LOCUS       sequence-427262-        6232 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For overexpression of Gal4-DBD EGR1 AroPATCHY3.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6232)
  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 6232)
  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-EGR1-AroPATCHY3
FEATURES             Location/Qualifiers
     source          1..6232
                     /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(1551..1569)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1552..1569)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1589..1606)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1595..1819
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1865..2293
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1952..1971)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2162..2183
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2302..2322)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2307..2636
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2487..2622
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2549..2568
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2703..3497
                     /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(2757..2776)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3367..3386
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3671..3792
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3708..3727)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3762..3781
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3841..3857)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3841..3857)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3854..3876)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3865..3881
                     /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(3889..3895,3896..3913,3914..3919))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3934..3955
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4072..4089)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4243..4831)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4323..4342)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5002..5793,5794..5862))
                     /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     5625..5644
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5863..5967)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6042..6061)
                     /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 gcgatattgc ggcagaagga caagaaagca gacaaaagtg ttgtggcctc
     1201 ttcggccacc tcctctctct cttcctacta ctattacccg tccccggttg ctacctctcc
     1261 gtccccggtt actacctctc catccccggc caccacctca ccatcccctg tgcccacctc
     1321 ctcctctccc ggctcctcga ccccatcccc tgtgcacagt ggcccctccc cgtcggtggc
     1381 caccacgtac ttcttctact cctctgttcc ccctgctccg gcccaggtca gcagcccttc
     1441 ctcagctgtc accaactcct tcttcttttt cagcgcctcc acagggcttt cggacatgac
     1501 agcaacctct cccaggacaa ttgaaatttg ctgagtctag agggccctat tctatagtgt
     1561 cacctaaatg ctagagctcg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     1621 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     1681 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     1741 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg
     1801 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat
     1861 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     1921 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     1981 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     2041 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     2101 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     2161 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     2221 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     2281 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     2341 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     2401 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     2461 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     2521 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     2581 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     2641 tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga ggatcgtttc
     2701 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     2761 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     2821 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     2881 tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     2941 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     3001 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     3061 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     3121 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     3181 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg
     3241 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     3301 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     3361 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     3421 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     3481 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     3541 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     3601 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     3661 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     3721 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     3781 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     3841 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     3901 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     3961 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4021 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     4081 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     4141 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     4201 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     4261 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     4321 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     4381 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     4441 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     4501 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     4561 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     4621 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     4681 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     4741 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     4801 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     4861 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     4921 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     4981 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     5041 ctgtctattt cgttcatcca tagttgcctg acttcccgtc gtgtagataa ctacgatacg
     5101 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     5161 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     5221 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     5281 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     5341 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     5401 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     5461 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     5521 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     5581 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     5641 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     5701 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     5761 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     5821 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     5881 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     5941 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcga
     6001 cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg ctctgatgcc
     6061 gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg
     6121 agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt
     6181 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg tt
//
