LOCUS       sequence-202550-        7216 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for pop-in/pop-out replacement of Vps10 with Vps10-msGFPx3..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7216)
  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 7216)
  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: YIplac211-VPS10-msGFPx3
FEATURES             Location/Qualifiers
     source          1..7216
                     /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"
     primer_bind     complement(3714..3731)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3714..3730)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3723..3745)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(3939..3958)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4058..4080
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(4198..5001)
                     /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(5002..5217)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(5224..5242)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5310..5414
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5415..5483,5484..6275)
                     /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(5633..5652)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6446..7034
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6935..6954
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7188..7205
                     /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 atgcctgcag gtcgactcta gaaagatccc tctatcaacc tgtgagggtg gcctgaaatt
      301 ggatgctccc tcatcaccac atgcttgccc aggaaaagaa aaagaattca aggaaaagta
      361 ctcagtaagt gccggtccct ttgcatttat tttcatttca attcttttaa taattttctt
      421 tgccgcatgg tttgtatatg acagaggtat cagaagaaat gggggatttg caaggtttgg
      481 agaaattagg ctaggtgacg atggtttaat agaaaacaat aatactgaca gagttgtcaa
      541 taacattgtg aaatcaggat tttacgtttt ctcaaatatc gggtctcttt tacagcacac
      601 aaaaactaat atagcgcatg ctatttccaa aattaaagga aggtttggaa acagaacagg
      661 tccaagctac tcatccctga tccatgatca atttttggat gaagcagatg acctgcttgc
      721 tggccacgat gaagacgcca atgacttatc cagtttcatg gatcagggta gtaattttga
      781 aatcgaagaa gatgatgttc caacacttga agaagagcat acatcatata cagatcaacc
      841 tacgaccacc gatgttccag atacattacc agaaggaaat gaggaaaaca tcgacaggcc
      901 tgattctaca gcgccatcta acgaaaacca gacggatcca ggaagttcgg ggtccagcgg
      961 ttcttcaggc gctagcatgg atagcactga gagcctgttc accggggtgg tgcccatcct
     1021 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgcgcggcg agggcgaggg
     1081 cgatgccacc aacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     1141 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     1201 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     1261 gcgcaccatc accttcaagg acgacggcac ctacaagacc cgcgccgagg tgaagttcga
     1321 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     1381 catcctgggg cacaagctgg agtacaactt caacagccac aacgtctata tcaccgccga
     1441 caagcagaag aacggcatca aggccaactt caagatccgc cacaacgtgg aggacggcag
     1501 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     1561 gcccgacaac cactacctga gcacccagtc caagctgagc aaagacccca acgagaagcg
     1621 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactcacg gcagctccgg
     1681 ctccagcggg tctagcatgg atagcactga gagcctgttc accggggtgg tgcccatcct
     1741 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgcgcggcg agggcgaggg
     1801 cgatgccacc aacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     1861 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     1921 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     1981 gcgcaccatc accttcaagg acgacggcac ctacaagacc cgcgccgagg tgaagttcga
     2041 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     2101 catcctgggg cacaagctgg agtacaactt caacagccac aacgtctata tcaccgccga
     2161 caagcagaag aacggcatca aggccaactt caagatccgc cacaacgtgg aggacggcag
     2221 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     2281 gcccgacaac cactacctga gcacccagtc caagctgagc aaagacccca acgagaagcg
     2341 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactcacg gcagctccgg
     2401 ctccagcggg tctagcatgg atagcactga gagcctgttc accggggtgg tgcccatcct
     2461 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgcgcggcg agggcgaggg
     2521 cgatgccacc aacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     2581 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     2641 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     2701 gcgcaccatc accttcaagg acgacggcac ctacaagacc cgcgccgagg tgaagttcga
     2761 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     2821 catcctgggg cacaagctgg agtacaactt caacagccac aacgtctata tcaccgccga
     2881 caagcagaag aacggcatca aggccaactt caagatccgc cacaacgtgg aggacggcag
     2941 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     3001 gcccgacaac cactacctga gcacccagtc caagctgagc aaagacccca acgagaagcg
     3061 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactcacg gcagctccgg
     3121 ctccagcggg taaagcggcc gcagagatta ctttacatag agtagataat tccatatact
     3181 tttcataaac aaaaacgagc agatccttta gacagaatat caagttcttt cataagccaa
     3241 agaaacttgg catgatcagt actggtgcaa ttcaaataac tgcaagtact gaatctccat
     3301 cttcaaagca gaaccgtaat aatgcgtaca aggaagcggg tgggatatat catatgttac
     3361 gggaaagtgg ttgtttgaca gaagatcatc gagaacgtca taacttcaat acataagtca
     3421 tcgcgtaatt ttacaagctt tactgttcag ctttatagga taatcagaat aagaaaaggc
     3481 agataaatag tttgataaat ggccagttgt gatggggaaa aagactttta gagaatggca
     3541 atatttcaag ttatcaatgt atgtatattt ttgacttttt gagtctcaac taccgaagag
     3601 aaataaacta ctaacgtact ttaatattta tagtacttca ttcgatcaag atgtggacga
     3661 tgcacatgct attgatcaaa tgacatggag gcaatggtac cgagctcgaa ttcactggcc
     3721 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca
     3781 gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc
     3841 caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct ccttacgcat
     3901 ctgtgcggta tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca
     3961 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     4021 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     4081 ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcg ggtaataact gatataatta
     4141 aattgaagct ctaatttgtg agtttagtat acatgcattt acttataata cagtttttta
     4201 gttttgctgg ccgcatcttc tcaaatatgc ttcccagcct gcttttctgt aacgttcacc
     4261 ctctacctta gcatcccttc cctttgcaaa tagtcctctt ccaacaataa taatgtcaga
     4321 tcctgtagag accacatcat ccacggttct atactgttga cccaatgcgt ctcccttgtc
     4381 atctaaaccc acaccgggtg tcataatcaa ccaatcgtaa ccttcatctc ttccacccat
     4441 gtctctttga gcaataaagc cgataacaaa atctttgtcg ctcttcgcaa tgtcaacagt
     4501 acccttagta tattctccag tagataggga gcccttgcat gacaattctg ctaacatcaa
     4561 aaggcctcta ggttcctttg ttacttcttc tgccgcctgc ttcaaaccgc taacaatacc
     4621 tgggcccacc acaccgtgtg cattcgtaat gtctgcccat tctgctattc tgtatacacc
     4681 cgcagagtac tgcaatttga ctgtattacc aatgtcagca aattttctgt cttcgaagag
     4741 taaaaaattg tacttggcgg ataatgcctt tagcggctta actgtgccct ccatggaaaa
     4801 atcagtcaag atatccacat gtgtttttag taaacaaatt ttgggaccta atgcttcaac
     4861 taactccagt aattccttgg tggtacgaac atccaatgaa gcacacaagt ttgtttgctt
     4921 ttcgtgcatg atattaaata gcttggcagc aacaggacta ggatgagtag cagcacgttc
     4981 cttatatgta gctttcgaca tgatttatct tcgtttcctg catgtttttg ttctgtgcag
     5041 ttgggttaag aatactgggc aatttcatgt ttcttcaaca ctacatatgc gtatatatac
     5101 caatctaagt ctgtgctcct tccttcgttc ttccttctgt tcggagatta ccgaatcaaa
     5161 aaaatttcaa ggaaaccgaa atcaaaaaaa agaataaaaa aaaaatgatg aattgaaaag
     5221 ctggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     5281 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     5341 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     5401 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     5461 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     5521 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     5581 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     5641 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     5701 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     5761 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     5821 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     5881 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     5941 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     6001 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     6061 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     6121 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     6181 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     6241 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     6301 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     6361 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     6421 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     6481 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     6541 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     6601 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     6661 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     6721 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     6781 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     6841 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     6901 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     6961 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     7021 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     7081 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     7141 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     7201 gcgaggaagc ggaaga
//
