LOCUS       sequence-202385-        7666 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Overexpresses iGFP-RCY1 from the TPI1 promoter. Integrating at 
            TRP1..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7666)
  AUTHORS   Day KJ, Casler JC, Glick BS
  TITLE     Budding Yeast Has a Minimal Endomembrane System.
  JOURNAL   Dev Cell. 2018 Jan 8;44(1):56-72.e4. doi: 
            10.1016/j.devcel.2017.12.014. Epub 2018 Jan 8.
  PUBMED    29316441
REFERENCE   2  (bases 1 to 7666)
  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: 
            "10.1016/j.devcel.2017.12.014"; journalName: "Dev Cell"; date: 
            "2018-01-8- 8"; volume: "44"; issue: "1"; pages: "56-72.e4"
COMMENT     Sequence Label: YIplac204-T|C-iGFP-RCY1
FEATURES             Location/Qualifiers
     source          1..7666
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    106..127
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(142..147,148..165,166..172)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    180..196
                     /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     185..207
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     204..220
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     204..220
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     terminator      complement(239..486)
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     452..470
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     3994..4022
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     regulatory      4049..4058
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(4082..4512)
                     /gene="S. cerevisiae TPI1"
                     /label=TPI1 promoter
                     /note="strong constitutive promoter for yeast triose 
                     phosphate isomerase"
     primer_bind     complement(4636..4655)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4755..4777
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4896..5570)
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     promoter        complement(5571..5672)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(5674..5692)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5760..5864
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5865..5933,5934..6725)
                     /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(6083..6102)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6896..7484
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7385..7404
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7638..7655
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttgc
      241 aaattaaagc cttcgagcgt cccaaaacct tctcaagcaa ggttttcagt ataatgttac
      301 atgcgtacac gcgtctgtac agaaaaaaaa gaaaaatttg aaatataaat aacgttctta
      361 atactaacat aactataaaa aaataaatag ggacctagac ttcaggttgt ctaactcctt
      421 ccttttcggt tagagcggat gtggggggag ggcgtgaatg taagcgtgac ataactaatt
      481 acatgatgac tctagaacta tgcgttagta aaagaataag tgtgaaaaaa aaaaaaatag
      541 aatagcgtat gaagaatgaa catactttcc taatagaaag ggagcgtgct gaaccatgtt
      601 atcgatgatc ttgtgtggaa tcctcttata aatggagtga acatttgtat gccccttgtg
      661 tgcgtgtggc taacagggaa aaatagagcc cgcgtaatct tgaataatct aatatgtgaa
      721 cagtctttta tttccgcact tcatacctat atagagtaat ataattcatc attgatagga
      781 atttacataa tgcagcaatc tcgtatgcct aggccgtaca ttaccttttc aacgtctttc
      841 cgtacacgaa cccaatcgct tctcctctga acaaattcat aaatttcctc ttgcgtaaaa
      901 atgccattga atttcccaag atcagaaata agctttccca actctttcga gtccttgcct
      961 gaaatgatgt ataactggcc aatagactta agcccaacaa aatagggaac aacatttttt
     1021 tgttttaatt tgttagcaat aaaatcgtaa aagtaattta aatcgcagat taaccaaatt
     1081 gcgccctccg tagatatgaa gcacttcttc agatgtttga cgatttcatt aaagaatctc
     1141 tcgccaattt cttgttggta tacgtcaata gtacccttgt ctgtggcacc agtcagtaga
     1201 aagcaatgat tagaaagtaa ctcgactacc ctgatagcac atttggttgg ctttatatcc
     1261 cttattggag aattcctaga gggtggcggt ggattataat cgtcaggcaa ttgaagagtc
     1321 ttgaatacaa atgagatttc atccattagt ttgtttatac cgatatttaa accttcggct
     1381 acaaaatcat cgatagaagt ctcaaaattt ttttttaatt ggataacatc attaagaaaa
     1441 tctttatttt tatcgataat ttttttggga atcaattcgt tcttgtaaaa aatggaaatc
     1501 atttgtaata ttatgtcacc tatattgata agttcagtga attgtactaa aggttcgact
     1561 tgagtattgg ctggcgaaag tgaattgaat tttatttcta tttgttctat ttcgttagga
     1621 ttatattttt gtagtagcag gattgccttt tcaaatgcag gtttaacgtg atttttattc
     1681 aattgtgtaa ttaaaatttt gaaaatctct tgacagcgta gcttgatcaa tggtagcaaa
     1741 ctttcaatcg tatgaaattt gtaaatttga tccgtctttt cgcttgtttc ctgtaaaatt
     1801 ttgtaacata aatcgagact tattaacgat ttaatgtttt gcaagttatt ggaaattaaa
     1861 tttaagttgt aggctagttg tagttgttgc tttagccttt gattttcatt gttcgagaac
     1921 ttgaaaatct tggtgaagga acttagaaag tctagtttat cattagaaat atcagttgaa
     1981 tgaactgatt gatggatcgt acttggtttt gaatccgctt ctgggacagt tttggaggtt
     2041 tcagtaatta ttgagaggtc attcggcaat tcattgttac ttgcactcga agcatccgtt
     2101 tgatctttta aagtgttata aatttttgcg ttttgctcac gctgcttgtc ttgaacttgt
     2161 gtgtcataat tagctaattg tgagttgatc aatgattcaa atactctagt tgtctgattc
     2221 atatagttga caacaaaggg ctcgagatac aggtttaaaa ggtcacgaac aactttacgg
     2281 aaatttgcgc ctgcgtttac tgagtcgttt aattcagtat caaacttttt tataaatgtg
     2341 aaataaagtt cgggtacgca ttcataattg gctttactat cctttccatt ttcttttaaa
     2401 aattcactat agcttaataa gatattgttt aaaatatcta ataaaatatc ttggataaag
     2461 gactcgatga actgtaatat tacagggaac tcatctttga ataaaatgtc cgctaattta
     2521 atcttttcgt tgaggaaatc ccttagctga ttgagaaacg tatctaaatt tttagaatta
     2581 aatatcacgg tactcccacc gctgtcctct tcccttggtt gttcggccca aaataattca
     2641 tcattagacg gcaaaaccgt taaactggga ggcaaatcag cctttgactt gaaaaattcg
     2701 atagcattgc tttcctcatt agatatcaat aaaattttca tgaaacgagc gacagcagct
     2761 aaattattag actggtaatt caattctgac tctttgagaa cagaattaat gaagatttcc
     2821 ctaagaatgt tgaaattagt ttcaatccta gtaaatgttt ctatatcatt tttattcgag
     2881 aaattgtact ttttgataaa atttagaatt cttgtttgtt tcaacggatc gttggaatat
     2941 gaagtaggaa agaaatttgc tagattatta ttatatagct tcctagcgta tgagttataa
     3001 caacggtaaa atttcacaaa gattttttta gaattttgag cagtgaaaga tttgtatatt
     3061 tcgaagatgc gaagggaatt ttgatcatca ttttcatcca gcaacgtaat ttcttcttct
     3121 tcatttgaat gcacctgttg gagacccatt ctagtcaatt tcaatgacca atactttgaa
     3181 tcatttttgc cgttaatgat tgcataaacc cgtttattca cttgttgaaa actcagataa
     3241 tctacagtag ataaatatga ggcaatattg gttacaatct cgggaacttt tagcagatca
     3301 tccatggatc cactagaccc tgagctacca cttccggacc cgctggagcc ggagctgcca
     3361 tgtgtaatcc cagcagctgt tacaaactct ttaaggacca tgtggtctct cttttcgttg
     3421 ggatctttcg aaagggcaga ttgtgtggac aggtaatggt tgtctggtaa aaggacaggg
     3481 ccatcgccaa ttggagtatt ttgttgataa tggtctgcta gttgaacgct tccatcttca
     3541 atgttgtgtc taattttgaa gttagctttg attccattct tttgtttgtc tgccatgatg
     3601 tatacattgt gtgagttata gttgtattcc aatttgtgcc caagaatgtt tccatcttct
     3661 ttaaaatcaa taccttttaa ctcgattcta ttaacaaggg tatcaccttc aaacttgact
     3721 tcagcacgtg tcttgtagtt cccgtcatct ttgaaaaata tagttctttc ctgtacataa
     3781 ccttcgggca tggcactctt gaaaaagtca tgctgtttca tatgatccgg gtatcttgaa
     3841 aagcactgaa caccataagt taaagtagtg acaagtgttg gccaaggaac aggtagtttt
     3901 ccagtagtgc aaataaattt aagggtaagt tttccgtatg ttgcatcacc ttcaccctct
     3961 ccactgacag aaaatttgtg cccattaaca tcaccatcta attcaacaag aattgggaca
     4021 actccagtga aaaggctctc agtgctatcc atggtggcga ccgggtaccg agctcgaatt
     4081 cttttagttt atgtatgtgt tttttgtagt tatagattta agcaagaaaa gaatacaaac
     4141 aaaaaattga aaaagattga tttagaatta aaaagaaaaa tatttacgta agaagggaaa
     4201 atagtaaatg ttgcaagttc actaaactcc taaattatgc tgccctttat attccctgtt
     4261 acagcagccg agccaaaggt atataggctc ctttgcatta gcatgcgtaa caaaccacct
     4321 gtcagtttca accgaggtgg tatccgagag aattgtgtga ttgctttaat taatttcgga
     4381 gaatctcaca tgccactgaa gattaaaaac tggatgccag aaaaggggtg tccaggtgta
     4441 acatcaatag aggaagctga aaagtcttag aacgggtaat cttccaccaa cctgatgggt
     4501 tcctagatat aatctcgaag ggaataagta gggtgatacc gcagaagtgt ctgaatgtat
     4561 taaggtccgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     4621 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     4681 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     4741 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     4801 cgcgagacga aagggcgatc ttttatgctt gcttttcaaa aggcttgcag gcaagtgcac
     4861 aaacaatact taaataaata ctactcagta ataacctatt tcttagcatt tttgacgaaa
     4921 tttgctattt tgttagagtc ttttacacca tttgtctcca cacctccgct tacatcaaca
     4981 ccaataacgc catttaatct aagcgcatca ccaacatttt ctggcgtcag tccaccagct
     5041 aacataaaat gtaagctctc ggggctctct tgccttccaa cccagtcaga aatcgagttc
     5101 caatccaaaa gttcacctgt cccacctgct tctgaatcaa acaagggaat aaacgaatga
     5161 ggtttctgtg aagctgcact gagtagtatg ttgcagtctt ttggaaatac gagtctttta
     5221 ataactggca aaccgaggaa ctcttggtat tcttgccacg actcatctcc atgcagttgg
     5281 acgatatcaa tgccgtaatc attgaccaga gccaaaacat cctccttagg ttgattacga
     5341 aacacgccaa ccaagtattt cggagtgcct gaactatttt tatatgcttt tacaagactt
     5401 gaaattttcc ttgcaataac cgggtcaatt gttctctttc tattgggcac acatataata
     5461 cccagcaagt cagcatcgga atctagtgca cattctgcgg cctctgtgct ctgcaagccg
     5521 caaactttca ccaatggacc agaactacct gtgaaattaa taacagacat actccaagct
     5581 gcctttgtgt gcttaatcac gtatactcac gtgctcaata gtcaccaatg ccctccctct
     5641 tggccctctc cttttctttt ttcgaccgaa ttggcctcgt gatacgccta tttttatagg
     5701 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     5761 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     5821 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     5881 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     5941 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     6001 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     6061 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     6121 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     6181 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     6241 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     6301 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     6361 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     6421 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     6481 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     6541 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     6601 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     6661 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     6721 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     6781 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     6841 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     6901 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     6961 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     7021 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     7081 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     7141 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     7201 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     7261 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     7321 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     7381 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     7441 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     7501 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     7561 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
     7621 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaaga
//
