LOCUS       sequence-429186-        6274 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For overexpression of Gal4-DBD HOXD4 AroPERFECT-1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6274)
  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 6274)
  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-1
FEATURES             Location/Qualifiers
     source          1..6274
                     /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(1593..1611)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1594..1611)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1631..1648)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1637..1861
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1907..2335
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1994..2013)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2204..2225
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2344..2364)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2349..2678
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2529..2664
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2591..2610
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2745..3539
                     /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(2799..2818)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3409..3428
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3713..3834
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3750..3769)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3804..3823
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3883..3899)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3883..3899)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3896..3918)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3907..3923
                     /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(3931..3937,3938..3955,3956..3961))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3976..3997
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4114..4131)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4285..4873)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4365..4384)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5044..5835,5836..5904))
                     /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     5667..5686
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5905..6009)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6084..6103)
                     /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 tgatgtcatc aatggtttac aattctaaag ttgatcctaa
     1201 accatttcca tgtgaagaac ttcaaggtgg gtatcttggg gaacaaggag ctgatggtta
     1261 tggaggggca caaggggcgg atcaatttcc acctggcctg cccaggccag attttggcga
     1321 acaaccaggc gggagtggtt ttcctggccc aggatcagcc cttccatacg caaggggaca
     1381 tggtcaggaa ccctacggag gtcctggcgg acatgctgcg tatcctgggg aaccatgtcc
     1441 ggcgccatat cccgcacccc cacctgcacc tctctatcca ggagcaaggg caagccaaag
     1501 ctatgatcct aaacaacccc cttcaggcta tacagccctg aaacaacctg cggtttgggt
     1561 accaatgaag aaagtcgtct agagggccct attctatagt gtcacctaaa tgctagagct
     1621 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     1681 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     1741 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     1801 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg
     1861 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc
     1921 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     1981 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2041 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     2101 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     2161 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     2221 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     2281 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     2341 tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta
     2401 tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag
     2461 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa
     2521 ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc catggctgac
     2581 taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt
     2641 agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat
     2701 ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg
     2761 gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac
     2821 aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg
     2881 ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc
     2941 ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg
     3001 aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc
     3061 accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc
     3121 ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta
     3181 ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg
     3241 cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg
     3301 tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat
     3361 tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc
     3421 gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta
     3481 tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag
     3541 cgggactctg gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt
     3601 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     3661 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     3721 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     3781 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     3841 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     3901 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     3961 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     4021 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4081 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4141 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4201 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4261 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4321 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4381 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4441 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4501 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4561 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4621 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4681 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     4741 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     4801 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     4861 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     4921 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     4981 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5041 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5101 catagttgcc tgacttcccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5161 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5221 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5281 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5341 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5401 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5461 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5521 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5581 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     5641 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     5701 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     5761 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     5821 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     5881 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     5941 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6001 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc
     6061 gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
     6121 ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca
     6181 acaaggcaag gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg
     6241 ctgcttcgcg atgtacgggc cagatatacg cgtt
//
