LOCUS       sequence-417280-        6935 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tomato reporter plasmid targeting the human HAND1 gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6935)
  AUTHORS   Lynch AT, Douglas M, Kimber SJ, Birket MJ
  TITLE     The generation and validation of a dual cardiac HAND1-Tomato 
            NKX2-5-GFP human embryonic stem cell line UMANe002-A-3.
  JOURNAL   Stem Cell Res. 2024 Mar 4;77:103342. doi: 10.1016/j.scr.2024.103342.
  PUBMED    38460234
REFERENCE   2  (bases 1 to 6935)
  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: "Stem Cell 
            Res."; date: "2024-03-4"; volume: "77:103342. doi"; pages: " 
            10.1016/j.scr.2024.103342"
COMMENT     Sequence Label: pHAND1-T2A-dTom-PGK-Neo
FEATURES             Location/Qualifiers
     source          1..6935
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(43..65)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     165..184
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     378..400
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     392..409
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     393..409
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1214..1267
                     /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             1268..1969
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     primer_bind     complement(1401..1419)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1669..1688
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1863..1886
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     protein_bind    2006..2039
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        2104..2603
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2139..2162)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2536..2555
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        2615..2662
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2681..3484
                     /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="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2744..2763)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3354..3373
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3516..3533)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3522..3746
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    3803..3836
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(3852..3868)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3852..3868)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(4728..4744)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4728..4744)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4741..4763)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4752..4768
                     /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)."
     promoter        complement(join(4776..4782,4783..4800,4801..4806))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4821..4842
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4959..4976)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5130..5718)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5210..5229)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5889..6680,6681..6749))
                     /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     6512..6531
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6750..6854)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     join(6922..6935,1..5)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gaggcccttt cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct
       61 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
      121 cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat
      181 tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata
      241 ccgcatcagg cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
      301 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
      361 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg tgacgaaagc
      421 tcatccaggg acgagggaaa gataaatagg catcgagtct ctaaacggct gccccggact
      481 ggggatggtg caggccggct tctttcgagg tgcggcgcct cgaaagaagt tgttattggg
      541 cggatcccgg cccccacgcg tcgccgcctc cttcaagtag caccgcttgg ttcctgctga
      601 aagagcggga aggaggacgc taggccagct cgactccccg ctcttttacc tccgtatctg
      661 aggctcccag ctccggctta gggggcccca agcctgcatc caagccctga tgtttaaacc
      721 aggttgggag gctggcggaa gtgtgtgggg tgcagcgaga gcgggagaac cgtggatttc
      781 cacccctccc ttccaatatc ctgggatctc ccttaggtgt cctaaagcca aatcaacgcg
      841 cgtcagtttc accagacgca gaacattttc cttaccaaac ctcccgaagc tgcccagccc
      901 atcccgggtc ccaggaactc cgcgcataaa ggcaaattta cgagcctgtt tgcctatttc
      961 gcaaacagaa atccgtcact tatttaaaga ctcccagtta ctgctcccga caccttcccc
     1021 tcctcctcat cttaccccat gccgggcgat cggagcagaa aaattaaaac ctgtctctcc
     1081 cgcctctttt cttctgtgtg tcctttggca gcagcagcac gaaggttttc ctcctgccct
     1141 gggcccagtc gagaagagga ttaaaggacg caccggctgg ccgcagcaag tctgggcgct
     1201 ggagttaaac caggagggca ggggaagcct gctgacatgc ggggacgtgg aggaaaatcc
     1261 cggccccatg gtgagcaagg gcgaggaggt catcaaagag ttcatgcgct tcaaggtgcg
     1321 catggagggc tccatgaacg gccacgagtt cgagatcgag ggcgagggcg agggccgccc
     1381 ctacgagggc acccagaccg ccaagctgaa ggtgaccaag ggcggccccc tgcccttcgc
     1441 ctgggacatc ctgtcccccc agttcatgta cggctccaag gcgtacgtga agcaccccgc
     1501 cgacatcccc gattacaaga agctgtcctt ccccgagggc ttcaagtggg agcgcgtgat
     1561 gaacttcgag gacggcggtc tggtgaccgt gacccaggac tcctccctgc aggacggcac
     1621 gctgatctac aaggtgaaga tgcgcggcac caacttcccc cccgacggcc ccgtaatgca
     1681 gaagaagacc atgggctggg aggcctccac cgagcgcctg tacccccgcg acggcgtgct
     1741 gaagggcgag atccaccagg ccctgaagct gaaggacggc ggccactacc tggtggagtt
     1801 caagaccatc tacatggcca agaagcccgt gcaactgccc ggctactact acgtggacac
     1861 caagctggac atcacctccc acaacgagga ctacaccatc gtggaacagt acgagcgctc
     1921 cgagggccgc caccacctgt tcctgtacgg catggacgag ctgtacaagt gagcccaatt
     1981 ccgatcatat tcaataaccc ttaatataac ttcgtataat gtatgctata cgaagttatt
     2041 aggtctgaag aggagtttac gtccagccaa gctagcttgg ctgcaggtcg tcgaaattct
     2101 accgggtagg ggaggcgctt ttcccaaggc agtctggagc atgcgcttta gcagccccgc
     2161 tgggcacttg gcgctacaca agtggcctct ggcctcgcac acattccaca tccaccggta
     2221 ggcgccaacc ggctccgttc tttggtggcc ccttcgcgcc accttctact cctcccctag
     2281 tcaggaagtt cccccccgcc ccgcagctcg cgtcgtgcag gacgtgacaa atggaagtag
     2341 cacgtctcac tagtctcgtg cagatggaca gcaccgctga gcaatggaag cgggtaggcc
     2401 tttggggcag cggccaatag cagctttgct ccttcgcttt ctgggctcag aggctgggaa
     2461 ggggtgggtc cgggggcggg ctcaggggcg ggctcagggg cggggcgggc gcccgaaggt
     2521 cctccggagg cccggcattc tgcacgcttc aaaagcgcac gtctgccgcg ctgttctcct
     2581 cttcctcatc tccgggcctt tcgacctgca gcctgttgac aattaatcat cggcatagta
     2641 tatcggcata gtataatacg acaaggtgag gaactaaacc atgggatcgg ccattgaaca
     2701 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     2761 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     2821 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aggacgaggc
     2881 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     2941 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     3001 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     3061 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     3121 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     3181 gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc atgcccgacg gcgatgatct
     3241 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     3301 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     3361 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     3421 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     3481 ctgaggggat caattctcta gagctcgctg atcagcctcg actgtgcctt ctagttgcca
     3541 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     3601 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     3661 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     3721 tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctcgac
     3781 tagagcttgc ggaaccctta atataacttc gtataatgta tgctatacga agttattagg
     3841 tccctcgagg ggatccacta gttctagatc agtgagccga ggcccgcgcc gaggacctgg
     3901 ccaggccagc cactcctgaa gccccgggag gagaggaagg cagcggcgaa cgccaggctc
     3961 tgggctccgg cgactggtgc tacgcatccc gcggagcttc tgctgagcgc cggcaggtcg
     4021 tcggctgcaa ccacacactt ggatcgcacg tgcaatgtcc tttgattttt tttaatacat
     4081 taagagaaag agaaatatat atatatccac ccccagccca accgagggcg gcccttggcg
     4141 gcaacatgca agaaggaggg actgtcgaac ccaagggctc aaagacgcac tcttccaccc
     4201 ttttggagcg aatttagaac ctcagcccta tctccatttc cctatctggc tctttctctc
     4261 ttgtccctcc atatgatccg ccccgacgcc gtcttctcta attaaaatgc aataaggaat
     4321 caattctttt ctgcctgaga aagagaacca gacgcaggaa gatgaaaggc tgccctttgt
     4381 tcttcgaatc gtggtggttt tattttattt ttctttttgt cgctgcactt cctgtttagt
     4441 tccaagggaa acactttctc tctttctctg tctctctctt ttcttccttc ttttccttcc
     4501 tttttgtttc tatctaaata aaagctttcc ctgtgttgga aagtttttat gtatttaaac
     4561 tacctaccat gcctgttgtg ctcaggtgtt tgttcatcct gccatcccca acccttttct
     4621 acctcaagtc tgtgtgacca ctcacaggta cctcgcgaat gcatctagat atcggatccc
     4681 gggcccgtcg actgcagagg cctgcatgca agcttggcgt aatcatggtc atagctgttt
     4741 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     4801 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     4861 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     4921 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     4981 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     5041 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     5101 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     5161 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     5221 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     5281 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     5341 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     5401 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     5461 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     5521 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     5581 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     5641 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     5701 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     5761 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     5821 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     5881 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     5941 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     6001 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     6061 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     6121 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     6181 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     6241 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     6301 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     6361 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     6421 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     6481 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     6541 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     6601 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     6661 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     6721 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     6781 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     6841 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga aaccattatt
     6901 atcatgacat taacctataa aaataggcgt atcac
//
