LOCUS       sequence-54339-e        7443 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 7443)
  AUTHORS   Zilio N, Wehrkamp-Richter S, Boddy MN
  TITLE     A new versatile system for rapid control of gene expression in the 
            fission yeast Schizosaccharomyces pombe.
  JOURNAL   Yeast. 2012 Aug 29. doi: 10.1002/yea.2920.
  PUBMED    22968950
REFERENCE   2  (bases 1 to 7443)
  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: "Yeast. 2012
            Aug 29. doi: 10.1002/yea.2920."
COMMENT     Sequence Label: pUG6SP-tTA
FEATURES             Location/Qualifiers
     source          1..7443
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     45..64
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        45..63
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    complement(117..150)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            complement(213..1569)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     terminator      complement(213..410)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(416..1225)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1139..1158
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(1226..1569)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     protein_bind    complement(1624..1657)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     enhancer        1763..2142
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2143..2346
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2297..2317
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2343..2367
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(2452..3072,3073..3459)
                     /codon_start=1
                     /product="tetracycline-controlled transactivator, 
                     comprising a fusion of the tetracycline repressor TetR with
                     the C-terminal activation domain of herpes simplex virus 
                     VP16"
                     /label=tTA
                     /note="In the Tet-Off(R) system, tTA binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTI
                     EGLLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHA
                     DALDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     primer_bind     complement(2525..2544)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        complement(5142..5160)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(5143..5160)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(5241..5260)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5360..5382
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5420..5438)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5506..5610
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5611..5679,5680..6471)
                     /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(5829..5848)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6642..7230
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7131..7150
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7384..7401
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 cgttggccga ttcattaatg caggttaacc tggcttatcg aaattaatac gactcactat
       61 agggagaccg gcagatccgc ggccgcatag gccactagtg gatctgatat cacctaataa
      121 cttcgtatag catacattat acgaagttat attaagggtt ctcgagagct cgttttcgac
      181 actggatggc ggcgttagta tcgaatcgac agcagtatag cgaccagcat tcacatacga
      241 ttgacgcatg atattacttt ctgcgcactt aacttcgcat ctgggcagat gatgtcgagg
      301 cgaaaaaaaa tataaatcac gctaacattt gattaaaata gaacaactac aatataaaaa
      361 aactatacaa atgacaagtt cttgaaaaca agaatctttt tattgtcagt actgattaga
      421 aaaactcatc gagcatcaaa tgaaactgca atttattcat atcaggatta tcaataccat
      481 atttttgaaa aagccgtttc tgtaatgaag gagaaaactc accgaggcag ttccatagga
      541 tggcaagatc ctggtatcgg tctgcgattc cgactcgtcc aacatcaata caacctatta
      601 atttcccctc gtcaaaaata aggttatcaa gtgagaaatc accatgagtg acgactgaat
      661 ccggtgagaa tggcaaaagc ttatgcattt ctttccagac ttgttcaaca ggccagccat
      721 tacgctcgtc atcaaaatca ctcgcatcaa ccaaaccgtt attcattcgt gattgcgcct
      781 gagcgagacg aaatacgcga tcgctgttaa aaggacaatt acaaacagga atcgaatgca
      841 accggcgcag gaacactgcc agcgcatcaa caatattttc acctgaatca ggatattctt
      901 ctaatacctg gaatgctgtt ttgccgggga tcgcagtggt gagtaaccat gcatcatcag
      961 gagtacggat aaaatgcttg atggtcggaa gaggcataaa ttccgtcagc cagtttagtc
     1021 tgaccatctc atctgtaaca tcattggcaa cgctaccttt gccatgtttc agaaacaact
     1081 ctggcgcatc gggcttccca tacaatcgat agattgtcgc acctgattgc ccgacattat
     1141 cgcgagccca tttataccca tataaatcag catccatgtt ggaatttaat cgcggcctcg
     1201 aaacgtgagt cttttcctta cccatggttg tttatgttcg gatgtgatgt gagaactgta
     1261 tcctagcaag attttaaaag gaagtatatg aaagaagaac ctcagtggca aatcctaacc
     1321 ttttatattt ctctacaggg gcgcggcgtg gggacaattc aacgcgtctg tgaggggagc
     1381 gtttccctgc tcgcaggtct gcagcgagga gccgtaattt ttgcttcgcg ccgtgcggcc
     1441 atcaaaatgt atggatgcaa atgattatac atggggatgt atgggctaaa tgtacgggcg
     1501 acagtcacat catgcccctg agctgcgcac gtcaagactg tcaaggaggg tattctgggc
     1561 ctccatgtcg ctggccgggt gacccggcgg ggacgaggca agctaaacag atctctagac
     1621 ctaataactt cgtatagcat acattatacg aagttatatt aagggttgtc gacgctcgag
     1681 gagcttggcc cattgcatac gttgtatcca tatcataata tgtacattta tattggctca
     1741 tgtccaacat taccgccatg ttgacattga ttattgacta gttattaata gtaatcaatt
     1801 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     1861 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     1921 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     1981 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     2041 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     2101 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     2161 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     2221 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     2281 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     2341 agagctcgtt tagtgaaccg tcagatcgcc tggagacgcc atccacgctg ttttgacctc
     2401 catagaagac accgggaccg atccagcctc cgcggccccg aattaattca tatgtctaga
     2461 ttagataaaa gtaaagtgat taacagcgca ttagagctgc ttaatgaggt cggaatcgaa
     2521 ggtttaacaa cccgtaaact cgcccagaag ctaggtgtag agcagcctac attgtattgg
     2581 catgtaaaaa ataagcgggc tttgctcgac gccttagcca ttgagatgtt agataggcac
     2641 catactcact tttgcccttt agaaggggaa agctggcaag attttttacg taataacgct
     2701 aaaagtttta gatgtgcttt actaagtcat cgcgatggag caaaagtaca tttaggtaca
     2761 cggcctacag aaaaacagta tgaaactctc gaaaatcaat tagccttttt atgccaacaa
     2821 ggtttttcac tagagaatgc attatatgca ctcagcgctg tggggcattt tactttaggt
     2881 tgcgtattgg aagatcaaga gcatcaagtc gctaaagaag aaagggaaac acctactact
     2941 gatagtatgc cgccattatt acgacaagct atcgaattat ttgatcacca aggtgcagag
     3001 ccagccttct tattcggcct tgaattgatc atatgcggat tagaaaaaca acttaaatgt
     3061 gaaagtgggt ccgcgtacag ccgcgcgcgt acgaaaaaca attacgggtc taccatcgag
     3121 ggcctgctcg atctcccgga cgacgacgcc cccgaagagg cggggctggc ggctccgcgc
     3181 ctgtcctttc tccccgcggg acacacgcgc agactgtcga cggccccccc gaccgatgtc
     3241 agcctggggg acgagctcca cttagacggc gaggacgtgg cgatggcgca tgccgacgcg
     3301 ctagacgatt tcgatctgga catgttgggg gacggggatt ccccgggtcc gggatttacc
     3361 ccccacgact ccgcccccta cggcgctctg gatatggccg acttcgagtt tgagcagatg
     3421 tttaccgatg cccttggaat tgacgagtac ggtgggtagg gggcgcgagg atcgtaataa
     3481 gcttcgacct gcagcgtacg aagcttcagc tggggtaaaa ggaatgtctc ccttgccagt
     3541 actgctaggg tttttctttc aaactatgga agcccattca agctgcatat tacgattttg
     3601 tttttcgctt ttagaaagtg gtttagatga gataatagaa aaattcttga tctccgacaa
     3661 cgagtacttt tatttttttt gctaatcact ttactcaata ttagctcgaa atcgtagaaa
     3721 cgtagacggg tgcgggatac cgagtggtgt agttaagaat ttttataaac cacgtggccc
     3781 aaaaatatga acccaaaacg tttatacatg agtatacttt aagaaggcta taccccttcg
     3841 tgttagatgt agttttagct acccaacccg agtctatgag cttgacttca gatgtagaag
     3901 gcattaaatc gttttgaata ttaattaaaa aacgatgaaa attaaatatt taaaagcaat
     3961 catacgctga aaatttagtg ctgtggctaa tccttcaaca tggaaatgcc ataaaagtga
     4021 ctttgacaaa aaaaaaagta tatacaggta gtaaactcat ctacttcatt gactttgttt
     4081 acagcatgtg gaaggaggaa tatttattgc taaatcgtag tttaacattc aataagtaat
     4141 actattgaaa ttcgacaaga ttggccgcat ggatgaaaaa gaggcatttt gctttgggag
     4201 aattagttca aattagaact gaaaaaaaaa actttacgag gcaaaaatgt cggattgaga
     4261 tcgtaaaagt tcgctcgtcg tcttttgctt tgtgattgtt ttcatggata catcttgctg
     4321 gatatttaaa ttttagtact atgtataaga tattctataa atgttttatc acccaaacct
     4381 gttagcgcct tcttaattct attcaatctg gcttttgctc tgagactact tcttggactt
     4441 tcactacttg ttagttatac ggaatttgtg taattagaag tgaaataatc ctttctatta
     4501 gtaatgcgag ctccttgatt tatacatgaa agcattcttt tgaaacaata ttttgttttc
     4561 tatttgattt cgcgttgaat agtcccaatg accctgatga gacaaaagca cttatcataa
     4621 aatttattat ctaagataat ctcgttaaaa aaaaaaaaaa aaaaaaaaaa aaaaaaaatt
     4681 tggcaagaag cccttctctt tcgttaaaac ttgtttcgct ttatgcaata ctgtaagtgt
     4741 aagcttgtaa tatagcgaat ctgtgaactc attgttcatt tcatggtagt tgttgatttc
     4801 acaagcatgc tgagttatca atatagtaac acaattggtt ttatttcagc tcgcttgaat
     4861 atagtgtcaa gagataattg aataaacaaa aaaaataact acacgttttt tagatacatt
     4921 ttatttacaa tgtattggtt ctttaaagaa tgggtggtaa ataaaaaaaa tattacaagg
     4981 tcgaaaaaaa ttcatccctt ttcattagtt ctataattaa atgctaaacc catgaatatt
     5041 cttttactgc taatcaatta acgattttgg tatttaaaag ctattgctga taagtattct
     5101 gacgttctgt atcaatatta ccatgcagct ggcggccgcg ttctatagtg tcacctaaat
     5161 cgtatgtgta tgatacataa ggttatgtat taattgtagc cgcgttctaa cgacaatatg
     5221 tccatatggt gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga
     5281 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
     5341 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
     5401 aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata
     5461 ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     5521 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     5581 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     5641 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     5701 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     5761 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     5821 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     5881 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     5941 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     6001 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     6061 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     6121 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     6181 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     6241 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     6301 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     6361 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     6421 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     6481 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     6541 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     6601 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     6661 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     6721 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     6781 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     6841 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     6901 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     6961 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     7021 ctacagcgtg agcattgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     7081 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     7141 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     7201 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     7261 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     7321 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     7381 cgcagcgagt cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc
     7441 gcg
//
