LOCUS       sequence-465585-        4912 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of N-terminally 6His tagged Gcn4 DBD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4912)
  AUTHORS   Bremer A, Lang WH, Kempen RP, Sweta K, Taylor AB, Borgia MB, Ansari 
            AZ, Mittag T
  TITLE     Reconciling competing models on the roles of condensates and soluble
            complexes in transcription factor function.
  JOURNAL   bioRxiv [Preprint]. 2024 Nov 22:2024.11.21.624739. doi: 
            10.1101/2024.11.21.624739.
  PUBMED    39605529
REFERENCE   2  (bases 1 to 4912)
  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"; journalName: "bioRxiv 
            [Preprint]. 2024 Nov 22:2024.11.21.624739. doi: 
            10.1101/2024.11.21.624739."
COMMENT     Sequence Label: pDest17_DBD
FEATURES             Location/Qualifiers
     source          1..4912
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     21..40
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        21..39
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             71..93
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             113..130
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    140..164
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             167..187
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     protein_bind    complement(381..405)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(458..476)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      472..519
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(885..890,891..907,908..913))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(940..958)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1026..1130
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1131..1199,1200..1991)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1349..1368)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2162..2750
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2651..2670
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2904..2921
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2936..3075
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3001..3020)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3161..3183
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3177..3368)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     4830..4849
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 agatctcgat cccgcgaaat taatacgact cactataggg agaccacaac ggtttccctc
       61 tagaaataat tttgtttaac tttaagaagg agatatacat atgtcgtact accatcacca
      121 tcaccatcac ctcgaatcaa caagtttgta caaaaaagca ggcttcgaga acctgtattt
      181 tcagggtagc atgagtccgg aaagcagcga tccggcagca ctgaaacgtg cccgtaatac
      241 cgaagcagca cgtcgtagcc gtgcacgtaa actgcagcgt atgaaacagt tagaagataa
      301 ggttgaagaa ctgctgtgca aaaactatca cctggaaaat gaagttgccc gtctgaaaaa
      361 actggttggt gaacgttaag acccagcttt cttgtacaaa gtggttgatt cgaggctgct
      421 aacaaagccc gaaaggaagc tgagttggct gctgccaccg ctgagcaata actagcataa
      481 ccccttgggg cctctaaacg ggtcttgagg ggttttttgc tgaaaggagg aactatatcc
      541 ggatatccac aggacgggtg tggtcgccat gatcgcgtag tcgatagtgg ctccaagtag
      601 cgaagcgagc aggactgggc ggcggccaaa gcggtcggac agtgctccga gaacgggtgc
      661 gcatagaaat tgcatcaacg catatagcgc tagcagcacg ccatagtgac tggcgatgct
      721 gtcggaatgg acgatatccc gcaagaggcc cggcagtacc ggcataacca agcctatgcc
      781 tacagcatcc agggtgacgg tgccgaggat gacgatgagc gcattgttag atttcataca
      841 cggtgcctga ctgcgttagc aatttaactg tgataaacta ccgcattaaa gcttatcgat
      901 gataagctgt caaacatgag aattcttgaa gacgaaaggg cctcgtgata cgcctatttt
      961 tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa
     1021 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     1081 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     1141 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     1201 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     1261 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     1321 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtgttgacg
     1381 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     1441 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     1501 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     1561 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     1621 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgcagcaa
     1681 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     1741 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     1801 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     1861 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     1921 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     1981 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     2041 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     2101 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     2161 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     2221 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     2281 tcagcagagc gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact
     2341 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     2401 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     2461 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     2521 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     2581 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     2641 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     2701 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     2761 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     2821 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     2881 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga
     2941 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag
     3001 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
     3061 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     3121 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     3181 aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg
     3241 tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc
     3301 agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt
     3361 ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg
     3421 aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa
     3481 cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag
     3541 ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat
     3601 cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt
     3661 tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag
     3721 cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa
     3781 ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggccag
     3841 gacccaacgc tgcccgagat gcgccgcgtg cggctgctgg agatggcgga cgcgatggat
     3901 atgttctgcc aagggttggt ttgcgcattc acagttctcc gcaagaattg attggctcca
     3961 attcttggag tggtgaatcc gttagcgagg tgccgccggc ttccattcag gtcgaggtgg
     4021 cccggctcca tgcaccgcga cgcaacgcgg ggaggcagac aaggtatagg gcggcgccta
     4081 caatccatgc caacccgttc catgtgctcg ccgaggcggc ataaatcgcc gtgacgatca
     4141 gcggtccaat gatcgaagtt aggctggtaa gagccgcgag cgatccttga agctgtccct
     4201 gatggtcgtc atctacctgc ctggacagca tggcctgcaa cgcgggcatc ccgatgccgc
     4261 cggaagcgag aagaatcata atggggaagg ccatccagcc tcgcgtcgcg aacgccagca
     4321 agacgtagcc cagcgcgtcg gccgccatgc cggcgataat ggcctgcttc tcgccgaaac
     4381 gtttggtggc gggaccagtg acgaaggctt gagcgagggc gtgcaagatt ccgaataccg
     4441 caagcgacag gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc
     4501 agagcgctgc cggcacctgt cctacgagtt gcatgataaa gaagacagtc ataagtgcgg
     4561 cgacgatagt catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg
     4621 gcatcggtcg atcgacgctc tcccttatgc gactcctgca ttaggaagca gcccagtagt
     4681 aggttgaggc cgttgagcac cgccgccgca aggaatggtg catgcaagga gatggcgccc
     4741 aacagtcccc cggccacggg gcctgccacc atacccacgc cgaaacaagc gctcatgagc
     4801 ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg cgatataggc gccagcaacc
     4861 gcacctgtgg cgccggtgat gccggccacg atgcgtccgg cgtagaggat cg
//
