LOCUS       sequence-199976-        5997 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for pop-in/pop-out replacement of Vps21 with msGFP-Vps21..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5997)
  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 5997)
  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-msGFP-VPS21
FEATURES             Location/Qualifiers
     source          1..5997
                     /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(619..640)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     1227..1248
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2490..2507)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2490..2506)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2499..2521)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2715..2734)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2834..2856
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2974..3777)
                     /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(3778..3998)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4005..4023)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4091..4195
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4196..4264,4265..5056)
                     /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(4414..4433)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5227..5815
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5716..5735
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5969..5986
                     /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 atgccggagg caacaacaaa gccaaaacct agattatgac gatataaact tatgctgagg
      301 taggtatata ccagaggatt tattttatag atattagtct gctgattatc tagtttcttg
      361 tcacctcata cgtgtttttt ttttttttgg ttttctcggt ttgcattatg gggctttttg
      421 ctaaaacaat atccccgaaa agacaaagaa agagaagtga ggtgaaaata cgagcagctt
      481 atcttataaa agtggcaacg attttaaaaa taaaaagcaa gatcgtcgac ttatctaagc
      541 tataaaaaaa tatccctttt atcacacaaa aaaatgaacg atagcactga gagcctgttc
      601 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
      661 gtgcgcggcg agggcgaggg cgatgccacc aacggcaagc tgaccctgaa gttcatctgc
      721 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac ctacggcgtg
      781 cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
      841 cccgaaggct acgtccagga gcgcaccatc accttcaagg acgacggcac ctacaagacc
      901 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
      961 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaactt caacagccac
     1021 aacgtctata tcaccgccga caagcagaag aacggcatca aggccaactt caagatccgc
     1081 cacaacgtgg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     1141 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc caagctgagc
     1201 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     1261 atcactcacg gcagctccgg ctccagcggg tccggaagtg gtagctcagg gtctagtgga
     1321 tccacatcag tcacttccat aaagttggta ctgttgggtg aggcagcagt tggtaaatcg
     1381 tcaatagtcc taaggtttgt atctaatgat tttgccgaaa ataaggagcc caccattggt
     1441 gcagcattcc tcactcaaag agttactata aatgaacata ctgttaagtt tgaaatatgg
     1501 gacactgctg ggcaagagag atttgcatct ttagcaccta tgtactatag aaacgcacaa
     1561 gctgctcttg tagtatacga cgttacaaaa ccacaatcct ttattaaagc gcgccactgg
     1621 gttaaggaat tacatgaaca ggcatcaaag gatatcatca ttgcgttagt tggtaacaag
     1681 atcgatatgc tgcaagaagg tggggaaaga aaagtcgcaa gggaagaagg tgaaaagcta
     1741 gcggaagaaa agggcttgtt gttctttgaa acaagtgcaa agacaggaga gaatgttaac
     1801 gatgtatttt tgggaatcgg tgaaaagata ccactgaaga cggctgaaga gcaaaatagt
     1861 gcaagtaacg aacgtgagtc gaacaatcaa agagtggatt tgaacgctgc taatgatggg
     1921 accagcgcaa acagtgcttg cagttgttag agaggggaaa ataaatatat ataaaaaaaa
     1981 agtctgggca aattgagata aatagatctt caaaacggga aagatgaaag tcataattta
     2041 tcgtacctag aaaggagagg gagttttcat tttttaccgt aatcgttgtt tttcttttct
     2101 tcttataatt tgtaaatttt ttaactgcat tagatcatta tattatcttt caaaacgcca
     2161 gtcgttttta ttagaccttt atatacgttt caaagtggtt tgtacacgtt agtttagtta
     2221 aacttttgtt ttcatgtaga ccaaaatata aacttatttt atgccgaaaa tcgggagatg
     2281 gattagttaa tattgtaaat aatgataaat taaaggaaca ttaaatttac aaagttgcaa
     2341 tgaggagcaa tatatatata tatatatgga tttctgagaa attaattaag cagtatttga
     2401 acacatgtcg tttactcttt cttatccttt atgtctaatt tttcgagtag atcggtaatt
     2461 acagctgtca aggtaccgag ctcgaattca ctggccgtcg ttttacaacg tcgtgactgg
     2521 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg
     2581 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc
     2641 gaatggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     2701 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     2761 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     2821 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     2881 gcgagacgaa agggcgggta ataactgata taattaaatt gaagctctaa tttgtgagtt
     2941 tagtatacat gcatttactt ataatacagt tttttagttt tgctggccgc atcttctcaa
     3001 atatgcttcc cagcctgctt ttctgtaacg ttcaccctct accttagcat cccttccctt
     3061 tgcaaatagt cctcttccaa caataataat gtcagatcct gtagagacca catcatccac
     3121 ggttctatac tgttgaccca atgcgtctcc cttgtcatct aaacccacac cgggtgtcat
     3181 aatcaaccaa tcgtaacctt catctcttcc acccatgtct ctttgagcaa taaagccgat
     3241 aacaaaatct ttgtcgctct tcgcaatgtc aacagtaccc ttagtatatt ctccagtaga
     3301 tagggagccc ttgcatgaca attctgctaa catcaaaagg cctctaggtt cctttgttac
     3361 ttcttctgcc gcctgcttca aaccgctaac aatacctggg cccaccacac cgtgtgcatt
     3421 cgtaatgtct gcccattctg ctattctgta tacacccgca gagtactgca atttgactgt
     3481 attaccaatg tcagcaaatt ttctgtcttc gaagagtaaa aaattgtact tggcggataa
     3541 tgcctttagc ggcttaactg tgccctccat ggaaaaatca gtcaagatat ccacatgtgt
     3601 ttttagtaaa caaattttgg gacctaatgc ttcaactaac tccagtaatt ccttggtggt
     3661 acgaacatcc aatgaagcac acaagtttgt ttgcttttcg tgcatgatat taaatagctt
     3721 ggcagcaaca ggactaggat gagtagcagc acgttcctta tatgtagctt tcgacatgat
     3781 ttatcttcgt ttcctgcatg tttttgttct gtgcagttgg gttaagaata ctgggcaatt
     3841 tcatgtttct tcaacactac atatgcgtat atataccaat ctaagtctgt gctccttcct
     3901 tcgttcttcc ttctgttcgg agattaccga atcaaaaaaa tttcaaagaa accgaaatca
     3961 aaaaaaagaa taaaaaaaaa atgatgaatt gaattgaaaa gctggcctcg tgatacgcct
     4021 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
     4081 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     4141 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     4201 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     4261 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     4321 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     4381 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     4441 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     4501 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     4561 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     4621 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     4681 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     4741 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     4801 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     4861 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     4921 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     4981 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     5041 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     5101 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     5161 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     5221 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     5281 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     5341 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     5401 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     5461 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     5521 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     5581 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     5641 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     5701 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     5761 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     5821 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     5881 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     5941 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaaga
//
