LOCUS       sequence-204983-        6658 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 6658)
  AUTHORS   Merksamer PI, Trusina A, Papa FR
  TITLE     Real-time redox measurements during endoplasmic reticulum stress 
            reveal interlinked protein folding functions.
  JOURNAL   Cell. 2008 Nov 28. 135(5):933-47.
  PUBMED    19026441
REFERENCE   2  (bases 1 to 6658)
  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: "Cell. 2008 
            Nov 28. 135(5):933-47."
COMMENT     Sequence Label: pPM28 (eroGFP CEN/ARS URA3)
FEATURES             Location/Qualifiers
     source          1..6658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(13..30)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(184..772)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(264..283)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(943..1734,1735..1803))
                     /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     1566..1585
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1804..1908)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    1945..2448
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     2489..2507
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2545..2567)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2667..2686
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        2710..2925
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             2926..3729
                     /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"
     rep_origin      complement(3860..4315)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3997..4018)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4209..4228
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4441..4463
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     4455..4472
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     4456..4472
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4479..4498
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4479..4497
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        4526..5168
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     5062..5085
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             join(5331..5333,5334..5336,5337..6047)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFISTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNCHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTCSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5376..5397)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5637..5656)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5984..6005
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      6100..6347
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(6116..6134)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(6356..6372)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6357..6373)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(6401..6419)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(6440..6456)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6440..6456)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6453..6475)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6464..6480
                     /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)."
     promoter        complement(join(6488..6494,6495..6512,6513..6518))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6533..6554
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
       61 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
      121 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
      181 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
      241 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
      301 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
      361 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
      421 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
      481 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
      541 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
      601 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
      661 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
      721 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
      781 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
      841 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
      901 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
      961 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     1021 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     1081 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     1141 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     1201 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     1261 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     1321 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     1381 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     1441 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     1501 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     1561 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     1621 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     1681 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     1741 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     1801 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     1861 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     1921 aaaagtgcca cctgggtcct tttcatcacg tgctataaaa ataattataa tttaaatttt
     1981 ttaatataaa tatataaatt aaaaatagaa agtaaaaaaa gaaattaaag aaaaaatagt
     2041 ttttgttttc cgaagatgta aaagactcta gggggatcgc caacaaatac taccttttat
     2101 cttgctcttc ctgctctcag gtattaatgc cgaattgttt catcttgtct gtgtagaaga
     2161 ccacacacga aaatcctgtg attttacatt ttacttatcg ttaatcgaat gtatatctat
     2221 ttaatctgct tttcttgtct aataaatata tatgtaaagt acgctttttg ttgaaatttt
     2281 ttaaaccttt gtttattttt ttttcttcat tccgtaactc ttctaccttc tttatttact
     2341 ttctaaaatc caaatacaaa acataaaaat aaataaacac agagtaaatt cccaaattat
     2401 tccatcatta aaagatacga ggcgcgtgta agttacaggc aagcgatccg tctaagaaac
     2461 cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc
     2521 gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga cggtcacagc
     2581 ttgtctgtaa gcggatgccg ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg
     2641 cgggtgtcgg ggctggctta actatgcggc atcagagcag attgtactga gagtgcacca
     2701 taccaccttt tcaattcatc attttttttt tattcttttt tttgatttcg gtttccttga
     2761 aatttttttg attcggtaat ctccgaacag aaggaagaac gaaggaagga gcacagactt
     2821 agattggtat atatacgcat atgtagtgtt gaagaaacat gaaattgccc agtattctta
     2881 acccaactgc acagaacaaa aacctgcagg aaacgaagat aaatcatgtc gaaagctaca
     2941 tataaggaac gtgctgctac tcatcctagt cctgttgctg ccaagctatt taatatcatg
     3001 cacgaaaagc aaacaaactt gtgtgcttca ttggatgttc gtaccaccaa ggaattactg
     3061 gagttagttg aagcattagg tcccaaaatt tgtttactaa aaacacatgt ggatatcttg
     3121 actgattttt ccatggaggg cacagttaag ccgctaaagg cattatccgc caagtacaat
     3181 tttttactct tcgaagacag aaaatttgct gacattggta atacagtcaa attgcagtac
     3241 tctgcgggtg tatacagaat agcagaatgg gcagacatta cgaatgcaca cggtgtggtg
     3301 ggcccaggta ttgttagcgg tttgaagcag gcggcagaag aagtaacaaa ggaacctaga
     3361 ggccttttga tgttagcaga attgtcatgc aagggctccc tatctactgg agaatatact
     3421 aagggtactg ttgacattgc gaagagcgac aaagattttg ttatcggctt tattgctcaa
     3481 agagacatgg gtggaagaga tgaaggttac gattggttga ttatgacacc cggtgtgggt
     3541 ttagatgaca agggagacgc attgggtcaa cagtatagaa ccgtggatga tgtggtctct
     3601 acaggatctg acattattat tgttggaaga ggactatttg caaagggaag ggatgctaag
     3661 gtagagggtg aacgttacag aaaagcaggc tgggaagcat atttgagaag atgcggccag
     3721 caaaactaaa aaactgtatt ataagtaaat gcatgtatac taaactcaca aattagagct
     3781 tcaatttaat tatatcagtt attaccctat gcggtgtgaa ataccgcaca gatgcgtaag
     3841 gagaaaatac cgcatcagga aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     3901 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     3961 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     4021 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     4081 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     4141 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     4201 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     4261 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtccatt
     4321 cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct tcgctattac
     4381 gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt
     4441 cccagtcacg acgttgtaaa acgacggcca gtgaattgta atacgactca ctatagggcg
     4501 aattggagct ccaccgcgga gtttatcatt atcaatactg ccatttcaaa gaatacgtaa
     4561 ataattaata gtagtgattt tcctaacttt atttagtcaa aaaattagcc ttttaattct
     4621 gctgtaaccc gtacatgccc aaaatagggg gcgggttaca cagaatatat aacatcgtag
     4681 gtgtctgggt gaacagttta ttcctggcat ccactaaata taatggagcc cgctttttaa
     4741 gctggcatcc agaaaaaaaa agaatcccag caccaaaata ttgttttctt caccaaccat
     4801 cagttcatag gtccattctc ttagcgcaac tacagagaac aggggcacaa acaggcaaaa
     4861 aacgggcaca acctcaatgg agtgatgcaa cctgcctgga gtaaatgatg acacaaggca
     4921 attgacccac gcatgtatct atctcatttt cttacacctt ctattacctt ctgctctctc
     4981 tgatttggaa aaagctgaaa aaaaaggttg aaaccagttc cctgaaatta ttcccctact
     5041 tgactaataa gtatataaag acggtaggta ttgattgtaa ttctgtaaat ctatttctta
     5101 aacttcttaa attctacttt tatagttagt ctttttttta gttttaaaac accagaactt
     5161 agtttcgagc tagcatgttt ttcaacagac taagcgctgg caagctgctg gtaccactct
     5221 ccgtggtcct gtacgccctt ttcgtggtaa tattaccttt acagaattct ttccactcct
     5281 ccaatgtttt agttagaggt gccgatgatg gggatccact agtagcaaca atggtgagta
     5341 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa
     5401 acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga
     5461 ccctgaagtt catctccacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
     5521 ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact
     5581 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
     5641 acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
     5701 tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
     5761 acaactacaa ctgccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg
     5821 tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc
     5881 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca
     5941 cctgctccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
     6001 tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagact cacggtatgg
     6061 acgaattgta caagcacgac gaattgtaat ctagagtcat catgtaatta gttatgtcac
     6121 gcttacattc acgccctccc cccacatccg ctctaaccga aaaggaagga gttagacaac
     6181 ctgaagtcta ggtccctatt tattttttta tagttatgtt agtattaaga acgttattta
     6241 tatttcaaat ttttcttttt tttctgtaca gacgcgtgta cgcatgtaac attatactga
     6301 aaaccttgct tgagaaggtt ttgggacgct cgaaggcttt aatttgcaag cttatcgata
     6361 ccgtcgacct cgaggggggg cccggtacca gcttttgttc cctttagtga gggttaattt
     6421 cgagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     6481 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     6541 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     6601 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcg
//
