LOCUS       sequence-447935-        6953 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tomato reporter plasmid targeting the human SOX17 gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6953)
  AUTHORS   Lynch AT, Phillips N, Douglas M, Dorgnach M, Lin IH, Adamson AD, 
            Darieva Z, Whittle J, Hanley NA, Bobola N, Birket MJ
  TITLE     HAND1 level controls the specification of multipotent cardiac and 
            extraembryonic progenitors from human pluripotent stem cells.
  JOURNAL   EMBO J. 2025 Mar 31. doi: 10.1038/s44318-025-00409-0.
  PUBMED    40164946
REFERENCE   2  (bases 1 to 6953)
  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: "EMBO J. 
            2025 Mar 31. doi: 10.1038/s44318-025-00409-0."
COMMENT     Sequence Label: pSOX17-T2A-dTom-PGK-Neo
FEATURES             Location/Qualifiers
     source          1..6953
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             903..920
                     /codon_start=1
                     /product="HQHQHQ metal affinity tag"
                     /label=HQ tag
                     /note="The HQ tag binds to nickel and other metal ions much
                     like the 6xHis tag, but may allow elution at lower 
                     concentrations of imidazole (Godat et al., 2008)."
                     /translation="HQHQHQ"
     CDS             909..926
                     /codon_start=1
                     /product="HQHQHQ metal affinity tag"
                     /label=HQ tag
                     /note="The HQ tag binds to nickel and other metal ions much
                     like the 6xHis tag, but may allow elution at lower 
                     concentrations of imidazole (Godat et al., 2008)."
                     /translation="HQHQHQ"
     CDS             1197..1250
                     /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             1251..1952
                     /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(1384..1402)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1652..1671
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     1846..1869
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     protein_bind    1989..2022
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        2087..2586
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2122..2145)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2519..2538
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     promoter        2598..2645
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2664..3467
                     /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(2727..2746)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3337..3356
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3499..3516)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3505..3729
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    3786..3819
                     /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(3835..3851)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3835..3851)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(4732..4748)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4732..4748)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4745..4767)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4756..4772
                     /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(4780..4786,4787..4804,4805..4810))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4825..4846
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4963..4980)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5134..5722)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5214..5233)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5893..6684,6685..6753))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MRIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6516..6535
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6754..6858)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6926..6944
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgtgaa gcgggtggag ggcggcttcc
      421 tgcacggcct ggctgagccg caggcggccg cgctgggccc cgagggcggc cgcgtggcca
      481 tggacggcct gggcctccag ttccccgagc agggcttccc cgccggcccg ccgctgctgc
      541 ctccgcacat gggcggccac taccgcgact gccagagtct gggcgcgcct ccgctcgacg
      601 gctacccgtt gcccacgccc gacacgtccc cgctggacgg cgtggacccc gacccggctt
      661 tcttcgccgc cccgatgccc ggggactgcc cggcggccgg cacctacagc tacgcgcagg
      721 tctcggacta cgctggcccc ccggagcctc ccgccggtcc catgcacccc cgactcggcc
      781 cagagcccgc gggtccctcg attccgggcc tcctggcgcc acccagcgcc cttcacgtgt
      841 actacggcgc gatgggctcg cccggggcgg gcggcgggcg cggcttccag atgcagccgc
      901 aacaccagca ccagcaccag caccagcacc accccccggg ccccggacag ccgtcgcccc
      961 ctccggaggc actgccctgc cgggacggca cggaccccag tcagcccgcc gagctcctcg
     1021 gggaggtgga ccgcacggaa tttgaacagt atctgcactt cgtgtgcaag cctgagatgg
     1081 gcctccccta ccaggggcat gactccggtg tgaatctccc cgacagccac ggggccattt
     1141 cctcggtggt gtccgacgcc agctccgcgg tatattactg caactatcct gacgtggagg
     1201 gcaggggaag cctgctgaca tgcggggacg tggaggaaaa tcccggcccc atggtgagca
     1261 agggcgagga ggtcatcaaa gagttcatgc gcttcaaggt gcgcatggag ggctccatga
     1321 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
     1381 ccgccaagct gaaggtgacc aagggcggcc ccctgccctt cgcctgggac atcctgtccc
     1441 cccagttcat gtacggctcc aaggcgtacg tgaagcaccc cgccgacatc cccgattaca
     1501 agaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
     1561 gtctggtgac cgtgacccag gactcctccc tgcaggacgg cacgctgatc tacaaggtga
     1621 agatgcgcgg caccaacttc ccccccgacg gccccgtaat gcagaagaag accatgggct
     1681 gggaggcctc caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc gagatccacc
     1741 aggccctgaa gctgaaggac ggcggccact acctggtgga gttcaagacc atctacatgg
     1801 ccaagaagcc cgtgcaactg cccggctact actacgtgga caccaagctg gacatcacct
     1861 cccacaacga ggactacacc atcgtggaac agtacgagcg ctccgagggc cgccaccacc
     1921 tgttcctgta cggcatggac gagctgtaca agtgagccca attccgatca tattcaataa
     1981 cccttaatat aacttcgtat aatgtatgct atacgaagtt attaggtctg aagaggagtt
     2041 tacgtccagc caagctagct tggctgcagg tcgtcgaaat tctaccgggt aggggaggcg
     2101 cttttcccaa ggcagtctgg agcatgcgct ttagcagccc cgctgggcac ttggcgctac
     2161 acaagtggcc tctggcctcg cacacattcc acatccaccg gtaggcgcca accggctccg
     2221 ttctttggtg gccccttcgc gccaccttct actcctcccc tagtcaggaa gttccccccc
     2281 gccccgcagc tcgcgtcgtg caggacgtga caaatggaag tagcacgtct cactagtctc
     2341 gtgcagatgg acagcaccgc tgagcaatgg aagcgggtag gcctttgggg cagcggccaa
     2401 tagcagcttt gctccttcgc tttctgggct cagaggctgg gaaggggtgg gtccgggggc
     2461 gggctcaggg gcgggctcag gggcggggcg ggcgcccgaa ggtcctccgg aggcccggca
     2521 ttctgcacgc ttcaaaagcg cacgtctgcc gcgctgttct cctcttcctc atctccgggc
     2581 ctttcgacct gcagcctgtt gacaattaat catcggcata gtatatcggc atagtataat
     2641 acgacaaggt gaggaactaa accatgggat cggccattga acaagatgga ttgcacgcag
     2701 gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg
     2761 gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca
     2821 agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc
     2881 tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg
     2941 actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg
     3001 ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta
     3061 cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag
     3121 ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac
     3181 tgttcgccag gctcaaggcg cgcatgcccg acggcgatga tctcgtcgtg acccatggcg
     3241 atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg
     3301 gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg
     3361 aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg
     3421 attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgaggg gatcaattct
     3481 ctagagctcg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     3541 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     3601 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     3661 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg
     3721 gctctatggc ttctgaggcg gaaagaacca gctggggctc gactagagct tgcggaaccc
     3781 ttaatataac ttcgtataat gtatgctata cgaagttatt aggtccctcg aggggatcca
     3841 ctagttctag acaggtccct gatccgcccc agcctgtagg ccagaagcag tgttacacac
     3901 ttcctggagg agctaaggaa atcctcagac tcctgggttt ttgttgttgc tgttgttgtt
     3961 ttttaaaagg tgtgttggca tataatttat ggtaatttat tttgtctgcc acttgaacag
     4021 tttggggggg tgaggtttca tttaaaattt gttcagagat ttgtttccca tagttggatt
     4081 gtcaaaaccc tatttccaag ttcaagttaa ctagctttga atgtgtccca aaacagcttc
     4141 ctccatttcc tgaaagttta ttgatcaaag aaatgttgtc ctgggtgtgt tttttcaatc
     4201 ttctaaaaaa taaaatctgg aatcctgctt ttttgctcta ctagtacctc tgtcacacta
     4261 gtcttatcaa aaaccagttc ttaagatcaa tgttaagttt attagttaat gtaaatttct
     4321 catcctcgaa aagggtgaac ataaatgcct ttaaggagta tatctaaaaa taaacattag
     4381 gatatctaag tttgatgtaa ttgtttcagg aaggaaaaaa gaaaagcatt ctggaatgag
     4441 cctacttcaa gtaatcttag tttctaaaac taacagttaa tattttcaat tccagtatat
     4501 cactttaagt agaaggggat gtccaagtaa ttttggtttt ctaactgttg aatcataagc
     4561 ttgacctgcc cccagaggct ttttggatgt ttttatctgt gttttgccat ctctttacac
     4621 tcctcgacat tcagtttacc ttaatcttca cgtacctcgc gaatgcatct agatatcgga
     4681 tcccgggccc gtcgactgca gaggcctgca tgcaagcttg gcgtaatcat ggtcatagct
     4741 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     4801 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     4861 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     4921 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     4981 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     5041 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5101 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     5161 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     5221 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     5281 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     5341 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     5401 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     5461 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     5521 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     5581 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     5641 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     5701 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     5761 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     5821 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     5881 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     5941 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6001 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6061 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6121 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6181 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6241 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6301 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6361 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     6421 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6481 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     6541 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     6601 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     6661 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     6721 aataagggcg acacggaaat gttgaatacg catactcttc ctttttcaat attattgaag
     6781 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     6841 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     6901 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtc
//
