LOCUS       sequence-55584-e        6987 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 6987)
  AUTHORS   Houser JR, Ford E, Chatterjea SM, Maleri S, Elston TC, Errede B
  TITLE     An improved short-lived fluorescent protein transcriptional reporter
            for Saccharomyces cerevisiae.
  JOURNAL   Yeast. 2012 Dec;29(12):519-30. doi: 10.1002/yea.2932. Epub 2012 Nov 
            21.
  PUBMED    23172645
REFERENCE   2  (bases 1 to 6987)
  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: "10.1002/yea.2932"; 
            journalName: "Yeast"; date: "2012-12"; volume: "29"; issue: "12"; 
            pages: "519-30"
COMMENT     Sequence Label: pNC1136
FEATURES             Location/Qualifiers
     source          1..6987
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             449..676
                     /codon_start=1
                     /product="Saccharomyces cerevisiae ubiquitin"
                     /label=ubiquitin
                     /translation="MQIFVKTLTGKTITLEVESSDTIDNVKSKIQDKEGIPPDQQRLIF
                     AGKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
     CDS             749..1459
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKAN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(776..804)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1399..1419
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      1483..1670
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1538..1562)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1745..2945
                     /label=HIS3MX6
                     /note="yeast selectable marker encoding the S. pombe his5 
                     gene, which corresponds to S. cerevisiae HIS3"
     promoter        1745..2088
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             2089..2742
                     /codon_start=1
                     /gene="Schizosaccharomyces pombe his5"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=S. pombe his5
                     /note="yeast auxotrophic marker; corresponds to S. 
                     cerevisiae HIS3"
                     /translation="MGRRAFVERNTNETKISVAIALDKAPLPEESNFIDELITSKHTNQ
                     KGEQVIQVDTGIGFLDHMYHALAKHAGWSLRLYSRGDLIIDDHHTAEDTAIALGIAFKQ
                     AMSNFAGVKRFGHAYCPLDEALSRSVVDLSGRPYAVIDLGLKREKVGELSCEMIPHLLY
                     SFSVAAGITLHVTCLYGSNDHHRAESAFKSLAVAMRAATSLTGSSEVPSTKGVL"
     terminator      2748..2945
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(3469..3486)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3469..3485)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3478..3500)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3698..4153
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3785..3804)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3995..4016
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4170..4189)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4289..4311
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4349..4367)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4435..4539
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4540..4608,4609..5400)
                     /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(4758..4777)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5571..6159
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6060..6079
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6313..6330
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6447..6468
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6483..6488,6489..6506,6507..6513)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6521..6537
                     /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     6526..6548
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6545..6561
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6545..6561
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ccccctcgag tgatgaaaca aacatgaaac gtctgtaatt tgaaacaaat cgagcagatc
       61 tcgccaggcg tgtatatagc gtggatggcc aggcaacttt agtgctgaca catacaggca
      121 tatatatatg tgtgcgacga cacatgatca tatggcatgc atgtgctctg tatgtatata
      181 aaactcttgt tttcttcttt tctctaaata ttctttcctt atacattagg tcctttgtag
      241 cataaattac tatacttcta tagacacgca aacacaaata cacacactaa attaccggat
      301 caattcgcct gcttatcttt cttcttccga aagtgctact tcagaaagag caagaactgt
      361 acggataagg ataagtatat cttctatcta ctacaatacg aaaagaactc tcgaactctc
      421 cctcccactt tactttaact aatagattat gcagattttc gtcaagactt tgaccggtaa
      481 aaccataaca ttggaagttg aatcttccga taccatcgac aacgttaagt cgaaaattca
      541 agacaaggaa ggtatccctc cagatcaaca aagattgatc tttgccggta agcagctaga
      601 agacggtaga acgctgtctg attacaacat tcagaaggag tccaccttac atcttgtgct
      661 aaggctaaga ggtggttatc acggatccgg agcttggctg ttgcccgtct cactggtgaa
      721 aagaaaaacc accctggcgc ccaatacgag taaaggagaa gaacttttca ctggagttgt
      781 cccaattctt gttgaattag atggtgatgt taatgggcac aaattttctg tcagtggaga
      841 gggtgaaggt gatgcaacat acggaaaact tacccttaaa tttatttgca ctactggaaa
      901 actacctgtt ccatggccaa cacttgtcac tactctcact tatggtgttc aatgcttttc
      961 aagataccca gatcacatga aacagcatga ctttttcaag agtgccatgc ccgaaggtta
     1021 tgtacaggaa agaactatat ttttcaaaga tgacgggaac tacaagacac gtgctgaagt
     1081 caagtttgaa ggtgataccc ttgttaatag aatcgagtta aaaggtattg attttaaaga
     1141 agatggaaac attcttggac acaaattgga atacaactat aactcacaca atgtatacat
     1201 catggcagac aaacaaaaga atggaatcaa agctaacttc aaaattagac acaacattga
     1261 agatggaagc gttcaactag cagaccatta tcaacaaaat actccaattg gcgatggccc
     1321 tgtcctttta ccagacaacc attacctgtc cacacaatct gccctttcga aagatcccaa
     1381 cgaaaagaga gaccacatgg tccttcttga gtttgtaaca gctgctggga ttacacatgg
     1441 catggatgaa ctatacaaat agggcgcgcc acttctaaat aagcgaattt cttatgattt
     1501 atgattttta ttattaaata agttataaaa aaaataagtg tatacaaatt ttaaagtgac
     1561 tcttaggttt taaaacgaaa attcttattc ttgagtaact ctttcctgta ggtcaggttg
     1621 ctttctcagg tatagtatga ggtcgctctt attgaccaca cctctaccgg cagatccgct
     1681 agggataaca gggtaatata gatctgttta gcttgcctcg tccccgccgg gtcacccggc
     1741 cagcgacatg gaggcccaga ataccctcct tgacagtctt gacgtgcgca gctcaggggc
     1801 atgatgtgac tgtcgcccgt acatttagcc catacatccc catgtataat catttgcatc
     1861 catacatttt gatggccgca cggcgcgaag caaaaattac ggctcctcgc tgcagacctg
     1921 cgagcaggga aacgctcccc tcacagacgc gttgaattgt ccccacgccg cgcccctgta
     1981 gagaaatata aaaggttagg atttgccact gaggttcttc tttcatatac ttccttttaa
     2041 aatcttgcta ggatacagtt ctcacatcac atccgaacat aaacaaccat gggtaggagg
     2101 gcttttgtag aaagaaatac gaacgaaacg aaaatcagcg ttgccatcgc tttggacaaa
     2161 gctcccttac ctgaagagtc gaattttatt gatgaactta taacttccaa gcatacaaac
     2221 caaaagggag aacaagtaat ccaagtagac acgggaattg gattcttgga tcacatgtat
     2281 catgcactgg ctaaacatgc aggctggagc ttacgacttt actcaagagg tgatttaatc
     2341 atcgatgatc atcacactgc agaagatact gctattgcac ttggtattgc attcaagcag
     2401 gctatgagta actttgccgg cgttaaaaga tttggacatg cttattgtcc acttgacgaa
     2461 gctctttcta gaagcgtagt tgacttgtcg ggacggccct atgctgttat cgatttggga
     2521 ttaaagcgtg aaaaggttgg ggaattgtcc tgtgaaatga tccctcactt actatattcc
     2581 ttttcggtag cagctggaat tactttgcat gttacctgct tatatggtag taatgaccat
     2641 catcgtgctg aaagcgcttt taaatctctg gctgttgcca tgcgcgcggc tactagtctt
     2701 actggaagtt ctgaagtccc aagcacgaag ggagtgttgt aaagagtact gacaataaaa
     2761 agattcttgt tttcaagaac ttgtcatttg tatagttttt ttatattgta gttgttctat
     2821 tttaatcaaa tgttagcgtg atttatattt tttttcgcct cgacatcatc tgcccagatg
     2881 cgaagttaag tgcgcagaaa gtaatatcat gcgtcaatcg tatgtgaatg ctggtcgcta
     2941 tactgctgtc gattcgatac taacgctcga cctgcaggca tgaagatggg aatctcggtc
     3001 gtaatgattt ctataatgac gaaaaaaaaa aaattggaaa gaaaaagctt catggccttt
     3061 ataaaaagga actatccaat acctcgccag aaccaagtaa cagtatttta cggggcacaa
     3121 atcaagaaca ataagacagg actgtaaaga tggacgcatt gaactccaaa gaacaacaag
     3181 agttccaaaa agtagtggaa caaaagcaaa tgaaggattt catgcgtttg tactctaatc
     3241 tggtagaaag atgtttcaca gactgtgtca atgacttcac aacatcaaag ctaaccaata
     3301 aggaacaaac atgcatcatg aagtgctcag aaaagttctt gaagcatagc gaacgtgtag
     3361 ggcagcgttt ccaagaacaa aacgctgcct tgggacaagg cttgggccga taaggtgtac
     3421 tggcgtatat atatctaatt atgtatctct ggtgtcgaca agcttggcac tggccgtcgt
     3481 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     3541 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     3601 gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat tttctcctta cgcatctgtg
     3661 cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta
     3721 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg
     3781 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     3841 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     3901 aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     3961 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     4021 acactcaacc ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggcc
     4081 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     4141 acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     4201 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     4261 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     4321 tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     4381 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     4441 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     4501 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     4561 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     4621 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     4681 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     4741 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     4801 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     4861 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     4921 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     4981 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     5041 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     5101 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     5161 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     5221 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     5281 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     5341 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     5401 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     5461 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     5521 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     5581 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     5641 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     5701 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     5761 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     5821 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     5881 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     5941 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     6001 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     6061 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     6121 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     6181 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     6241 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     6301 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg
     6361 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
     6421 aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag
     6481 gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt
     6541 cacacaggaa acagctatga ccatgattac gaattcgagc tctggagaat atactaaggg
     6601 tactgttgac attgcgaaga gcgacaaaga ttttgttatc ggctttattg ctcaaagaga
     6661 catgggtgga agagatgaag gttacgattg gttgattatg acacccggtg tgggtttaga
     6721 tgacaaggga gacgcattgg gtcaacagta tagaaccgtg gatgatgtgg tctctacagg
     6781 atctgacatt attattgttg gaagaggact atttgcaaag ggaagggatg ctaaggtaga
     6841 gggtgaacgt tacagaaaag caggctggga agcatatttg agaagatgcg gccagcaaaa
     6901 ctaaaaaact gtattataag taaatgcatg tatactaaac tcacaaatta gagcttcaat
     6961 ttaattatat cagttattac ccggtac
//
