LOCUS       sequence-9482-ex        6663 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 6663)
  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 6663)
  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..6663
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        190..410
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             411..1214
                     /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(1343..1798)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1480..1501)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1692..1711
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1928..1950
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1942..1959
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1943..1959
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1966..1985
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1966..1984
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2013..2655
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     2549..2572
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     CDS             join(2818..2820,2821..2823,2824..3534)
                     /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(2863..2884)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3124..3143)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3471..3492
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      3587..3834
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3603..3621)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(3843..3859)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3844..3860)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(3889..3907)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3928..3944)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3928..3944)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3941..3963)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3952..3968
                     /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(3976..3982,3983..4000,4001..4006))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4159..4176)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4330..4918)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5089..5880,5881..5949))
                     /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     5712..5731
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5950..6054)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    6091..6594
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     6636..6654
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accacgcttt tcaattcaat tcatcatttt ttttttattc ttttttttga tttcggtttc
      241 tttgaaattt ttttgattcg gtaatctccg aacagaagga agaacgaagg aaggagcaca
      301 gacttagatt ggtatatata cgcatatgta gtgttgaaga aacatgaaat tgcccagtat
      361 tcttaaccca actgcacaga acaaaaacct gcaggaaacg aagataaatc atgtcgaaag
      421 ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag ctatttaata
      481 tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc accaaggaat
      541 tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca catgtggata
      601 tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta tccgccaagt
      661 acaatttttt actcttcgaa gacagaaaat ttgctgacat tggtaataca gtcaaattgc
      721 agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat gcacacggtg
      781 tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta acaaaggaac
      841 ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct actggagaat
      901 atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc ggctttattg
      961 ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg acacccggtg
     1021 tgggtttaga tgacaaggga gacgcattgg gtcaacagta tagaaccgtg gatgatgtgg
     1081 tctctacagg atctgacatt attattgttg gaagaggact atttgcaaag ggaagggatg
     1141 ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg agaagatgcg
     1201 gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac tcacaaatta
     1261 gagcttcaat ttaattatat cagttattac cctgcggtgt gaaataccgc acagatgcgt
     1321 aaggagaaaa taccgcatca ggaaattgta aacgttaata ttttgttaaa attcgcgtta
     1381 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     1441 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     1501 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     1561 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     1621 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     1681 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     1741 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcg
     1801 cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg
     1861 ctattacgcc agctggcgaa ggggggatgt gctgcaaggc gattaagttg ggtaacgcca
     1921 gggttttccc agtcacgacg ttgtaaaacg acggccagtg aattgtaata cgactcacta
     1981 tagggcgaat tggagctcca ccgcggagtt tatcattatc aatactgcca tttcaaagaa
     2041 tacgtaaata attaatagta gtgattttcc taactttatt tagtcaaaaa attagccttt
     2101 taattctgct gtaacccgta catgcccaaa atagggggcg ggttacacag aatatataac
     2161 atcgtaggtg tctgggtgaa cagtttattc ctggcatcca ctaaatataa tggagcccgc
     2221 tttttaagct ggcatccaga aaaaaaaaga atcccagcac caaaatattg ttttcttcac
     2281 caaccatcag ttcataggtc cattctctta gcgcaactac agagaacagg ggcacaaaca
     2341 ggcaaaaaac gggcacaacc tcaatggagt gatgcaacct gcctggagta aatgatgaca
     2401 caaggcaatt gacccacgca tgtatctatc tcattttctt acaccttcta ttaccttctg
     2461 ctctctctga tttggaaaaa gctgaaaaaa aaggttgaaa ccagttccct gaaattattc
     2521 ccctacttga ctaataagta tataaagacg gtaggtattg attgtaattc tgtaaatcta
     2581 tttcttaaac ttcttaaatt ctacttttat agttagtctt ttttttagtt ttaaaacacc
     2641 agaacttagt ttcgagctag catgtttttc aacagactaa gcgctggcaa gctgctggta
     2701 ccactctccg tggtcctgta cgcccttttc gtggtaatat tacctttaca gaattctttc
     2761 cactcctcca atgttttagt tagaggtgcc gatgatgggg atccactagt agcaacaatg
     2821 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     2881 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     2941 aagctgaccc tgaagttcat ctccaccacc ggcaagctgc ccgtgccctg gcccaccctc
     3001 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     3061 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     3121 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     3181 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     3241 ctggagtaca actacaactg ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     3301 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     3361 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     3421 ctgagcacct gctccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     3481 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagactcac
     3541 ggtatggacg aattgtacaa gcacgacgaa ttgtaatcta gagtcatcat gtaattagtt
     3601 atgtcacgct tacattcacg ccctcccccc acatccgctc taaccgaaaa ggaaggagtt
     3661 agacaacctg aagtctaggt ccctatttat ttttttatag ttatgttagt attaagaacg
     3721 ttatttatat ttcaaatttt tctttttttt ctgtacagac gcgtgtacgc atgtaacatt
     3781 atactgaaaa ccttgcttga gaaggttttg ggacgctcga aggctttaat ttgcaagctt
     3841 atcgataccg tcgacctcga gggggggccc ggtacccagc ttttgttccc tttagtgagg
     3901 gttaattccg agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     3961 gctcacaatt ccacacaaca taggagccgg aagcataaag tgtaaagcct ggggtgccta
     4021 atgagtgagg taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     4081 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     4141 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     4201 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     4261 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     4321 gctggcgttt ttccataggc tcggcccccc tgacgagcat cacaaaaatc gacgctcaag
     4381 tcagaggtgg cgaaacccga caggactata aagataccag gcgttccccc ctggaagctc
     4441 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     4501 ttcgggaagc gtggcgcttt ctcaatgctc acgctgtagg tatctcagtt cggtgtaggt
     4561 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     4621 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     4681 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     4741 gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa
     4801 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     4861 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     4921 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     4981 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     5041 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     5101 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     5161 gcccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     5221 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     5281 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     5341 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     5401 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     5461 ccaacgatca aggcgagtta catgatcccc catgttgtga aaaaaagcgg ttagctcctt
     5521 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     5581 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     5641 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     5701 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     5761 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     5821 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     5881 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     5941 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     6001 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     6061 tccccgaaaa gtgccacctg ggtccttttc atcacgtgct ataaaaataa ttataattta
     6121 aattttttaa tataaatata taaattaaaa atagaaagta aaaaaagaaa ttaaagaaaa
     6181 aatagttttt gttttccgaa gatgtaaaag actctagggg gatcgccaac aaatactacc
     6241 ttttatcttg ctcttcctgc tctcaggtat taatgccgaa ttgtttcatc ttgtctgtgt
     6301 agaagaccac acacgaaaat cctgtgattt tacattttac ttatcgttaa tcgaatgtat
     6361 atctatttaa tctgcttttc ttgtctaata aatatatatg taaagtacgc tttttgttga
     6421 aattttttaa acctttgttt attttttttt cttcattccg taactcttct accttcttta
     6481 tttactttct aaaatccaaa tacaaaacat aaaaataaat aaacacagag taaattccca
     6541 aattattcca tcattaaaag atacgaggcg cgtgtaagtt acaggcaagc gatccgtcct
     6601 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     6661 gtc
//
