LOCUS       sequence-212482-        6383 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for pop-in/pop-out replacement of Vph1 with Vph1-mEGFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6383)
  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 6383)
  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-VPH1-mEGFP
FEATURES             Location/Qualifiers
     source          1..6383
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     395..412
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    529..550
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(565..570,571..588,589..595)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    603..619
                     /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     608..630
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     627..643
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     627..643
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(1191..1193,1194..1196,1197..1910)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1236..1257)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1497..1516)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1844..1865
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2924..2941)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2924..2940)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2933..2955)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(3149..3168)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3268..3290
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3408..4211)
                     /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(4212..4427)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4434..4452)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4520..4624
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4625..4693,4694..5485)
                     /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(4843..4862)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5656..6244
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
       61 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
      121 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
      181 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
      241 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
      301 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
      361 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
      421 agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
      481 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
      541 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
      601 aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt acgccaagct
      661 tgcatgcgcc atcgactgaa gcagatgcta gttctgaaga tttggaagca caacaattaa
      721 tttccgcgat ggacgccgat gacgctgaag aagaagaagt tggttctgga tctcatggtg
      781 aagactttgg tgatattatg attcatcaag ttattcatac aattgaattc tgtttgaatt
      841 gtgtttcgca cactgcatcc tatttacgtt tatgggcctt atcattggca catgctcaat
      901 tgtctagtgt tttatggaca atgacaattc aaattgcctt tggatttaga ggatttgtgg
      961 gtgtgtttat gacggttgca ctttttgcca tgtggttcgc actaacatgt gcagttcttg
     1021 ttttgatgga aggtacatct gccatgcttc attccttacg tttgcactgg gttgaatcta
     1081 tgtccaagtt tttcgtgggt gaaggtttac catacgaacc attcgcattt gagtataaag
     1141 acatggaagt cgctgttgct agtgcaagct cttccgcttc aagcggatcc atggtgagca
     1201 agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa
     1261 acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga
     1321 ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca
     1381 ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact
     1441 tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg
     1501 acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca
     1561 tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt
     1621 acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg
     1681 tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc
     1741 agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca
     1801 cccagtccaa gctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt
     1861 tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagtaa agcggccgca
     1921 ttttgaggac ttttaaaaaa agcgaaacat ttaagtactt cattaaataa taaatgatgc
     1981 tactatatag tgagtaaacc ttattatata ctatcaatgc taattcaata aagtatttaa
     2041 agtatcaacc aatgcttaaa aagagtcaaa aaaaagggaa aaagaaatag gaaagccaaa
     2101 cttatctcat gaaaacgtta gacttgcaat ccaatccacc agtgaccatc atttgccttg
     2161 gggtaacgtt agaatcgaaa tccacatcca ggtctaaaca gttgacaaat cctgtatgca
     2221 acttttccca aacgtgtaag cactgccccg tgttcagatc ccaacatctg acagatttat
     2281 catcagcaca agaaaacaag tattggcctc gaatactaat atctctaacc cacgaaagat
     2341 gccccttcaa cgtaagaaca catctgaaat ttgaatctgt aggattcggt atgggtgctc
     2401 tatgggccat taacgtaggt agaggtatct cccagatttt tatcgttcta tctcttgacg
     2461 ctgagacaca atattgcatc ccccaattct tatatcgatc cgttgatggt gttctaaacc
     2521 ttatctccgg agagtcttcg atgaaatgaa taaatttaac tttctctatg ggaaattcat
     2581 gacccgtgcc aacactaaga ccattacctg atggccaatg tgttaacctt aatgtagtgt
     2641 catgtgaacc agagatgatg taatcgccta gaacgtctat ggaacgaacc cattgggaat
     2701 gaggctgaaa tgttttcaac gaccaaccat tgtggaagtc ccaaatttta acagtttgat
     2761 ctctggaaca ggacgctata tggacatcat tatttttctg ccagattttt accgcagaga
     2821 caatatgctc atggcccaat aaagacctaa tttgttggaa cttgcattcc tcggagaccc
     2881 acttgaaaac atgaatctgc aaatctttag acgcggtggt accactggcc gtcgttttac
     2941 aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc
     3001 ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc
     3061 gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     3121 tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     3181 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     3241 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     3301 catcaccgaa acgcgcgaga cgaaagggcg ggtaataact gatataatta aattgaagct
     3361 ctaatttgtg agtttagtat acatgcattt acttataata cagtttttta gttttgctgg
     3421 ccgcatcttc tcaaatatgc ttcccagcct gcttttctgt aacgttcacc ctctacctta
     3481 gcatcccttc cctttgcaaa tagtcctctt ccaacaataa taatgtcaga tcctgtagag
     3541 accacatcat ccacggttct atactgttga cccaatgcgt ctcccttgtc atctaaaccc
     3601 acaccgggtg tcataatcaa ccaatcgtaa ccttcatctc ttccacccat gtctctttga
     3661 gcaataaagc cgataacaaa atctttgtcg ctcttcgcaa tgtcaacagt acccttagta
     3721 tattctccag tagataggga gcccttgcat gacaattctg ctaacatcaa aaggcctcta
     3781 ggttcctttg ttacttcttc tgccgcctgc ttcaaaccgc taacaatacc tgggcccacc
     3841 acaccgtgtg cattcgtaat gtctgcccat tctgctattc tgtatacacc cgcagagtac
     3901 tgcaatttga ctgtattacc aatgtcagca aattttctgt cttcgaagag taaaaaattg
     3961 tacttggcgg ataatgcctt tagcggctta actgtgccct ccatggaaaa atcagtcaag
     4021 atatccacat gtgtttttag taaacaaatt ttgggaccta atgcttcaac taactccagt
     4081 aattccttgg tggtacgaac atccaatgaa gcacacaagt ttgtttgctt ttcgtgcatg
     4141 atattaaata gcttggcagc aacaggacta ggatgagtag cagcacgttc cttatatgta
     4201 gctttcgaca tgatttatct tcgtttcctg catgtttttg ttctgtgcag ttgggttaag
     4261 aatactgggc aatttcatgt ttcttcaaca ctacatatgc gtatatatac caatctaagt
     4321 ctgtgctcct tccttcgttc ttccttctgt tcggagatta ccgaatcaaa aaaatttcaa
     4381 ggaaaccgaa atcaaaaaaa agaataaaaa aaaaatgatg aattgaaaag ctggcctcgt
     4441 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     4501 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     4561 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     4621 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     4681 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     4741 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     4801 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     4861 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     4921 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     4981 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     5041 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     5101 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     5161 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     5221 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     5281 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     5341 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     5401 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     5461 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     5521 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     5581 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     5641 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     5701 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     5761 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     5821 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     5881 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     5941 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     6001 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     6061 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     6121 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     6181 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     6241 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     6301 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     6361 agctgatacc gctcgccgca gcc
//
