LOCUS       sequence-222003-        7205 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express Erg11-iGFPx3 at the endogenous locus via pop-in pop-out 
            mutagenesis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7205)
  AUTHORS   Casler JC, Papanikou E, Barrero JJ, Glick BS
  TITLE     Maturation-driven transport and AP-1-dependent recycling of a 
            secretory cargo in the Golgi.
  JOURNAL   J Cell Biol. 2019 May 6;218(5):1582-1601. doi: 
            10.1083/jcb.201807195. Epub 2019 Mar 11.
  PUBMED    30858194
REFERENCE   2  (bases 1 to 7205)
  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: 
            "10.1083/jcb.201807195"; journalName: "J Cell Biol"; date: 
            "2019-05-6- 6"; volume: "218"; issue: "5"; pages: "1582-1601"
COMMENT     Sequence Label: YIplac211-Erg11-iGFPx3
FEATURES             Location/Qualifiers
     source          1..7205
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     44..63
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     257..279
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     271..288
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     272..288
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3771..3787)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3771..3787)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3784..3806)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3795..3811
                     /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(3819..3825,3826..3843,3844..3849))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3864..3885
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4002..4019)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4173..4761)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4253..4272)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4932..5723,5724..5792))
                     /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     5555..5574
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5793..5897)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5965..5983
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5990..6205
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             6206..7009
                     /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"
     primer_bind     complement(7127..7149)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc agagcagatt
       61 gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
      121 cgcatcaggc gccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg
      181 gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg
      241 gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga attcgagctc
      301 ggtaccgcac gtccaaacac ttcttatgtc atcccagcag gttatcacgt tttggtttct
      361 ccaggttaca ctcatttaag agacgaatac ttccctaatg ctcaccaatt caacattcac
      421 cgttggaaca aagattctgc ctcctcttat tccgtcggtg aagaagtcga ttacggtttc
      481 ggtgccattt ctaagggtgt cagctctcca tacttacctt tcggtggtgg tagacacaga
      541 tgtatcggtg aacactttgc ttactgtcag ctaggtgttc taatgtccat ttttatcaga
      601 acattaaaat ggcattaccc agagggtaag accgttccac ctcctgactt tacatctatg
      661 gttactcttc caaccggtcc agccaagatc atctgggaaa agagaaatcc agaacaaaag
      721 atctgggatc caggaagttc ggggtccagc ggttcttcag gcgctagcat ggatagcact
      781 gagagccttt tcactggagt tgtcccaatt cttgttgaat tagatggtga tgttaatggg
      841 cacaaatttt ctgtcagtgg agagggtgaa ggtgatgcaa catacggaaa acttaccctt
      901 aaatttattt gcactactgg aaaactacct gttccttggc caacacttgt cactacttta
      961 acttatggtg ttcagtgctt ttcaagatac ccggatcata tgaaacagca tgactttttc
     1021 aagagtgcca tgcccgaagg ttatgtacag gaaagaacta tatttttcaa agatgacggg
     1081 aactacaaga cacgtgctga agtcaagttt gaaggtgata cccttgttaa tagaatcgag
     1141 ttaaaaggta ttgattttaa agaagatgga aacattcttg ggcacaaatt ggaatacaac
     1201 tataactcac acaatgtata catcatggca gacaaacaaa agaatggaat caaagctaac
     1261 ttcaaaatta gacacaacat tgaagatgga agcgttcaac tagcagacca ttatcaacaa
     1321 aatactccaa ttggcgatgg ccctgtcctt ttaccagaca accattacct gtccacacaa
     1381 tctgcccttt cgaaagatcc caacgaaaag agagaccaca tggtccttaa agagtttgta
     1441 acagctgctg ggattacaca tggcagctcc ggctccagcg ggtctagcat ggatagcact
     1501 gagagccttt tcactggagt tgtcccaatt cttgttgaat tagatggtga tgttaatggg
     1561 cacaaatttt ctgtcagtgg agagggtgaa ggtgatgcaa catacggaaa acttaccctt
     1621 aaatttattt gcactactgg aaaactacct gttccttggc caacacttgt cactacttta
     1681 acttatggtg ttcagtgctt ttcaagatac ccggatcata tgaaacagca tgactttttc
     1741 aagagtgcca tgcccgaagg ttatgtacag gaaagaacta tatttttcaa agatgacggg
     1801 aactacaaga cacgtgctga agtcaagttt gaaggtgata cccttgttaa tagaatcgag
     1861 ttaaaaggta ttgattttaa agaagatgga aacattcttg ggcacaaatt ggaatacaac
     1921 tataactcac acaatgtata catcatggca gacaaacaaa agaatggaat caaagctaac
     1981 ttcaaaatta gacacaacat tgaagatgga agcgttcaac tagcagacca ttatcaacaa
     2041 aatactccaa ttggcgatgg ccctgtcctt ttaccagaca accattacct gtccacacaa
     2101 tctgcccttt cgaaagatcc caacgaaaag agagaccaca tggtccttaa agagtttgta
     2161 acagctgctg ggattacaca tggcagctcc ggctccagcg ggtctagcat ggatagcact
     2221 gagagccttt tcactggagt tgtcccaatt cttgttgaat tagatggtga tgttaatggg
     2281 cacaaatttt ctgtcagtgg agagggtgaa ggtgatgcaa catacggaaa acttaccctt
     2341 aaatttattt gcactactgg aaaactacct gttccttggc caacacttgt cactacttta
     2401 acttatggtg ttcagtgctt ttcaagatac ccggatcata tgaaacagca tgactttttc
     2461 aagagtgcca tgcccgaagg ttatgtacag gaaagaacta tatttttcaa agatgacggg
     2521 aactacaaga cacgtgctga agtcaagttt gaaggtgata cccttgttaa tagaatcgag
     2581 ttaaaaggta ttgattttaa agaagatgga aacattcttg ggcacaaatt ggaatacaac
     2641 tataactcac acaatgtata catcatggca gacaaacaaa agaatggaat caaagctaac
     2701 ttcaaaatta gacacaacat tgaagatgga agcgttcaac tagcagacca ttatcaacaa
     2761 aatactccaa ttggcgatgg ccctgtcctt ttaccagaca accattacct gtccacacaa
     2821 tctgcccttt cgaaagatcc caacgaaaag agagaccaca tggtccttaa agagtttgta
     2881 acagctgctg ggattacaca tggcagctcc ggctccagcg ggtaaagcgg ccgcggtttg
     2941 taactgagaa aaaaaaaaaa aacagaaaaa gagagaagtt gtacattcac gtgggcctta
     3001 tcgccacact catccatctc caatgattta ttttatttgc cagtacgttt gatatgacat
     3061 cgagttatgt tctttagtta ctttcaaact tggacaacca tcccgcatta tttatgagtg
     3121 gtatggtttc acttctttat ttataaatat ttatactacg aaattatata tagattacat
     3181 ttctaaaaca acatcaaata taaggtttat ttatgcaaca tgcctactgg attagaagag
     3241 agaacacaca gctcacaggc caagcggaaa taaagcagtc ttttcgcaaa cttgtaaatc
     3301 tttttgcaat ttaaattaat caagagaggg cccaggagga gaggagtttt ggggaggggt
     3361 cactgaggaa gctctcttaa atatggattc actgaaataa atgtatataa gattcattac
     3421 tgctttttat agtgataaat ccaaatgatg agtgaattga accatcctta gtatccattg
     3481 ttttataaag caccgctaaa aataatatat tgctacaaaa atgatataat aataatatta
     3541 cgtaaaatac ctgaaaccgc cataaaaata atacctgaat taaccttaaa attctatccc
     3601 ataagctttt ttgtaaaggg ccagataatc ttgagtctcc ttcaaatgat tctcacccag
     3661 aagttcaaag tccatttgct tctgtaacat ctcattaatg gaaatcatat tgctttcctt
     3721 tgactgattg attgcgtcct cctcgggcat gcaagcttgg cgtaatcatg gtcatagctg
     3781 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     3841 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     3901 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     3961 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     4021 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     4081 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     4141 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     4201 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     4261 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     4321 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     4381 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     4441 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     4501 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     4561 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     4621 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     4681 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     4741 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     4801 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     4861 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     4921 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     4981 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     5041 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     5101 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     5161 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     5221 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     5281 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     5341 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     5401 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     5461 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     5521 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     5581 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     5641 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     5701 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     5761 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     5821 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     5881 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt
     5941 attatcatga cattaaccta taaaaatagg cgtatcacga ggccagcttt tcaattcatc
     6001 attttttttt tattcttttt tttgatttcg gtttccttga aatttttttg attcggtaat
     6061 ctccgaacag aaggaagaac gaaggaagga gcacagactt agattggtat atatacgcat
     6121 atgtagtgtt gaagaaacat gaaattgccc agtattctta acccaactgc acagaacaaa
     6181 aacatgcagg aaacgaagat aaatcatgtc gaaagctaca tataaggaac gtgctgctac
     6241 tcatcctagt cctgttgctg ccaagctatt taatatcatg cacgaaaagc aaacaaactt
     6301 gtgtgcttca ttggatgttc gtaccaccaa ggaattactg gagttagttg aagcattagg
     6361 tcccaaaatt tgtttactaa aaacacatgt ggatatcttg actgattttt ccatggaggg
     6421 cacagttaag ccgctaaagg cattatccgc caagtacaat tttttactct tcgaagacag
     6481 aaaatttgct gacattggta atacagtcaa attgcagtac tctgcgggtg tatacagaat
     6541 agcagaatgg gcagacatta cgaatgcaca cggtgtggtg ggcccaggta ttgttagcgg
     6601 tttgaagcag gcggcagaag aagtaacaaa ggaacctaga ggccttttga tgttagcaga
     6661 attgtcatgc aagggctccc tatctactgg agaatatact aagggtactg ttgacattgc
     6721 gaagagcgac aaagattttg ttatcggctt tattgctcaa agagacatgg gtggaagaga
     6781 tgaaggttac gattggttga ttatgacacc cggtgtgggt ttagatgaca agggagacgc
     6841 attgggtcaa cagtatagaa ccgtggatga tgtggtctct acaggatctg acattattat
     6901 tgttggaaga ggactatttg caaagggaag ggatgctaag gtagagggtg aacgttacag
     6961 aaaagcaggc tgggaagcat atttgagaag atgcggccag caaaactaaa aaactgtatt
     7021 ataagtaaat gcatgtatac taaactcaca aattagagct tcaatttaat tatatcagtt
     7081 attacccgcc ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc
     7141 agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc
     7201 agggc
//
