LOCUS       sequence-275843-        5588 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for pop-in/pop-out replacement of Mup1 with Mup1-mScarlet2I.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5588)
  AUTHORS   Casler JC, Glick BS
  TITLE     A microscopy-based kinetic analysis of yeast vacuolar protein 
            sorting.
  JOURNAL   Elife. 2020 Jun 25;9. pii: 56844. doi: 10.7554/eLife.56844.
  PUBMED    32584255
REFERENCE   2  (bases 1 to 5588)
  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"; journalName: "Elife."; 
            date: "2020-06-25"; volume: "9. pii: 56844. doi"; pages: " 
            10.7554/eLife.56844"
COMMENT     Sequence Label: YIplac211-Mup1-mScarlet2I
FEATURES             Location/Qualifiers
     source          1..5588
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    127..148
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(163..168,169..186,187..193)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    201..217
                     /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     206..228
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     225..241
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     225..241
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2107..2124)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2107..2123)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2116..2138)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2332..2351)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2451..2473
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2591..3394)
                     /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(3395..3610)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(3617..3635)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3703..3807
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3808..3876,3877..4668)
                     /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(4026..4045)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4839..5427
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5328..5347
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     join(5581..5588,1..10)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc
       61 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
      121 cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
      181 ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga
      241 ccatgattac gccaagcttg catgcctgca ggacgtgaag gtgtgttgcc cttttctaat
      301 ttctttgcct cttctaagcc attcaactcc ccaatggttg gtttgtttca acattttatt
      361 gtttgtacag taaccatttt agccccacct ccaggtgatg catatttgct agtccaaaat
      421 ttaatttcct acccaatgaa tattattaat tttgccatta gcgcgggttt gctttggata
      481 tactggcaac gtagacaagg taaaattgaa tggaacccac caattaaggc cggtgttttt
      541 gttacgggat ttttcacatt atccaattta tacctgatta ttgctcctta cgttccacct
      601 tcaaatggcg agtcggtgta ttcaagcatg ccatactgga tccactgtgt aattgcttgg
      661 ggtattttct tctttggtgg tgtttactac gttgtttggg cccaattatt gccaagatgg
      721 ggccactaca aattggtttc caaggatgtg cttggtgaag atgggttctg gagagtcaaa
      781 attgccaaag tttatgatga cactattggc gatgtcgata cacaagaaga cggcgttatt
      841 gaaacgaata taatcgaaca ttacaaaagt gaacaagaaa aatcgctgaa agatccaact
      901 agtatggata gtacagaagc agttataaag gaattcatga gattcaaagt ccacatggaa
      961 ggtagtatga acggtcacga attcgaaata gaaggtgaag gtgaaggtag accatatgaa
     1021 ggtactcaaa cagctaaatt gaaggttact aaaggtggtc cattgccatt ttcttgggat
     1081 atcttgtctc cacaattcat gtacggttca agagctttta ttaaacatcc agcagatatt
     1141 ccagattact acaagcaatc atttccagaa ggttttaaat gggaaagagt tatgaacttc
     1201 gaagatggtg gtgcagttac tgttacacaa gatacttctt tggaagatgg tacattgatc
     1261 tataaggtta aattgagagg tacaaatttt ccaccagatg gtccagttat gcaaaagaaa
     1321 actatgggtt gggaagcatc tacagaaaga ttgtacccag aagatggtgt tttgaagggt
     1381 gacatcaaga tggcattgag attgaaggat ggtggtagat atttggctga tttcaagact
     1441 acatacaagg ctaagaaacc agttcaaatg ccaggtgctt acaacgttga tagaaaattg
     1501 gatatcactt ctcataacga agattacaca gttgttgaac aatacgaaag aagtgaaggt
     1561 agacatagta caggtggtag tggtgctagc taagcggccg cccaaaaagg tacttaccat
     1621 ctcatctcga ttcttataat cacgtatgaa cattcttata attgcatttt ctttatacaa
     1681 aatatatagg taccaaacct cattataatt tctgccattt aatagtttaa gatgctattc
     1741 cgtttcgtat tttttcaatt tttataccta ctattgcgcg aaatcaaaat agcatactaa
     1801 gaaaaagcaa tatcatgaat tgctgatgag atagattcaa attggaatta actaaatcta
     1861 tcagcaacgc tataaacaca gataagcgct ctataaaatc gtttgtctaa cgttttatat
     1921 agttctttat tgctccctta ccctctttcc gtcaatagtg attgttgtct cgggaaagct
     1981 ttttttttta ttaatcaaag gcccaattga gttacaactg aaattaagct gagttggcgg
     2041 aacagtgtgg tattagcgaa gggaaaatct gtgaagtgcc ctagctacaa aacagagctc
     2101 gaattcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact
     2161 taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac
     2221 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggcgcctga tgcggtattt
     2281 tctccttacg catctgtgcg gtatttcaca ccgcatatgg tgcactctca gtacaatctg
     2341 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
     2401 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     2461 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcgggtaata
     2521 actgatataa ttaaattgaa gctctaattt gtgagtttag tatacatgca tttacttata
     2581 atacagtttt ttagttttgc tggccgcatc ttctcaaata tgcttcccag cctgcttttc
     2641 tgtaacgttc accctctacc ttagcatccc ttccctttgc aaatagtcct cttccaacaa
     2701 taataatgtc agatcctgta gagaccacat catccacggt tctatactgt tgacccaatg
     2761 cgtctccctt gtcatctaaa cccacaccgg gtgtcataat caaccaatcg taaccttcat
     2821 ctcttccacc catgtctctt tgagcaataa agccgataac aaaatctttg tcgctcttcg
     2881 caatgtcaac agtaccctta gtatattctc cagtagatag ggagcccttg catgacaatt
     2941 ctgctaacat caaaaggcct ctaggttcct ttgttacttc ttctgccgcc tgcttcaaac
     3001 cgctaacaat acctgggccc accacaccgt gtgcattcgt aatgtctgcc cattctgcta
     3061 ttctgtatac acccgcagag tactgcaatt tgactgtatt accaatgtca gcaaattttc
     3121 tgtcttcgaa gagtaaaaaa ttgtacttgg cggataatgc ctttagcggc ttaactgtgc
     3181 cctccatgga aaaatcagtc aagatatcca catgtgtttt tagtaaacaa attttgggac
     3241 ctaatgcttc aactaactcc agtaattcct tggtggtacg aacatccaat gaagcacaca
     3301 agtttgtttg cttttcgtgc atgatattaa atagcttggc agcaacagga ctaggatgag
     3361 tagcagcacg ttccttatat gtagctttcg acatgattta tcttcgtttc ctgcatgttt
     3421 ttgttctgtg cagttgggtt aagaatactg ggcaatttca tgtttcttca acactacata
     3481 tgcgtatata taccaatcta agtctgtgct ccttccttcg ttcttccttc tgttcggaga
     3541 ttaccgaatc aaaaaaattt caaggaaacc gaaatcaaaa aaaagaataa aaaaaaaatg
     3601 atgaattgaa aagctggcct cgtgatacgc ctatttttat aggttaatgt catgataata
     3661 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     3721 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     3781 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     3841 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     3901 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     3961 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     4021 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     4081 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     4141 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     4201 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     4261 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     4321 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     4381 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     4441 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     4501 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     4561 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     4621 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     4681 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     4741 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     4801 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     4861 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     4921 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     4981 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     5041 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     5101 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     5161 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     5221 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     5281 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     5341 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     5401 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     5461 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     5521 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     5581 agcgagtc
//
