LOCUS       sequence-422156-        7327 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 7327)
  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 7327)
  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-Neo-TNNT2-Blast
FEATURES             Location/Qualifiers
     source          1..7327
                     /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..1708
                     /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="GTMIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQG
                     RPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDL
                     LSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEE
                     HQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQD
                     IALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(968..987)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1578..1597
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1728..1745)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1734..1958
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     regulatory      2605..2614
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2611..3006
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     primer_bind     complement(3119..3138)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3184..3239
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3238..3257)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     misc_feature    3795..4324
                     /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(4470..4489)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4471..4489)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4496..4513)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4496..4512)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4505..4527)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4654..5109
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4741..4760)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4951..4972
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5135..5239
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5240..5308,5309..6100)
                     /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(5458..5477)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6271..6859
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6760..6779
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7013..7030
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7147..7168
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7183..7188,7189..7206,7207..7213)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7221..7237
                     /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     7226..7248
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7245..7261
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7245..7261
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        7282..7300
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    join(7313..7327,1..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 gctcgagggt accatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
      961 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     1021 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     1081 cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     1141 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     1201 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     1261 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     1321 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     1381 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     1441 gcgcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     1501 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     1561 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     1621 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     1681 ctatcgcctt cttgacgagt tcttctgagt ttaaacccgc tgatcagcct cgactgtgcc
     1741 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     1801 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     1861 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga
     1921 caatagcagg catgctgggg atgcggtggg ctctatggac tagcggtggc ggcctcgaca
     1981 ttgattattg actagtgtca tggagaagac ccaccttgca gatgtcctca ctggggctgg
     2041 cagagccggc aacctgccca aggctgctca gtccattagg agccagtagc ctggaagatg
     2101 tctttacccc cagcatcagt tcaagtggag cagcacataa ctcttgccct ctgccttcca
     2161 agattctggt gctgagactt atggagtgtc ttggaggttg ccttctgccc cccaaccctg
     2221 ctcccagctg gccctcccag gcctgggttg ctggcctctg ctttatcagg attctcaaga
     2281 gggacagctg gtttatgttg catgactgtt ccctgcatat ctgctctggt tttaaatagc
     2341 ttatctgagc agctggagga ccacatgggc ttatatggcg tggggtacat gttcctgtag
     2401 ccttgtccct ggcacctgcc aaaatagcag ccaacacccc ccacccccac cgccatcccc
     2461 ctgccccacc cgtcccctgt cgcacattcc tccctccgca gggctggctc accaggcccc
     2521 agcccacatg cctgcttaaa gccctctcca tcctctgcct cacccagtcc ccgctgagac
     2581 tgagcagacg cctccaggat cgcggccgcc atggccaagc ctttgtctca agaagaatcc
     2641 accctcattg aaagagcaac ggctacaatc aacagcatcc ccatctctga agactacagc
     2701 gtcgccagcg cagctctctc tagcgacggc cgcatcttca ctggtgtcaa tgtatatcat
     2761 tttactgggg gaccttgtgc agaactcgtg gtgctgggca ctgctgctgc tgcggcagct
     2821 ggcaacctga cttgtatcgt cgcgatcgga aatgagaaca ggggcatctt gagcccctgc
     2881 ggacggtgcc gacaggtgct tctcgatctg catcctggga tcaaagccat agtgaaggac
     2941 agtgatggac agccgacggc agttgggatt cgtgaattgc tgccctctgg ttatgtgtgg
     3001 gagggcctgc agctgcagta gcctcgaggc aattggaatt cactcctcag gtgcaggctg
     3061 cctatcagaa ggtggtggct ggtgtggcca atgccctggc tcacaaatac cactgagatc
     3121 tttttccctc tgccaaaaat tatggggaca tcatgaagcc ccttgagcat ctgacttctg
     3181 gctaataaag gaaatttatt ttcattgcaa tagtgtgttg gaattttttg tgtctctcac
     3241 tcggaaggac atatgggagg gcaaatcatt taaaacatca gaatgagtat ttggtttaga
     3301 gtttggcaac atatgcccat atgctggctg ccatgaacaa aggttggcta taaagaggtc
     3361 atcagtatat gaaacagccc cctgctgtcc attccttatt ccatagaaaa gccttgactt
     3421 gaggttagat tttttttata ttttgttttg tgttattttt ttctttaaca tccctaaaat
     3481 tttccttaca tgttttacta gccagatttt tcctcctctc ctgactactc ccagtcatag
     3541 ctgtccctct tctcttatgg agatccctcg acctgcagcc caagcttgga tccctcgagt
     3601 taattaaatc tagaagtcga cagtactaag ctttgacaga aaagccccat ccttaggcct
     3661 cctccttcct agtctcctga tattgggtct aacccccacc tcctgttagg cagattcctt
     3721 atctggtgac acacccccat ttcctggagc catctctctc cttgccagaa cctctaaggt
     3781 ttgcttacga tggagccaga gaggatcctg ggagggagag cttggcaggg ggtgggaggg
     3841 aaggggggga tgcgtgacct gcccggttct cagtggccac cctgcgctac cctctcccag
     3901 aacctgagct gctctgacgc ggctgtctgg tgcgtttcac tgatcctggt gctgcagctt
     3961 ccttacactt cccaagagga gaagcagttt ggaaaaacaa aatcagaata agttggtcct
     4021 gagttctaac tttggctctt cacctttcta gtccccaatt tatattgttc ctccgtgcgt
     4081 cagttttacc tgtgagataa ggccagtagc cagccccgtc ctggcagggc tgtggtgagg
     4141 aggggggtgt ccgtgtggaa aactcccttt gtgagaatgg tgcgtcctag gtgttcacca
     4201 ggtcgtggcc gcctctactc cctttctctt tctccatcct tctttcctta aagagtcccc
     4261 agtgctatct gggacatatt cctccgccca gagcagggtc ccgcttccct aaggccctgc
     4321 tctgggcttc tgggtttgag tccttggcaa gcccaggaga ggcgctcagg cttccctgtc
     4381 ccccttcctc gtccaccatc tcatgcccct ggctctcctg ccccttccct acaggggttc
     4441 ctggctctgc tctagcgatc gccaattcgc cctatagtga gtcgtattac aattcactgg
     4501 ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt aatcgccttg
     4561 cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc gatcgccctt
     4621 cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg
     4681 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     4741 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     4801 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     4861 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
     4921 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
     4981 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
     5041 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc
     5101 ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     5161 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     5221 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     5281 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     5341 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     5401 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     5461 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     5521 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     5581 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     5641 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     5701 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     5761 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     5821 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     5881 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     5941 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     6001 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     6061 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     6121 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     6181 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     6241 aaaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     6301 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     6361 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     6421 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     6481 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     6541 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     6601 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     6661 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     6721 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     6781 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     6841 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     6901 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     6961 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     7021 agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc
     7081 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
     7141 cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
     7201 ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga
     7261 ccatgattac gccaagctcg aaattaaccc tcactaaagg gaacaaaagc tgtgctttct
     7321 ctgacca
//
