LOCUS       sequence-422163-        7291 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 7291)
  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 7291)
  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-Zeo
FEATURES             Location/Qualifiers
     source          1..7291
                     /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..2985
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     primer_bind     complement(3083..3102)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3148..3203
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3202..3221)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     misc_feature    3759..4288
                     /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(4434..4453)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4435..4453)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4460..4477)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4460..4476)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4469..4491)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4618..5073
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4705..4724)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4915..4936
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5099..5203
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5204..5272,5273..6064)
                     /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(5422..5441)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6235..6823
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6724..6743
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6977..6994
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7111..7132
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7147..7152,7153..7170,7171..7177)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7185..7201
                     /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     7190..7212
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7209..7225
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7209..7225
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        7246..7264
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    join(7277..7291,1..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 atggccaagt tgaccagtgc cgttccggtg
     2641 ctcaccgcgc gcgacgtcgc cggagcggtc gagttctgga ccgaccggct cgggttctcc
     2701 cgggacttcg tggaggacga cttcgccggt gtggtccggg acgacgtgac cctgttcatc
     2761 agcgcggtcc aggaccaggt ggtgccggac aacaccctgg cctgggtgtg ggtgcgcggc
     2821 ctggacgagc tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg ggacgcctcc
     2881 gggccggcca tgaccgagat cggcgagcag ccgtgggggc gggagttcgc cctgcgcgac
     2941 ccggccggca actgcgtgca cttcgtggcc gaggagcagg actgacctcg aggaagggcg
     3001 aattcactcc tcaggtgcag gctgcctatc agaaggtggt ggctggtgtg gccaatgccc
     3061 tggctcacaa ataccactga gatctttttc cctctgccaa aaattatggg gacatcatga
     3121 agccccttga gcatctgact tctggctaat aaaggaaatt tattttcatt gcaatagtgt
     3181 gttggaattt tttgtgtctc tcactcggaa ggacatatgg gagggcaaat catttaaaac
     3241 atcagaatga gtatttggtt tagagtttgg caacatatgc ccatatgctg gctgccatga
     3301 acaaaggttg gctataaaga ggtcatcagt atatgaaaca gccccctgct gtccattcct
     3361 tattccatag aaaagccttg acttgaggtt agattttttt tatattttgt tttgtgttat
     3421 ttttttcttt aacatcccta aaattttcct tacatgtttt actagccaga tttttcctcc
     3481 tctcctgact actcccagtc atagctgtcc ctcttctctt atggagatcc ctcgacctgc
     3541 agcccaagct tggatccctc gagttaatta aatctagaag tcgacagtac taagctttga
     3601 cagaaaagcc ccatccttag gcctcctcct tcctagtctc ctgatattgg gtctaacccc
     3661 cacctcctgt taggcagatt ccttatctgg tgacacaccc ccatttcctg gagccatctc
     3721 tctccttgcc agaacctcta aggtttgctt acgatggagc cagagaggat cctgggaggg
     3781 agagcttggc agggggtggg agggaagggg gggatgcgtg acctgcccgg ttctcagtgg
     3841 ccaccctgcg ctaccctctc ccagaacctg agctgctctg acgcggctgt ctggtgcgtt
     3901 tcactgatcc tggtgctgca gcttccttac acttcccaag aggagaagca gtttggaaaa
     3961 acaaaatcag aataagttgg tcctgagttc taactttggc tcttcacctt tctagtcccc
     4021 aatttatatt gttcctccgt gcgtcagttt tacctgtgag ataaggccag tagccagccc
     4081 cgtcctggca gggctgtggt gaggaggggg gtgtccgtgt ggaaaactcc ctttgtgaga
     4141 atggtgcgtc ctaggtgttc accaggtcgt ggccgcctct actccctttc tctttctcca
     4201 tccttctttc cttaaagagt ccccagtgct atctgggaca tattcctccg cccagagcag
     4261 ggtcccgctt ccctaaggcc ctgctctggg cttctgggtt tgagtccttg gcaagcccag
     4321 gagaggcgct caggcttccc tgtccccctt cctcgtccac catctcatgc ccctggctct
     4381 cctgcccctt ccctacaggg gttcctggct ctgctctagc gatcgccaat tcgccctata
     4441 gtgagtcgta ttacaattca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     4501 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     4561 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg
     4621 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4681 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     4741 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     4801 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     4861 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     4921 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     4981 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     5041 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     5101 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     5161 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     5221 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     5281 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     5341 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     5401 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     5461 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     5521 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     5581 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     5641 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     5701 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     5761 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     5821 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     5881 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     5941 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     6001 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     6061 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     6121 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     6181 tgagttttcg ttccactgag cgtcaaaccc cgtagaaaag atcaaaggat cttcttgaga
     6241 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     6301 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     6361 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     6421 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     6481 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     6541 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     6601 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     6661 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     6721 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     6781 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     6841 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     6901 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     6961 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     7021 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
     7081 ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc
     7141 ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca
     7201 atttcacaca ggaaacagct atgaccatga ttacgccaag ctcgaaatta accctcacta
     7261 aagggaacaa aagctgtgct ttctctgacc a
//
