LOCUS       sequence-9609-ex        7601 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 7601)
  AUTHORS   Hackett EA, Esch RK, Maleri S, Errede B
  TITLE     A family of destabilized cyan fluorescent proteins as 
            transcriptional reporters in S. cerevisiae.
  JOURNAL   Yeast. 2006 Apr 15. 23(5):333-49.
  PUBMED    16598699
REFERENCE   2  (bases 1 to 7601)
  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. 2006
            Apr 15. 23(5):333-49."
COMMENT     Sequence Label: pNC951
FEATURES             Location/Qualifiers
     source          1..7601
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        866..1307
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    866..983
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     1278..1301
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             1710..1937
                     /codon_start=1
                     /product="Saccharomyces cerevisiae ubiquitin"
                     /label=ubiquitin
                     /translation="MQIFVKTLTGKTITLEVESSDTIDNVKSKIQDKEGIPPDQQRLIF
                     AGKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
     CDS             2010..2720
                     /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
                     ICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKAH
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     complement(2037..2065)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     2660..2680
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      2744..2931
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(2799..2823)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            3006..4206
                     /label=HIS3MX6
                     /note="yeast selectable marker encoding the S. pombe his5 
                     gene, which corresponds to S. cerevisiae HIS3"
     promoter        3006..3349
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             3350..4003
                     /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      4009..4206
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(4730..4747)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4730..4746)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4739..4761)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4959..5414
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5046..5065)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5256..5277
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5431..5450)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5550..5572
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5610..5628)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5696..5800
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5801..5869,5870..6661)
                     /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(6019..6038)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6832..7420
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7321..7340
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7574..7591
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gaattcgagc
      241 tctggagaat atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc
      301 ggctttattg ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg
      361 acacccggtg tgggtttaga tgacaaggga gacgcattgg gtcaacagta tagaaccgtg
      421 gatgatgtgg tctctacagg atctgacatt attattgttg gaagaggact atttgcaaag
      481 ggaagggatg ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg
      541 agaagatgcg gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac
      601 tcacaaatta gagcttcaat ttaattatat cagttattac ccggtacccc cctcgaggaa
      661 ttttcaaaaa ttcttacttt ttttttggat ggacgcaaag aagtttaata atcatattac
      721 atggcattac caccatatac atatccatat ctaatcttac ttatatgttg tggaaatgta
      781 aagagcccca ttatcttagc ctaaaaaaac cttctctttg gaactttcag taatacgctt
      841 aactgctcat tgctatattg aagtacggat tagaagccgc cgagcgggcg acagccctcc
      901 gacggaagac tctcctccgt gcgtcctcgt cttcaccggt cgcgttcctg aaacgcagat
      961 gtgcctcgcg ccgcactgct ccgaacaata aagattctac aatactagct tttatggtta
     1021 tgaagaggaa aaattggcag taacctggcc ccacaaacct tcaaattaac gaatcaaatt
     1081 aacaaccata ggatgataat gcgattagtt ttttagcctt atttctgggg taattaatca
     1141 gcgaagcgat gatttttgat ctattaacag atatataaat ggaaaagctg cataaccact
     1201 ttaactaata ctttcaacat tttcagtttg tattacttct tattcaaatg tcataaaagt
     1261 atcaacaaaa aattgttaat atacctctat actttaacgt caaggagaaa aaaccccgga
     1321 tctgccaggc gtgtatatag cgtggatggc caggcaactt tagtgctgac acatacaggc
     1381 atatatatat gtgtgcgacg acacatgatc atatggcatg catgtgctct gtatgtatat
     1441 aaaactcttg ttttcttctt ttctctaaat attctttcct tatacattag gtcctttgta
     1501 gcataaatta ctatacttct atagacacgc aaacacaaat acacacacta aattaccgga
     1561 tcaattcgcc tgcttatctt tcttcttccg aaagtgctac ttcagaaaga gcaagaactg
     1621 tacggataag gataagtata tcttctatct actacaatac gaaaagaact ctcgaactct
     1681 ccctcccact ttactttaac taatagatta tgcagatttt cgtcaagact ttgaccggta
     1741 aaaccataac attggaagtt gaatcttccg ataccatcga caacgttaag tcgaaaattc
     1801 aagacaagga aggtatccct ccagatcaac aaagattgat ctttgccggt aagcagctag
     1861 aagacggtag aacgctgtct gattacaaca ttcagaagga gtccacctta catcttgtgc
     1921 taaggctaag aggtggttat cacggatccg gagcttggct gttgcccgtc tcactggtga
     1981 aaagaaaaac caccctggcg cccaatacga gtaaaggaga agaacttttc actggagttg
     2041 tcccaattct tgttgaatta gatggtgatg ttaatgggca caaattttct gtcagtggag
     2101 agggtgaagg tgatgcaaca tacggaaaac ttacccttaa atttatttgc actactggaa
     2161 aactacctgt tccatggcca acacttgtca ctactttgac ttggggtgtt caatgctttt
     2221 caagataccc agatcatatg aaacggcatg actttttcaa gagtgccatg cccgaaggtt
     2281 atgtacagga aagaactata tttttcaaag atgacgggaa ctacaagaca cgtgctgaag
     2341 tcaagtttga aggtgatacc cttgttaata gaatcgagtt aaaaggtatt gattttaaag
     2401 aagatggaaa cattcttgga cacaaattgg aatacaacta tatttcacac aatgtataca
     2461 tcactgcaga caaacaaaag aatggaatca aagctcattt caaaattaga cacaacattg
     2521 aagatggaag cgttcaacta gcagaccatt atcaacaaaa tactccaatt ggcgatggcc
     2581 ctgtcctttt accagacaac cattacctgt ccacacaatc tgccctttcg aaagatccca
     2641 acgaaaagag agaccacatg gtccttcttg agtttgtaac agctgctggg attacacatg
     2701 gcatggatga actatacaaa tagggcgcgc cacttctaaa taagcgaatt tcttatgatt
     2761 tatgattttt attattaaat aagttataaa aaaaataagt gtatacaaat tttaaagtga
     2821 ctcttaggtt ttaaaacgaa aattcttatt cttgagtaac tctttcctgt aggtcaggtt
     2881 gctttctcag gtatagtatg aggtcgctct tattgaccac acctctaccg gcagatccgc
     2941 tagggataac agggtaatat agatctgttt agcttgcctc gtccccgccg ggtcacccgg
     3001 ccagcgacat ggaggcccag aataccctcc ttgacagtct tgacgtgcgc agctcagggg
     3061 catgatgtga ctgtcgcccg tacatttagc ccatacatcc ccatgtataa tcatttgcat
     3121 ccatacattt tgatggccgc acggcgcgaa gcaaaaatta cggctcctcg ctgcagacct
     3181 gcgagcaggg aaacgctccc ctcacagacg cgttgaattg tccccacgcc gcgcccctgt
     3241 agagaaatat aaaaggttag gatttgccac tgaggttctt ctttcatata cttcctttta
     3301 aaatcttgct aggatacagt tctcacatca catccgaaca taaacaacca tgggtaggag
     3361 ggcttttgta gaaagaaata cgaacgaaac gaaaatcagc gttgccatcg ctttggacaa
     3421 agctccctta cctgaagagt cgaattttat tgatgaactt ataacttcca agcatacaaa
     3481 ccaaaaggga gaacaagtaa tccaagtaga cacgggaatt ggattcttgg atcacatgta
     3541 tcatgcactg gctaaacatg caggctggag cttacgactt tactcaagag gtgatttaat
     3601 catcgatgat catcacactg cagaagatac tgctattgca cttggtattg cattcaagca
     3661 ggctatgagt aactttgccg gcgttaaaag atttggacat gcttattgtc cacttgacga
     3721 agctctttct agaagcgtag ttgacttgtc gggacggccc tatgctgtta tcgatttggg
     3781 attaaagcgt gaaaaggttg gggaattgtc ctgtgaaatg atccctcact tactatattc
     3841 cttttcggta gcagctggaa ttactttgca tgttacctgc ttatatggta gtaatgacca
     3901 tcatcgtgct gaaagcgctt ttaaatctct ggctgttgcc atgcgcgcgg ctactagtct
     3961 tactggaagt tctgaagtcc caagcacgaa gggagtgttg taaagagtac tgacaataaa
     4021 aagattcttg ttttcaagaa cttgtcattt gtatagtttt tttatattgt agttgttcta
     4081 ttttaatcaa atgttagcgt gatttatatt ttttttcgcc tcgacatcat ctgcccagat
     4141 gcgaagttaa gtgcgcagaa agtaatatca tgcgtcaatc gtatgtgaat gctggtcgct
     4201 atactgctgt cgattcgata ctaacgctcg acctgcaggc atgaagatgg gaatctcggt
     4261 cgtaatgatt tctataatga cgaaaaaaaa aaaattggaa agaaaaagct tcatggcctt
     4321 tataaaaagg aactatccaa tacctcgcca gaaccaagta acagtatttt acggggcaca
     4381 aatcaagaac aataagacag gactgtaaag atggacgcat tgaactccaa agaacaacaa
     4441 gagttccaaa aagtagtgga acaaaagcaa atgaaggatt tcatgcgttt gtactctaat
     4501 ctggtagaaa gatgtttcac agactgtgtc aatgacttca caacatcaaa gctaaccaat
     4561 aaggaacaaa catgcatcat gaagtgctca gaaaagttct tgaagcatag cgaacgtgta
     4621 gggcagcgtt tccaagaaca aaacgctgcc ttgggacaag gcttgggccg ataaggtgta
     4681 ctggcgtata tatatctaat tatgtatctc tggtgtcgac aagcttggca ctggccgtcg
     4741 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     4801 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     4861 agttgcgcag cctgaatggc gaatggcgcc tgatgcggta ttttctcctt acgcatctgt
     4921 gcggtatttc acaccgcata cgtcaaagca accatagtac gcgccctgta gcggcgcatt
     4981 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     5041 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     5101 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     5161 caaaaaactt gatttgggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     5221 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     5281 aacactcaac cctatctcgg gctattcttt tgatttataa gggattttgc cgatttcggc
     5341 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     5401 aacgtttaca attttatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     5461 ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     5521 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     5581 gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa
     5641 tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg
     5701 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     5761 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     5821 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     5881 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     5941 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     6001 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     6061 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     6121 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     6181 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     6241 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     6301 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     6361 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     6421 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     6481 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     6541 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     6601 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     6661 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     6721 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     6781 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     6841 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     6901 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     6961 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     7021 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     7081 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     7141 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     7201 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     7261 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     7321 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     7381 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     7441 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
     7501 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
     7561 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa g
//
