LOCUS       sequence-430434-        6106 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For overexpression of Gal4-DBD NGN2 AroPERFECT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6106)
  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 6106)
  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-NGN2-AroPERFECT
FEATURES             Location/Qualifiers
     source          1..6106
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     217..236
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     470..487
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    604..625
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(640..645,646..663,664..670)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    678..694
                     /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)."
     primer_bind     683..705
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     702..718
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     702..718
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    767..888
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(778..797)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     832..851
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1062..1856)
                     /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(1173..1192)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1783..1802
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1923..2252)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1937..2072
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(1991..2010)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     2237..2257
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(2266..2694)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2376..2397)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2588..2607
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    2740..2964
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     2953..2970
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     promoter        2990..3008
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2990..3007
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             complement(3283..3723)
                     /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"
     primer_bind     complement(3785..3804)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3786..3804)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(3849..4052)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3878..3898)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        4053..4432
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     4604..4623
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4698..4802
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4803..4871,4872..5663)
                     /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     complement(5021..5040)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(5834..6106,1..316)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
       61 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
      121 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
      181 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
      241 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
      301 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
      361 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
      421 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
      481 gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat
      541 tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc
      601 aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc
      661 tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca
      721 tgattacgcc aagctctagc tagaggtcga cggtatacag acatgataag atacattgat
      781 gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt
      841 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttgg ggtgggcgaa
      901 gaactccagc atgagatccc cgcgctggag gatcatccag ccggcgtccc ggaaaacgat
      961 tccgaagccc aacctttcat agaaggcggc ggtggaatcg aaatctcgtg atggcaggtt
     1021 gggcgtcgct tggtcggtca tttcgaaccc cagagtcccg ctcagaagaa ctcgtcaaga
     1081 aggcgataga aggcgatgcg ctgcgaatcg ggagcggcga taccgtaaag cacgaggaag
     1141 cggtcagccc attcgccgcc aagctcttca gcaatatcac gggtagccaa cgctatgtcc
     1201 tgatagcggt ccgccacacc cagccggcca cagtcgatga atccagaaaa gcggccattt
     1261 tccaccatga tattcggcaa gcaggcatcg ccatgggtca cgacgagatc ctcgccgtcg
     1321 ggcatgcgcg ccttgagcct ggcgaacagt tcggctggcg cgagcccctg atgctcttcg
     1381 tccagatcat cctgatcgac aagaccggct tccatccgag tacgtgctcg ctcgatgcga
     1441 tgtttcgctt ggtggtcgaa tgggcaggta gccggatcaa gcgtatgcag ccgccgcatt
     1501 gcatcagcca tgatggatac tttctcggca ggagcaaggt gagatgacag gagatcctgc
     1561 cccggcactt cgcccaatag cagccagtcc cttcccgctt cagtgacaac gtcgagcaca
     1621 gctgcgcaag gaacgcccgt cgtggccagc cacgatagcc gcgctgcctc gtcctgcagt
     1681 tcattcaggg caccggacag gtcggtcttg acaaaaagaa ccgggcgccc ctgcgctgac
     1741 agccggaaca cggcggcatc agagcagccg attgtctgtt gtgcccagtc atagccgaat
     1801 agcctctcca cccaagcggc cggagaacct gcgtgcaatc catcttgttc aatcatgcga
     1861 aacgatcctc atcctgtctc ttgatcagat ccgaaaatgg atatacaagc tcccgggagc
     1921 tttttgcaaa agcctaggcc tccaaaaaag cctcctcact acttctggaa tagctcagag
     1981 gcagaggcgg cctcggcctc tgcataaata aaaaaaatta gtcagccatg gggcggagaa
     2041 tgggcggaac tgggcggagt taggggcggg atgggcggag ttaggggcgg gactatggtt
     2101 gctgactaat tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt
     2161 ccacacctgg ttgctgacta attgagatgc atgctttgca tacttctgcc tgctggggag
     2221 cctggggact ttccacaccc taactgacac acattccaca gaattaattc gcgttaaatt
     2281 tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat
     2341 caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat
     2401 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     2461 tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     2521 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     2581 gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca
     2641 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtggggat
     2701 accccctaga gccccagctg gttctttccg cctcagaagc catagagccc accgcatccc
     2761 cagcatgcct gctattgtct tcccaatcct cccccttgct gtcctgcccc accccacccc
     2821 ccagaataga atgacaccta ctcagacaat gcgatgcaat ttcctcattt tattaggaaa
     2881 ggacagtggg agtggcacct tccagggtca aggaaggcac gggggagggg caaacaacag
     2941 atggctggca actagaaggc acagtcgagg ctgatcagcg agctctagca tttaggtgac
     3001 actatagaat agggccctct agacgataca atccctggct atggggaggt gaggtgcata
     3061 gcggtgcttg tcaggcggag gaggttgatc catatcgcta ccccaagcag gggatgcagg
     3121 ggagagggta caactaggag acgtactgta gttagatgat acgctacttg aaggggccgg
     3181 tgaatttgta caataggacg tgctagcagg gcttggggaa tcgccacttg aggacaacca
     3241 agcagcacta gccccgcctg gactaagcag tacagctatc gccgatacag tcaactgtct
     3301 ttgacctttg ttactactct cttccgatga tgatgtcgca cttattctat gctgtctcaa
     3361 tgttagaggc atatcagtct ccactgaagc caatctatct gtgacggcat ctttattcac
     3421 attatcttgt acaaataatc ctgttaacaa tgcttttata tcctgtaaag aatccatttt
     3481 caaaatcatg tcaaggtctt ctcgaggaaa aatcagtaga aatagctgtt ccagtctttc
     3541 tagccttgat tccacttctg tcagatgtgc cctagtcagc ggagaccttt tggttttggg
     3601 agagtagcga cactcccagt tgttcttcag acacttggcg cacttcggtt tttctttgga
     3661 gcacttgagc tttttaagtc ggcaaatatc gcatgcttgt tcgatagaag acagtagctt
     3721 catgaattcc agcacactgg cggccgttac tagtggatcc gagctcggta ccaagcttgg
     3781 gtctccctat agtgagtcgt attaatttcg ataagccagt aagcagtggg ttctctagtt
     3841 agccagagag ctctgcttat atagacctcc caccgtacac gcctaccgcc catttgcgtc
     3901 aatggggcgg agttgttacg acattttgga aagtcccgtt gattttggtg ccaaaacaaa
     3961 ctcccattga cgtcaatggg gtggagactt ggaaatcccc gtgagtcaaa ccgctatcca
     4021 cgcccattga tgtactgcca aaaccgcatc accatggtaa tagcgatgac taatacgtag
     4081 atgtactgcc aagtaggaaa gtcccataag gtcatgtact gggcataatg ccaggcgggc
     4141 catttaccgt cattgacgtc aatagggggc gtacttggca tatgatacac ttgatgtact
     4201 gccaagtggg cagtttaccg taaatactcc acccattgac gtcaatggaa agtccctatt
     4261 ggcgttacta tgggaacata cgtcattatt gacgtcaatg ggcgggggtc gttgggcggt
     4321 cagccaggcg ggccatttac cgtaagttat gtaacgcgga actccatata tgggctatga
     4381 actaatgacc ccgtaattga ttactattaa taactagtca ataatcaatg tcaacgcgta
     4441 tatctggccc gtacatcgcg aagcagcgca aaacgcctaa ccctaagcag attcttcatg
     4501 caattgtcgg tcaagccttg ccttgttgta gcttaaattt tgctcgcgca ctactcagcg
     4561 acctccaaca cacaagcagg gagcagatac tggcttaact atgcggcatc agagcagatt
     4621 gtactgagag tgcaccatag gggatcggga gatctcccga tccgtcgacg tcaggtggca
     4681 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     4741 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     4801 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     4861 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     4921 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     4981 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     5041 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     5101 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     5161 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     5221 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     5281 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     5341 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     5401 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     5461 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     5521 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     5581 acacgacggg aagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     5641 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     5701 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     5761 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     5821 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     5881 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     5941 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     6001 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     6061 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggtt
//
