LOCUS       sequence-426928-        6277 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For overexpression of Gal4-DBD HOXD4 AroPERFECT shuffled 1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6277)
  AUTHORS   Naderi J, Magalhaes AP, Kibar G, Stik G, Zhang Y, Mackowiak SD, 
            Wieler HM, Rossi F, Buschow R, Christou-Kent M, Alcoverro-Bertran M,
            Graf T, Vingron M, Hnisz D
  TITLE     An activity-specificity trade-off encoded in human transcription 
            factors.
  JOURNAL   Nat Cell Biol. 2024 Aug;26(8):1309-1321. doi: 
            10.1038/s41556-024-01411-0. Epub 2024 Jul 5.
  PUBMED    38969762
REFERENCE   2  (bases 1 to 6277)
  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"; doi: 
            "10.1038/s41556-024-01411-0"; journalName: "Nat Cell Biol"; date: 
            "2024-08"; volume: "26"; issue: "8"; pages: "1309-1321"
COMMENT     Sequence Label: pGAL4-DBD-HOXD4-AroPERFECT_shuffled1
FEATURES             Location/Qualifiers
     source          1..6277
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             710..1150
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="DNA binding domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 DNA binding domain
                     /translation="MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTK
                     RSPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNK
                     DAVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVS"
     promoter        complement(1596..1614)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1597..1614)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1634..1651)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1640..1864
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1910..2338
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1997..2016)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2207..2228
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2347..2367)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2352..2681
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2532..2667
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2594..2613
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2748..3542
                     /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(2802..2821)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3412..3431
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3716..3837
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3753..3772)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3807..3826
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3886..3902)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3886..3902)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3899..3921)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3910..3926
                     /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(3934..3940,3941..3958,3959..3964))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3979..4000
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4117..4134)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4288..4876)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4368..4387)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5047..5838,5839..5907))
                     /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     5670..5689
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5908..6012)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6087..6106)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctctctgg ctaactagag
      601 aacccactgc ttactggctt atcgaaatta atacgactca ctatagggag acccaagctt
      661 ggtaccgagc tcggatccac tagtaacggc cgccagtgtg ctggaattca tgaagctact
      721 gtcttctatc gaacaagcat gcgatatttg ccgacttaaa aagctcaagt gctccaaaga
      781 aaaaccgaag tgcgccaagt gtctgaagaa caactgggag tgtcgctact ctcccaaaac
      841 caaaaggtct ccgctgacta gggcacatct gacagaagtg gaatcaaggc tagaaagact
      901 ggaacagcta tttctactga tttttcctcg agaagacctt gacatgattt tgaaaatgga
      961 ttctttacag gatataaaag cattgttaac aggattattt gtacaagata atgtgaataa
     1021 agatgccgtc acagatagat tggcttcagt ggagactgat atgcctctaa cattgagaca
     1081 gcatagaata agtgcgacat catcatcgga agagagtagt aacaaaggtc aaagacagtt
     1141 gactgtatcg gcgatcatgg gagtccaggc ccccaaagcg tatatgggaa agcctccaga
     1201 tggcgaattc cctgggctgc tggacggttc tcaatatgga ccaaaagacc tcgcggggcc
     1261 atacaaggaa ccccaggccc aggtggactt cccccccggt catcagagcg cctctttcgc
     1321 acccctgaag ccgccgccaa actttggggc agaggcagct ggcccctcct acccaggcct
     1381 gggcgtgggc gggtcttatc cagaggccca acaggaggcg gggtatccta gccccgggat
     1441 gtccatgaca tatcgctgtc agcggccggc gtcaccctat cttaagcctg gtccatgcgc
     1501 aggatacgtt cacggagagc ctccgccaga ttacgtgggt ggcccaggcc caagggtctg
     1561 gcagccagga gcaggttagg tctagagggc cctattctat agtgtcacct aaatgctaga
     1621 gctcgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     1681 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     1741 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     1801 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     1861 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     1921 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     1981 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     2041 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     2101 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     2161 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     2221 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     2281 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     2341 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     2401 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     2461 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     2521 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     2581 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     2641 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     2701 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     2761 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     2821 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     2881 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
     2941 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     3001 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     3061 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     3121 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     3181 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     3241 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
     3301 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     3361 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     3421 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     3481 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     3541 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     3601 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     3661 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
     3721 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     3781 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     3841 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc
     3901 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     3961 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     4021 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     4081 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     4141 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     4201 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     4261 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     4321 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     4381 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     4441 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     4501 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     4561 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     4621 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     4681 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     4741 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     4801 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     4861 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     4921 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     4981 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5041 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5101 atccatagtt gcctgacttc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     5161 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     5221 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     5281 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     5341 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     5401 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     5461 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     5521 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     5581 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     5641 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     5701 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     5761 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     5821 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     5881 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     5941 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6001 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat cgggagatct
     6061 cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     6121 tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa aatttaagct
     6181 acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt taggcgtttt
     6241 gcgctgcttc gcgatgtacg ggccagatat acgcgtt
//
