccccct tcctcgtcca ccatctcatg cccctggctc tcctgcccct
     4621 tccctacagg ggttcctggc tctgctctag cgatcgccaa ttcgccctat agtgagtcgt
     4681 attacaattc actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc
     4741 aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc
     4801 gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatgggac gcgccctgta
     4861 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     4921 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     4981 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     5041 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     5101 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     5161 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     5221 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     5281 acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     5341 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     5401 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     5461 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     5521 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     5581 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     5641 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     5701 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     5761 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     5821 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     5881 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     5941 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     6001 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     6061 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     6121 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     6181 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     6241 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     6301 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     6361 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     6421 gttccactga gcgtcaaacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     6481 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     6541 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     6601 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     6661 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     6721 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     6781 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     6841 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     6901 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     6961 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     7021 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     7081 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     7141 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     7201 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct
     7261 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     7321 gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt
     7381 acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac
     7441 aggaaacagc tatgaccatg attacgccaa gctcgaaatt aaccctcact aaagggaaca
     7501 aaagctgtgc tttctctgac ca
//
