LOCUS       sequence-429188-        6247 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For overexpression of Gal4-DBD NANOG AroLITE A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6247)
  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 6247)
  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-NANOG-AroLITE_A
FEATURES             Location/Qualifiers
     source          1..6247
                     /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(1566..1584)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1567..1584)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1604..1621)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1610..1834
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1880..2308
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1967..1986)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2177..2198
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2317..2337)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2322..2651
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2502..2637
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2564..2583
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2718..3512
                     /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(2772..2791)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3382..3401
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3686..3807
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3723..3742)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3777..3796
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3856..3872)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3856..3872)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3869..3891)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3880..3896
                     /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(3904..3910,3911..3928,3929..3934))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3949..3970
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4087..4104)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4258..4846)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4338..4357)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5017..5808,5809..5877))
                     /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     5640..5659
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5878..5982)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6057..6076)
                     /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 gcgataaaac aggtgaagac cgcagcacag aaccagagaa tgaaatctaa
     1201 gagggcacag aaaaacaacg caccgaagaa tagcaatggt gtgacgcaga aggcctcagc
     1261 acctaccgca cccagccttg catcttccgc acaccaggga tgcctggtga acccgactgg
     1321 gaaccttcca atggcaagca accagaccgc aaacaattca accgcaagca accagaccca
     1381 gaacatccag tccgcaagca accactccgc aaacactcag accgcatgca cccaatccgc
     1441 aaacaatcag gccgcaaaca gtcccgcagc aaactgtgga gaggaatctc tgcagtcctg
     1501 catgcaggca cagccaaatt ctcctgccag tgacttggag gctgccttgg tctagagggc
     1561 cctattctat agtgtcacct aaatgctaga gctcgctgat cagcctcgac tgtgccttct
     1621 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     1681 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     1741 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     1801 agcaggcatg ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg
     1861 ggctctaggg ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     1921 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     1981 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     2041 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     2101 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     2161 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     2221 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     2281 ctgatttaac aaaaatttaa cgcgaattaa ttctgtggaa tgtgtgtcag ttagggtgtg
     2341 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     2401 caaccaggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     2461 tcaattagtc agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc
     2521 ccagttccgc ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg
     2581 aggccgcctc tgcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag
     2641 gcttttgcaa aaagctcccg ggagcttgta tatccatttt cggatctgat caagagacag
     2701 gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt
     2761 gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg
     2821 ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg
     2881 gtgccctgaa tgaactgcag gacgaggcag cgcggctatc gtggctggcc acgacgggcg
     2941 ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg
     3001 gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca
     3061 tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc
     3121 accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc
     3181 aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca
     3241 aggcgcgcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga
     3301 atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg
     3361 cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg
     3421 aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg
     3481 ccttctatcg ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga
     3541 ccaagcgacg cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag
     3601 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     3661 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     3721 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     3781 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     3841 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     3901 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     3961 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     4021 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     4081 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     4141 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     4201 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     4261 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     4321 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     4381 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     4441 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     4501 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     4561 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     4621 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     4681 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4741 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4801 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     4861 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4921 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     4981 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     5041 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgacttc ccgtcgtgta
     5101 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     5161 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     5221 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     5281 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     5341 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     5401 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     5461 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     5521 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     5581 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     5641 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     5701 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     5761 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     5821 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     5881 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     5941 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6001 gccacctgac gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca
     6061 atctgctctg atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc
     6121 gctgagtagt gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc
     6181 atgaagaatc tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat
     6241 acgcgtt
//
