LOCUS       sequence-7371-ex        5834 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5834)
  AUTHORS   Meng X, Brodsky MH, Wolfe SA
  TITLE     A bacterial one-hybrid system for determining the DNA-binding 
            specificity of transcription factors.
  JOURNAL   Nat Biotechnol. 2005 Aug . 23(8):988-94.
  PUBMED    16041365
REFERENCE   2  (bases 1 to 5834)
  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: "Nat 
            Biotechnol. 2005 Aug . 23(8):988-94."
COMMENT     Sequence Label: pH3U3-mcs
FEATURES             Location/Qualifiers
     source          1..5834
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      12..98
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(633..1583)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLRNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKHDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      1631..1853
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(2345..2439)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2470..3264)
                     /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(2581..2600)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3191..3210
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     terminator      3419..3505
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      3597..3624
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     primer_bind     3702..3720
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(3785..3790,3791..3808,3809..3815)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     3825..3847
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3844..3860
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3856..4518
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     CDS             4566..5369
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLATGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     rep_origin      complement(5378..5833)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5515..5536)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5727..5746
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 tctagaggca tcaaataaaa cgaaaggctc agtcgaaaga ctgggccttt cgttttatct
       61 gttgtttgtc ggtgaacgct ctcctgagta ggacaaatcc gccgccctag acctagggta
      121 cgggttttgc tgcccgcaaa cgggctgttc tggtgttgct agtttgttat cagaatcgca
      181 gatccggctt caggtttgcc ggctgaaagc gctatttctt ccagaattgc catgattttt
      241 tccccacggg aggcgtcact ggctcccgtg ttgtcggcag ctttgattcg ataagcagca
      301 tcgcctgttt caggctgtct atgtgtgact gttgagctgt aacaagttgt ctcaggtgtt
      361 caatttcatg ttctagttgc tttgttttac tggtttcacc tgttctatta ggtgttacat
      421 gctgttcatc tgttacattg tcgatctgtt catggtgaac agctttaaat gcaccaaaaa
      481 ctcgtaaaag ctctgatgta tctatctttt ttacaccgtt ttcatctgtg catatggaca
      541 gttttccctt tgatatctaa cggtgaacag ttgttctact tttgtttgtt agtcttgatg
      601 cttcactgat agatacaaga gccataagaa cctcagatcc ttccgtattt agccagtatg
      661 ttctctagtg tggttcgttg tttttgcgtg agccatgaga acgaaccatt gagatcatgc
      721 ttactttgca tgtcactcaa aaattttgcc tcaaaactgg tgagctgaat ttttgcagtt
      781 aaagcatcgt gtagtgtttt tcttagtccg ttacgtaggt aggaatctga tgtaatggtt
      841 gttggtattt tgtcaccatt catttttatc tggttgttct caagttcggt tacgagatcc
      901 atttgtctat ctagttcaac ttggaaaatc aacgtatcag tcgggcggcc tcgcttatca
      961 accaccaatt tcatattgct gtaagtgttt aaatctttac ttattggttt caaaacccat
     1021 tggttaagcc ttttaaactc atggtagtta ttttcaagca ttaacatgaa cttaaattca
     1081 tcaaggctaa tctctatatt tgccttgtga gttttctttt gtgttagttc ttttaataac
     1141 cactcataaa tcctcataga gtatttgttt tcaaaagact taacatgttc cagattatat
     1201 tttatgaatt tttttaactg gaaaagataa ggcaatatct cttcactaaa aactaattct
     1261 aatttttcgc ttgagaactt ggcatagttt gtccactgga aaatctcaaa gcctttaacc
     1321 aaaggattcc tgatttccac agttctcgtc atcagctctc tggttgcttt agctaataca
     1381 ccataagcat tttccctact gatgttcatc atctgagcgt attggttata agtgaacgat
     1441 accgtccgtt ctttccttgt agggttttca atcgtggggt tgagtagtgc cacacagcat
     1501 aaaattagct tggtttcatg ctccgttaag tcatagcgac taatcgctag ttcatttgct
     1561 ttgaaaacaa ctaattcaga catacatctc aattggtcta ggtgatttta atcactatac
     1621 caattgagat gggctagtca atgataatta ctagtccttt tcctttgagt tgtgggtatc
     1681 tgtaaattct gctagacctt tgctggaaaa cttgtaaatt ctgctagacc ctctgtaaat
     1741 tccgctagac ctttgtgtgt tttttttgtt tatattcaag tggttataat ttatagaata
     1801 aagaaagaat aaaaaaagat aaaaagaata gatcccagcc ctgtgtataa ctcactactt
     1861 tagtcagttc cgcagtatta caaaaggatg tcgcaaacgc tgtttgctcc tctacaaaac
     1921 agaccttaaa accctaaagg cttaagtagc accctcgcaa gctcgggcaa atcgctgaat
     1981 attccttttg tctccgacca tcaggcacct gagtcgctgt ctttttcgtg acattcagtt
     2041 cgctgcgctc acggctctgg cagtgaatgg gggtaaatgg cactacaggc gccttttatg
     2101 gattcatgca aggaaactac ccataataca agaaaagccc gtcacgggct tctcagggcg
     2161 ttttatggcg ggtctgctat gtggtgctat ctgacttttt gctgttcagc agttcctgcc
     2221 ctctgatttt ccagtctgac cacttcggat tatcccgtga caggtcattc agactggcta
     2281 atgcacccag taaggcagcg gtatcatcaa caggcttacc cgtcttactg tccctagtgc
     2341 ttggattctc accaataaaa aacgcccggc ggcaaccgag cgttctgaac aaatccagat
     2401 ggagttctga ggtcattact ggatctatca acaggagtcc aagcgagctc tcgaacccca
     2461 gagtcccgct cagaagaact cgtcaagaag gcgatagaag gcgatgcgct gcgaatcggg
     2521 agcggcgata ccgtaaagca cgaggaagcg gtcagcccat tcgccgccaa gctcttcagc
     2581 aatatcacgg gtagccaacg ctatgtcctg atagcggtcc gccacaccca gccggccaca
     2641 gtcgatgaat ccagaaaagc ggccattttc caccatgata ttcggcaagc aggcatcgcc
     2701 atgggtcacg acgagatcct cgccgtcggg catgcgcgcc ttgagcctgg cgaacagttc
     2761 ggctggcgcg agcccctgat gctcttcgtc cagatcatcc tgatcgacaa gaccggcttc
     2821 catccgagta cgtgctcgct cgatgcgatg tttcgcttgg tggtcgaatg ggcaggtagc
     2881 cggatcaagc gtatgcagcc gccgcattgc atcagccatg atggatactt tctcggcagg
     2941 agcaaggtga gatgacagga gatcctgccc cggcacttcg cccaatagca gccagtccct
     3001 tcccgcttca gtgacaacgt cgagcacagc tgcgcaagga acgcccgtcg tggccagcca
     3061 cgatagccgc gctgcctcgt cctgcagttc attcagggca ccggacaggt cggtcttgac
     3121 aaaaagaacc gggcgcccct gcgctgacag ccggaacacg gcggcatcag agcagccgat
     3181 tgtctgttgt gcccagtcat agccgaatag cctctccacc caagcggccg gagaacctgc
     3241 gtgcaatcca tcttgttcaa tcatgcgaaa cgatcctcat cctgtctctt gatcagatct
     3301 tgatcccctg cgccatcaga tccttggcgg caagaaagcc atccagttta ctttgcaggg
     3361 cttcccaacc ttaccagagg gcgccccagc tggcaattcc gacgtcaccg gtaggcatca
     3421 aataaaacga aaggctcagt cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt
     3481 gaacgctctc ctgagtagga caaatccgcc gggagcggat ttgaacgttg cgaagcaacg
     3541 gcccggaggg tggcgggcag gacgcccgcc ataaactgcc aggcatcaaa ttaagcagaa
     3601 ggccatcctg acggatggcc tttttgcgtt tctacaaact cttttgttta tttttctaaa
     3661 tacattcaaa tatgtatccg ctcatgacat taacctataa aaataggcgt atcacgaggc
     3721 cctttcgtct tcaaacgcgt gtacacccgg gcggccgcgt gcactccgga ggcgcgccga
     3781 attctttaca ctttatgctt ccggctcgta tgttgtgtcg accgagcgga taacaatttc
     3841 acacaggaaa cagctatgac agagcagaaa gccctagtaa agcgtattac aaatgaaacc
     3901 aagattcaga ttgcgatctc tttaaagggt ggtcccctag cgatagagca ctcgatcttc
     3961 ccagaaaaag aggcagaagc agtagcagaa caggccacac aatcgcaagt gattaacgtc
     4021 cacacaggta tagggtttct ggaccatatg atacatgctc tggccaagca ttccggctgg
     4081 tcgctaatcg ttgagtgcat tggtgactta cacatagacg accatcacac cactgaagac
     4141 tgcgggattg ctctcggtca agcttttaaa gaggccctag gggccgtgcg tggagtaaaa
     4201 aggtttggat caggatttgc gcctttggat gaggcacttt ccagagcggt ggtagatctt
     4261 tcgaacaggc cgtacgcagt tgtcgaactt ggtttgcaaa gggagaaagt aggagatctc
     4321 tcttgcgaga tgatcccgca ttttcttgaa agctttgcag aggctagcag aattaccctc
     4381 cacgttgatt gtctgcgagg caagaatgat catcaccgta gtgagagtgc gttcaaggct
     4441 cttgcggttg ccataagaga agccacctcg cccaatggta ccaacgatgt tccctccacc
     4501 aaaggtgttc ttatgtagtg acaccgatta tttaaagctg cagggatccc acacaggaaa
     4561 cagctatgtc gaaagctaca tataaggaac gtgctgctac tcatcctagt cctgttgctg
     4621 ccaagctatt taatatcatg cacgaaaagc aaacaaactt gtgtgcttca ttggatgttc
     4681 gtaccaccaa ggaattactg gagttagttg aagcattagg tcccaaaatt tgtttactaa
     4741 aaacacatgt ggatatcttg actgattttt ccatggaggg cacagttaag ccgctaaagg
     4801 cattatccgc caagtacaat tttttactct tcgaagacag aaaatttgct gacattggta
     4861 atacagtcaa attgcagtac tctgcgggtg tatacagaat agcagaatgg gcagacatta
     4921 cgaatgcaca cggtgtggtg ggcccaggta ttgttagcgg tttgaagcag gcggcggaag
     4981 aagtaacaaa ggaacctaga ggccttttga tgttagcaga attgtcatgc aagggctccc
     5041 tagctactgg agaatatact aagggtactg ttgacattgc gaagagcgac aaagattttg
     5101 ttatcggctt tattgctcaa agagacatgg gtggaagaga tgaaggttac gattggttga
     5161 ttatgacacc cggtgtgggt ttagatgaca agggagacgc attgggtcaa cagtatagaa
     5221 ccgtggatga tgtggtctct acaggatctg acattattat tgttggaaga ggactatttg
     5281 caaagggaag ggatgctaag gtagagggtg aacgttacag aaaagcaggc tgggaagcat
     5341 atttgagaag atgcggccag caaaactaac tcgagctaaa ttgtaagcgt taatattttg
     5401 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     5461 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
     5521 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
     5581 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
     5641 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     5701 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     5761 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     5821 ctacagggcg cgtc
//
