LOCUS       sequence-356299-        8055 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses human FOXC1 and miRFP670 via T2A linker under control of a
            CMV promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8055)
  AUTHORS   Abatti LE, Lado-Fernandez P, Huynh L, Collado M, Hoffman MM, 
            Mitchell JA
  TITLE     Epigenetic reprogramming of a distal developmental enhancer cluster 
            drives SOX2 overexpression in breast and lung adenocarcinoma.
  JOURNAL   Nucleic Acids Res. 2023 Sep 22:gkad734. doi: 10.1093/nar/gkad734.
  PUBMED    37738673
REFERENCE   2  (bases 1 to 8055)
  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: "Nucleic 
            Acids Res. 2023 Sep 22:gkad734. doi: 10.1093/nar/gkad734."
COMMENT     Sequence Label: CMV-FOXC1-T2A-miRFP670
FEATURES             Location/Qualifiers
     source          1..8055
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        82..461
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        462..665
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     616..636
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     662..686
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             2432..2485
                     /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             2492..3436
                     /codon_start=1
                     /product="bright monomeric near-infrared fluorescent 
                     protein engineered from a bacterial phytochrome, with an 
                     emission maximum of 670 nm (Shcherbakova et al., 2016)"
                     /label=miRFP670
                     /label=ORF frame 1
                     /translation="VAGHASGSPAFGTASHSNCEHEEIHLAGSIQPHGALLVVSEHDHR
                     VIQASANAAEFLNLGSVLGVPLAEIDGDLLIKILPHLDPTAEGMPVAVRCRIGNPSTEY
                     CGLMHRPPEGGLIIELERAGPSIDLSGTLAPALERIRTAGSLRALCDDTVLLFQQCTGY
                     DRVMVYRFDEQGHGLVFSECHVPGLESYFGNRYPSSTVPQMARQLYVRQRVRVLVDVTY
                     QPVPLEPRLSPLTGRDLDMSGCFLRSMSPCHLQFLKDMGVRATLAVSLVVGGKLWGLVV
                     CHHYLPRFIRFELRAICKRLAERIATRITALES"
     primer_bind     complement(3473..3492)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3517..3565
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      3767..4195
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3854..3873)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4064..4085
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4204..4224)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4209..4538
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4389..4524
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4451..4470
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4605..5399
                     /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(4659..4678)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5269..5288
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5575..5696
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5612..5631)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5666..5685
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5745..5761)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5745..5761)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5758..5780)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5769..5785
                     /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(5793..5799,5800..5817,5818..5823))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5838..5859
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5976..5993)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6147..6735)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6227..6246)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6906..7697,7698..7766))
                     /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     7529..7548
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7767..7871)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7946..7965)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
       61 tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta
      121 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
      181 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
      241 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
      301 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
      361 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
      421 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
      481 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
      541 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
      601 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
      661 gagctcgttt agtgaaccgt cagatcgcct ggagacgcca tccacgctgt tttgacctcc
      721 atagaagaca ccgggaccga tccagcctcc ggactctaga gccaccatgc aggcgcgcta
      781 ctccgtgtcc agccccaact ccctgggagt ggtgccctac ctcggcggcg agcagagcta
      841 ctaccgcgcg gcggccgcgg cggccggggg cggctacacc gccatgccgg cccccatgag
      901 cgtgtactcg caccctgcgc acgccgagca gtacccgggc ggcatggccc gcgcctacgg
      961 gccctacacg ccgcagccgc agcccaagga catggtgaag ccgccctata gctacatcgc
     1021 gctcatcacc atggccatcc agaacgcccc ggacaagaag atcaccctga acggcatcta
     1081 ccagttcatc atggaccgct tccccttcta ccgggacaac aagcagggct ggcagaacag
     1141 catccgccac aacctctcgc tcaacgagtg cttcgtcaag gtgccgcgcg acgacaagaa
     1201 gccgggcaag ggcagctact ggacgctgga cccggactcc tacaacatgt tcgagaacgg
     1261 cagcttcctg cggcggcggc ggcgcttcaa gaagaaggac gcggtgaagg acaaggagga
     1321 gaaggacagg ctgcacctca aggagccgcc cccgcccggc cgccagcccc cgcccgcgcc
     1381 gccggagcag gccgacggca acgcgcccgg tccgcagccg ccgcccgtgc gcatccagga
     1441 catcaagacc gagaacggta cgtgcccctc gccgccccag cccctgtccc cggccgccgc
     1501 cctgggcagc ggcagcgccg ccgcggtgcc caagatcgag agccccgaca gcagcagcag
     1561 cagcctgtcc agcgggagca gccccccggg cagcctgccg tcggcgcggc cgctcagcct
     1621 ggacggtgcg gattccgcgc cgccgccgcc cgcgccctcc gccccgccgc cgcaccatag
     1681 ccagggcttc agcgtggaca acatcatgac gtcgctgcgg gggtcgccgc agagcgcggc
     1741 cgcggagctc agctccggcc ttctggcctc ggcggccgcg tcctcgcgcg cggggatcgc
     1801 acccccgctg gcgctcggcg cctactcgcc cggccagagc tccctctaca gctccccctg
     1861 cagccagacc tccagcgcgg gcagctcggg cggcggcggc ggcggcgcgg gggccgcggg
     1921 gggcgcgggc ggcgccggga cctaccactg caacctgcaa gccatgagcc tgtacgcggc
     1981 cggcgagcgc gggggccact tgcagggcgc gcccgggggc gcgggcggct cggccgtgga
     2041 cgaccccctg cccgactact ctctgcctcc ggtcaccagc agcagctcgt cgtccctgag
     2101 tcacggcggc ggcggcggcg gcggcggggg aggccaggag gccggccacc accctgcggc
     2161 ccaccaaggc cgcctcacct cgtggtacct gaaccaggcg ggcggagacc tgggccactt
     2221 ggcgagcgcg gcggcggcgg cggcggccgc aggctacccg ggccagcagc agaacttcca
     2281 ctcggtgcgg gagatgttcg agtcacagag gatcggcttg aacaactctc cagtgaacgg
     2341 gaatagtagc tgtcaaatgg ccttcccttc cagccagtct ctgtaccgca cgtccggagc
     2401 tttcgtctac gactgtagca agtttgctag cgagggcaga ggaagtcttc taacatgcgg
     2461 tgacgtggag gagaatcccg gccctggtac cgtagcaggt catgcctctg gcagccccgc
     2521 attcgggacc gcctctcatt cgaattgcga acatgaagag atccacctcg ccggctcgat
     2581 ccagccgcat ggcgcgcttc tggtcgtcag cgaacatgat catcgcgtca tccaggccag
     2641 cgccaacgcc gcggaatttc tgaatctcgg aagcgtactc ggcgttccgc tcgccgagat
     2701 cgacggcgat ctgttgatca agatcctgcc gcatctcgat cccaccgccg aaggcatgcc
     2761 ggtcgcggtg cgctgccgga tcggcaatcc ctctacggag tactgcggtc tgatgcatcg
     2821 gcctccggaa ggcgggctga tcatcgaact cgaacgtgcc ggcccgtcga tcgatctgtc
     2881 aggcacgctg gcgccggcgc tggagcggat ccgcacggcg ggttcactgc gcgcgctgtg
     2941 cgatgacacc gtgctgctgt ttcagcagtg caccggctac gaccgggtga tggtgtatcg
     3001 tttcgatgag caaggccacg gcctggtatt ctccgagtgc catgtgcctg ggctcgaatc
     3061 ctatttcggc aaccgctatc cgtcgtcgac tgtcccgcag atggcgcggc agctgtacgt
     3121 gcggcagcgc gtccgcgtgc tggtcgacgt cacctatcag ccggtgccgc tggagccgcg
     3181 gctgtcgccg ctgaccgggc gcgatctcga catgtcgggc tgcttcctgc gctcgatgtc
     3241 gccgtgccat ctgcagttcc tgaaggacat gggcgtgcgc gccaccctgg cggtgtcgct
     3301 ggtggtcggc ggcaagctgt ggggcctggt tgtctgtcac cattatctgc cgcgcttcat
     3361 ccgtttcgag ctgcgggcga tctgcaaacg gctcgccgaa aggatcgcga cgcggatcac
     3421 cgcgcttgag agctaaaccg gttagtaatg agtttaaacg ggggaggcta actgaaacac
     3481 ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga cagaataaaa
     3541 cgcacgggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg ctggcactct
     3601 gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc cttttcccca
     3661 ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg ggcggcaggc
     3721 cctgccatag cagatctgcg cagctggggc tctagggggt atccccacgc gccctgtagc
     3781 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     3841 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     3901 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     3961 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     4021 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     4081 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     4141 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     4201 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     4261 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     4321 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     4381 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     4441 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     4501 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     4561 ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     4621 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     4681 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     4741 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     4801 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     4861 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     4921 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     4981 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     5041 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     5101 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     5161 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     5221 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     5281 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     5341 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     5401 cgggactctg gggttcgcga aatgaccgac caagcgacgc ccaacctgcc atcacgagat
     5461 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc
     5521 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg
     5581 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     5641 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat
     5701 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct
     5761 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     5821 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     5881 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     5941 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     6001 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     6061 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     6121 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     6181 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     6241 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     6301 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     6361 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     6421 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     6481 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     6541 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     6601 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     6661 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     6721 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     6781 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     6841 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     6901 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     6961 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     7021 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     7081 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     7141 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     7201 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     7261 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     7321 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     7381 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     7441 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     7501 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     7561 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     7621 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     7681 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     7741 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     7801 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     7861 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc
     7921 ccgatcccct atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
     7981 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
     8041 caacaaggca aggct
//
