LOCUS       sequence-9614-ex        6309 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 6309)
  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 6309)
  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: pNC1053
FEATURES             Location/Qualifiers
     source          1..6309
                     /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"
     CDS             718..1428
                     /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(745..773)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1368..1388
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     terminator      1452..1639
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(1507..1531)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     gene            1714..2914
                     /label=HIS3MX6
                     /note="yeast selectable marker encoding the S. pombe his5 
                     gene, which corresponds to S. cerevisiae HIS3"
     promoter        1714..2057
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             2058..2711
                     /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      2717..2914
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(3438..3455)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3438..3454)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3447..3469)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3667..4122
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3754..3773)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3964..3985
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4139..4158)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4258..4280
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4318..4336)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4404..4508
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4509..4577,4578..5369)
                     /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(4727..4746)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5540..6128
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6029..6048
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6282..6299
                     /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 ccggtacccg gggatccgga
      661 gcttggctgt tgcccgtctc actggtgaaa agaaaaacca ccctggcgcc caatacgagt
      721 aaaggagaag aacttttcac tggagttgtc ccaattcttg ttgaattaga tggtgatgtt
      781 aatgggcaca aattttctgt cagtggagag ggtgaaggtg atgcaacata cggaaaactt
      841 acccttaaat ttatttgcac tactggaaaa ctacctgttc catggccaac acttgtcact
      901 actttgactt ggggtgttca atgcttttca agatacccag atcatatgaa acggcatgac
      961 tttttcaaga gtgccatgcc cgaaggttat gtacaggaaa gaactatatt tttcaaagat
     1021 gacgggaact acaagacacg tgctgaagtc aagtttgaag gtgataccct tgttaataga
     1081 atcgagttaa aaggtattga ttttaaagaa gatggaaaca ttcttggaca caaattggaa
     1141 tacaactata tttcacacaa tgtatacatc actgcagaca aacaaaagaa tggaatcaaa
     1201 gctcatttca aaattagaca caacattgaa gatggaagcg ttcaactagc agaccattat
     1261 caacaaaata ctccaattgg cgatggccct gtccttttac cagacaacca ttacctgtcc
     1321 acacaatctg ccctttcgaa agatcccaac gaaaagagag accacatggt ccttcttgag
     1381 tttgtaacag ctgctgggat tacacatggc atggatgaac tatacaaata gggcgcgcca
     1441 cttctaaata agcgaatttc ttatgattta tgatttttat tattaaataa gttataaaaa
     1501 aaataagtgt atacaaattt taaagtgact cttaggtttt aaaacgaaaa ttcttattct
     1561 tgagtaactc tttcctgtag gtcaggttgc tttctcaggt atagtatgag gtcgctctta
     1621 ttgaccacac ctctaccggc agatccgcta gggataacag ggtaatatag atctgtttag
     1681 cttgcctcgt ccccgccggg tcacccggcc agcgacatgg aggcccagaa taccctcctt
     1741 gacagtcttg acgtgcgcag ctcaggggca tgatgtgact gtcgcccgta catttagccc
     1801 atacatcccc atgtataatc atttgcatcc atacattttg atggccgcac ggcgcgaagc
     1861 aaaaattacg gctcctcgct gcagacctgc gagcagggaa acgctcccct cacagacgcg
     1921 ttgaattgtc cccacgccgc gcccctgtag agaaatataa aaggttagga tttgccactg
     1981 aggttcttct ttcatatact tccttttaaa atcttgctag gatacagttc tcacatcaca
     2041 tccgaacata aacaaccatg ggtaggaggg cttttgtaga aagaaatacg aacgaaacga
     2101 aaatcagcgt tgccatcgct ttggacaaag ctcccttacc tgaagagtcg aattttattg
     2161 atgaacttat aacttccaag catacaaacc aaaagggaga acaagtaatc caagtagaca
     2221 cgggaattgg attcttggat cacatgtatc atgcactggc taaacatgca ggctggagct
     2281 tacgacttta ctcaagaggt gatttaatca tcgatgatca tcacactgca gaagatactg
     2341 ctattgcact tggtattgca ttcaagcagg ctatgagtaa ctttgccggc gttaaaagat
     2401 ttggacatgc ttattgtcca cttgacgaag ctctttctag aagcgtagtt gacttgtcgg
     2461 gacggcccta tgctgttatc gatttgggat taaagcgtga aaaggttggg gaattgtcct
     2521 gtgaaatgat ccctcactta ctatattcct tttcggtagc agctggaatt actttgcatg
     2581 ttacctgctt atatggtagt aatgaccatc atcgtgctga aagcgctttt aaatctctgg
     2641 ctgttgccat gcgcgcggct actagtctta ctggaagttc tgaagtccca agcacgaagg
     2701 gagtgttgta aagagtactg acaataaaaa gattcttgtt ttcaagaact tgtcatttgt
     2761 atagtttttt tatattgtag ttgttctatt ttaatcaaat gttagcgtga tttatatttt
     2821 ttttcgcctc gacatcatct gcccagatgc gaagttaagt gcgcagaaag taatatcatg
     2881 cgtcaatcgt atgtgaatgc tggtcgctat actgctgtcg attcgatact aacgctcgac
     2941 ctgcaggcat gaagatggga atctcggtcg taatgatttc tataatgacg aaaaaaaaaa
     3001 aattggaaag aaaaagcttc atggccttta taaaaaggaa ctatccaata cctcgccaga
     3061 accaagtaac agtattttac ggggcacaaa tcaagaacaa taagacagga ctgtaaagat
     3121 ggacgcattg aactccaaag aacaacaaga gttccaaaaa gtagtggaac aaaagcaaat
     3181 gaaggatttc atgcgtttgt actctaatct ggtagaaaga tgtttcacag actgtgtcaa
     3241 tgacttcaca acatcaaagc taaccaataa ggaacaaaca tgcatcatga agtgctcaga
     3301 aaagttcttg aagcatagcg aacgtgtagg gcagcgtttc caagaacaaa acgctgcctt
     3361 gggacaaggc ttgggccgat aaggtgtact ggcgtatata tatctaatta tgtatctctg
     3421 gtgtcgacaa gcttggcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc
     3481 gttacccaac ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa
     3541 gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggcgcctg
     3601 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac
     3661 catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     3721 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     3781 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     3841 gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta
     3901 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     3961 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcgggc tattcttttg
     4021 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     4081 aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta
     4141 caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg
     4201 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     4261 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc
     4321 tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag
     4381 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     4441 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     4501 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     4561 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     4621 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     4681 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     4741 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     4801 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     4861 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     4921 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     4981 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     5041 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     5101 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     5161 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     5221 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     5281 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     5341 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     5401 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     5461 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     5521 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     5581 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     5641 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
     5701 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     5761 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     5821 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     5881 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     5941 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     6001 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     6061 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     6121 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     6181 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
     6241 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
     6301 aagcggaag
//
