LOCUS       sequence-56725-e        7528 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 7528)
  AUTHORS   Chu D, Kazana E, Bellanger N, Singh T, Tuite MF, von der Haar T
  TITLE     Translation elongation can control translation initiation on 
            eukaryotic mRNAs.
  JOURNAL   EMBO J. 2014 Jan 1;33(1):21-34. doi: 10.1002/embj.201385651. Epub 
            2013 Dec 19.
  PUBMED    24357599
REFERENCE   2  (bases 1 to 7528)
  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/embj.201385651"; journalName: "EMBO J"; date: "2014-01-1- 
            1"; volume: "33"; issue: "1"; pages: "21-34"
COMMENT     Sequence Label: pTH730-staRLuc
FEATURES             Location/Qualifiers
     source          1..7528
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    70..573
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        610..714
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(715..783,784..1575)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(933..952)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1746..2334
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2488..2505
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2622..2643
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2658..2663,2664..2681,2682..2688)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2696..2712
                     /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     2701..2723
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2720..2736
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2720..2736
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2757..2775
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     terminator      complement(2823..3122)
                     /label=ADH2 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 2 (ADH2) gene"
     promoter        3564..4208
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     4101..4124
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             4216..5148
                     /codon_start=1
                     /product="luciferase from the anthozoan coelenterate 
                     Renilla reniformis (sea pansy)"
                     /label=Rluc
                     /translation="MTSKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     terminator      5295..5482
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(5350..5374)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     primer_bind     complement(5544..5563)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5545..5563)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5570..5587)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5570..5586)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5579..5601)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5731..6186
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5818..5837)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6028..6049
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(6315..7118)
                     /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
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(7119..7339)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(7359..7378)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7478..7500
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttaggacgg atcgcttgcc tgtaacttac acgcgcctcg tatcttttaa tgatggaata
      121 atttgggaat ttactctgtg tttatttatt tttatgtttt gtatttggat tttagaaagt
      181 aaataaagaa ggtagaagag ttacggaatg aagaaaaaaa aataaacaaa ggtttaaaaa
      241 atttcaacaa aaagcgtact ttacatatat atttattaga caagaaaagc agattaaata
      301 gatatacatt cgattaacga taagtaaaat gtaaaatcac aggattttcg tgtgtggtct
      361 tctacacaga caagatgaaa caattcggca ttaatacctg agagcaggaa gagcaagata
      421 aaaggtagta tttgttggcg atccccctag agtcttttac atcttcggaa aacaaaaact
      481 attttttctt taatttcttt ttttactttc tatttttaat ttatatattt atattaaaaa
      541 atttaaatta taattatttt tatagcacgt gatgaaaagg acccaggtgg cacttttcgg
      601 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
      661 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
      721 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
      781 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
      841 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
      901 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
      961 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     1021 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     1081 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     1141 ccgaaggagc taaccgcttt ttttcacaac atgggggatc atgtaactcg ccttgatcgt
     1201 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     1261 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     1321 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     1381 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     1441 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     1501 ggcagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     1561 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     1621 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     1681 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     1741 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     1801 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     1861 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     1921 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     1981 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     2041 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     2101 acgacctaca ccgaactgag atacctacag cgtgagcatt gagaaagcgc cacgcttccc
     2161 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     2221 agggagcttc caggggggaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     2281 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc cgagcctatg gaaaaacgcc
     2341 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     2401 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     2461 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     2521 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
     2581 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact
     2641 cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg tggaattgtg
     2701 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gctcgaaatt
     2761 aaccctcact aaagggaaca aaagctggta ccgggccccc cctcgagggc atgcgaagga
     2821 aaatgagaaa tatcgaggga gacgattcag aggagcagga caaactataa ccgactgttt
     2881 gttggaggat gccgtacata acgaacactg ctgaagctac catgtctaca gtttagagga
     2941 atgggtacaa ctcacaggcg agggatggtg ttcactcgtg ctagcaaacg cggtgggagc
     3001 aaaaagtaga atattatctt ttattcgtga aacttcgaac actgtcatct aaagatgcta
     3061 tatactaata taggcatact tgataatgaa aactataaat cgtaaagaca taagagatcc
     3121 gcttatttag agtgtcgaat tcgagctcgg tacccgggga gttgattgta gcttggtata
     3181 gcttgaaata ttgtgcagaa aaagaaacaa ggaagaaagg gaacgagaac aatgacgagg
     3241 aaacaaaaga ttaataattg caggtctatt tatacttgat agcaagacag caaacttttt
     3301 ttatttcaaa ttcaagtaac tggaaggaag gccgtatacc gttgctcatt agagagtagt
     3361 gtgcgtgaat gaaggaagga aaaagtttcg tgtgttcgaa gatacccctc atcagctctg
     3421 gaacaacgac atctgttggt gctgtctttg tcgttaattt tttcctttag tgtcttccat
     3481 catttttttt gtcattgcgg atatggtgag acaacaacgg gggagagaga aaagaaaaaa
     3541 aaagaaaaga atccgggagt ttatcattat caatactgcc atttcaaaga atacgtaaat
     3601 aattaatagt agtgattttc ctaactttat ttagtcaaaa aattagcctt ttaattctgc
     3661 tgtaacccgt acatgcccaa aatagggggc gggttacaca gaatatataa catcgtaggt
     3721 gtctgggtga acagtttatt cctggcatcc actaaatata atggagcccg ctttttaagc
     3781 tggcatccag aaaaaaaaag aatcccagca ccaaaatatt gttttcttca ccaaccatca
     3841 gttcataggt ccattctctt agcgcaacta cagagaacag gggcacaaac aggcaaaaaa
     3901 cgggcacaac ctcaatggag tgatgcaacc tgcctggagt aaatgatgac acaaggcaat
     3961 tgacccacgc atgtatctat ctcattttct tacaccttct attaccttct gctctctctg
     4021 atttggaaaa agctgaaaaa aaaggttgaa accagttccc tgaaattatt cccctacttg
     4081 actaataagt atataaagaa cggtaggtat tgattgtaat tctgtaaatc tatttcttaa
     4141 acttcttaaa ttctactttt atagttagtc ttttttttag ttttaaaaca ccaagaactt
     4201 agtttcgacg gatccatgac ttcgaaagtt tatgatccag aacaaaggaa acggatgata
     4261 actggtccgc agtggtgggc cagatgtaaa caaatgaatg ttcttgattc atttattaat
     4321 tattatgatt cagaaaaaca tgcagaaaat gctgttattt ttttacatgg taacgcggcc
     4381 tcttcttatt tatggcgaca tgttgtgcca catattgagc cagtagcgcg gtgtattata
     4441 ccagacctta ttggtatggg caaatcaggc aaatctggta atggttctta taggttactt
     4501 gatcattaca aatatcttac tgcatggttt gaacttctta atttaccaaa gaagatcatt
     4561 tttgtcggcc atgattgggg tgcttgtttg gcatttcatt atagctatga gcatcaagat
     4621 aagatcaaag caatagttca cgctgaaagt gtagtagatg tgattgaatc atgggatgaa
     4681 tggcctgata ttgaagaaga tattgcgttg atcaaatctg aagaaggaga aaaaatggtt
     4741 ttggagaata acttcttcgt ggaaaccatg ttgccatcaa aaatcatgag aaagttagaa
     4801 ccagaagaat ttgcagcata tcttgaacca ttcaaagaga aaggtgaagt tcgtcgtcca
     4861 acattatcat ggcctcgtga aatcccgtta gtaaaaggtg gtaaacctga cgttgtacaa
     4921 attgttagga attataatgc ttatctacgt gcaagtgatg atttaccaaa aatgtttatt
     4981 gaatcggacc caggattctt ttccaatgct attgttgaag gtgccaagaa gtttcctaat
     5041 actgaatttg tcaaagtaaa aggtcttcat ttttcgcaag aagatgcacc tgatgaaatg
     5101 ggaaaatata tcaaatcgtt cgttgagcga gttctcaaaa atgaacaaat gtcgtaaaag
     5161 ctttggactt cttcgccaga ggtttggtca agtctccaat caaggttgtc ggcttgtcta
     5221 ccttgccaga aatttacgaa aagatggaaa agggtcaaat cgttggtaga tacgttgttg
     5281 acacttctaa ataagcgaat ttcttatgat ttatgatttt tattattaaa taagttataa
     5341 aaaaaataag tgtatacaaa ttttaaagtg actcttaggt tttaaaacga aaattcttgt
     5401 tcttgagtaa ctctttcctg taggtcaggt tgctttctca ggtatagcat gaggtcgctc
     5461 ttattgacca cacctctacc ggcatgcccg aaattcccct accctatgaa aagcggccgc
     5521 caccgcggtg gagctccaat tcgccctata gtgagtcgta ttacaattca ctggccgtcg
     5581 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     5641 atcccccctt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     5701 agttgcgcag cctgaatggc gaatggcgcg acgcgccctg tagcggcgca ttaagcgcgg
     5761 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     5821 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     5881 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     5941 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     6001 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     6061 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     6121 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta
     6181 caatttcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcagggt
     6241 aataactgat ataattaaat tgaagctcta atttgtgagt ttagtataca tgcatttact
     6301 tataatacag ttttttagtt ttgctggccg catcttctca aatatgcttc ccagcctgct
     6361 tttctgtaac gttcaccctc taccttagca tcccttccct ttgcaaatag tcctcttcca
     6421 acaataataa tgtcagatcc tgtagagacc acatcatcca cggttctata ctgttgaccc
     6481 aatgcgtctc ccttgtcatc taaacccaca ccgggtgtca taatcaacca atcgtaacct
     6541 tcatctcttc cacccatgtc tctttgagca ataaagccga taacaaaatc tttgtcgctc
     6601 ttcgcaatgt caacagtacc cttagtatat tctccagtag atagggagcc cttgcatgac
     6661 aattctgcta acatcaaaag gcctctaggt tcctttgtta cttcttctgc cgcctgcttc
     6721 aaaccgctaa caatacctgg gcccaccaca ccgtgtgcat tcgtaatgtc tgcccattct
     6781 gctattctgt atacacccgc agagtactgc aatttgactg tattaccaat gtcagcaaat
     6841 tttctgtctt cgaagagtaa aaaattgtac ttggcggata atgcctttag cggcttaact
     6901 gtgccctcca tggaaaaatc agtcaagata tccacatgtg tttttagtaa acaaattttg
     6961 ggacctaatg cttcaactaa ctccagtaat tccttggtgg tacgaacatc caatgaagca
     7021 cacaagtttg tttgcttttc gtgcatgata ttaaatagct tggcagcaac aggactagga
     7081 tgagtagcag cacgttcctt atatgtagct ttcgacatga tttatcttcg tttcctgcag
     7141 gtttttgttc tgtgcagttg ggttaagaat actgggcaat ttcatgtttc ttcaacacta
     7201 catatgcgta tatataccaa tctaagtctg tgctccttcc ttcgttcttc cttctgttcg
     7261 gagattaccg aatcaaaaaa atttcaaaga aaccgaaatc aaaaaaaaga ataaaaaaaa
     7321 aatgatgaat tgaattgaaa agcgtggtgc actctcagta caatctgctc tgatgccgca
     7381 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     7441 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     7501 ttttcaccgt catcaccgaa acgcgcga
//
