LOCUS       sequence-199974-        5965 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integrating construct for expressing GFP-Snc1(L96V) from the TPI1 
            promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5965)
  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 5965)
  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: pRS406-GFP-SNC1(L96V)
FEATURES             Location/Qualifiers
     source          1..5965
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(144..149,150..167,168..174)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    182..198
                     /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     187..209
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     206..222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     206..222
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     241..261
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        243..261
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        306..717
                     /gene="S. cerevisiae TPI1"
                     /label=TPI1 promoter
                     /note="strong constitutive promoter for yeast triose 
                     phosphate isomerase"
     CDS             728..1441
                     /codon_start=1
                     /gene="mgfp5"
                     /product="GFP with folding enhancement mutations"
                     /label=mgfp5
                     /note="suitable for expression in plants due to removal of 
                     a cryptic intron"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKA
                     NFKTRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(758..786)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     1381..1401
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     complement(2003..2020)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2003..2019)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2012..2034)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2160..2615
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2247..2266)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2457..2478
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(2746..3549)
                     /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(3550..3770)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(3796..3815)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3915..3937
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3975..3993)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4061..4165
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4166..4234,4235..5026)
                     /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(4384..4403)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5197..5785
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5686..5705
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5939..5956
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
       61 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttac
      121 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcctat gttgtgtgga
      181 attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta cgccaagcgc
      241 gcaattaacc ctcactaaag ggaacaaaag ctggagctcc accgcggtgg cggccccccc
      301 tcgagccatc aggttggtgg aagattaccc gttctaagac ttttcagctt cctctattga
      361 tgttacacct ggacacccct tttctggcat ccagttttta atcttcagtg gcatgtgaga
      421 ttctccgaaa ttaattaaag caatcacaca attctctcgg ataccacctc ggttgaaact
      481 gacaggtggt ttgttacgca tgctaatgca aaggagccta tatacctttg gctcggctgc
      541 tgtaacaggg aatataaagg gcagcataat ttaggagttt agtgaacttg caacatttac
      601 tatttttcct tcttacgtaa atatttttct ttttaattct aaatcaatct ttttcaattt
      661 tttgtttgta ttcttttctt gcttaaatct ataactacaa aaaacacata cataaacaag
      721 ctagcttatg agtaaaggag aagaactttt cactggagtt gtcccaattc ttgttgaatt
      781 agatggtgat gttaatggac acaaattttc tgtcagtgga gagggtgaag gtgatgcaac
      841 atacggaaaa cttaccctta aatttatttg cactactgga aaactacctg ttccatggcc
      901 aacacttgtc actactttct cttatggtgt tcaatgcttt tcaagatacc cagatcatat
      961 gaagcggcac gacttcttca agagcgccat gcctgaggga tacgtgcagg agaggaccat
     1021 cttcttcaag gacgacggga actacaagac acgtgctgaa gtcaagtttg agggagacac
     1081 cctcgtcaac aggatcgagc ttaagggaat cgatttcaag gaggacggaa acatcctcgg
     1141 ccacaagttg gaatacaact acaactccca caacgtatac atcatggccg acaaacaaaa
     1201 gaatggaatc aaagctaact tcaaaactag acacaacatt gaagatggag gcgttcaact
     1261 agcagaccat tatcaacaaa atactccaat tggcgatggc cctgtccttt taccagacaa
     1321 ccattacctg tccacacaat ctgccctttc gaaagatccc aacgaaaaga gagaccacat
     1381 ggtccttctt gagtttgtaa cagctgctgg gattacacat ggcatggatg aactatacaa
     1441 atcgaattcc atgtcgtcat ctactccctt tgacccttat gctctatccg agcacgatga
     1501 agaacgaccc cagaatgtac agtctaagtc aaggactgcg gaactgcagg ctgaaattga
     1561 tgataccgtg ggaataatga gagataacat aaataaagta gctgagcgag gtgaaagatt
     1621 aacgtccatt gaagataaag ccgataacct agcggtctca gcccaaggct ttaagagggg
     1681 tgccaatagg gtcagaaaag ccatgtggta caaggatcta aaaatgaaga tgtgtgtggc
     1741 tttagtaatc atcatattgc ttgttgtaat catcgtcccc attgctgttc actttagtcg
     1801 atagaagttc actcgcaatg ctatatatat agggagcttc catatgtagt gtaggatata
     1861 tgtatatata catatataca tatatgggcg tatatttact ttgttcttat tttctgtcta
     1921 attttataat tttactgaca gtagctaagc cctctgtatt gctgggatcc ccatggattc
     1981 aaagccgata ccaaacgcgc tcactggccg tcgttttaca acgtcgtgac tgggaaaacc
     2041 ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata
     2101 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatgga
     2161 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2221 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2281 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     2341 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     2401 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     2461 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     2521 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     2581 cgcgaatttt aacaaaatat taacgtttac aatttcctga tgcggtattt tctccttacg
     2641 catctgtgcg gtatttcaca ccgcataggg taataactga tataattaaa ttgaagctct
     2701 aatttgtgag tttagtatac atgcatttac ttataataca gttttttagt tttgctggcc
     2761 gcatcttctc aaatatgctt cccagcctgc ttttctgtaa cgttcaccct ctaccttagc
     2821 atcccttccc tttgcaaata gtcctcttcc aacaataata atgtcagatc ctgtagagac
     2881 cacatcatcc acggttctat actgttgacc caatgcgtct cccttgtcat ctaaacccac
     2941 accgggtgtc ataatcaacc aatcgtaacc ttcatctctt ccacccatgt ctctttgagc
     3001 aataaagccg ataacaaaat ctttgtcgct cttcgcaatg tcaacagtac ccttagtata
     3061 ttctccagta gatagggagc ccttgcatga caattctgct aacatcaaaa ggcctctagg
     3121 ttcctttgtt acttcttctg ccgcctgctt caaaccgcta acaatacctg ggcccaccac
     3181 accgtgtgca ttcgtaatgt ctgcccattc tgctattctg tatacacccg cagagtactg
     3241 caatttgact gtattaccaa tgtcagcaaa ttttctgtct tcgaagagta aaaaattgta
     3301 cttggcggat aatgccttta gcggcttaac tgtgccctcc atggaaaaat cagtcaagat
     3361 atccacatgt gtttttagta aacaaatttt gggacctaat gcttcaacta actccagtaa
     3421 ttccttggtg gtacgaacat ccaatgaagc acacaagttt gtttgctttt cgtgcatgat
     3481 attaaatagc ttggcagcaa caggactagg atgagtagca gcacgttcct tatatgtagc
     3541 tttcgacatg atttatcttc gtttcctgca ggtttttgtt ctgtgcagtt gggttaagaa
     3601 tactgggcaa tttcatgttt cttcaacact acatatgcgt atatatacca atctaagtct
     3661 gtgctccttc cttcgttctt ccttctgttc ggagattacc gaatcaaaaa aatttcaaag
     3721 aaaccgaaat caaaaaaaag aataaaaaaa aaatgatgaa ttgaattgaa aagctgtggt
     3781 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     3841 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     3901 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     3961 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     4021 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     4081 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     4141 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     4201 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     4261 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     4321 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     4381 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     4441 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     4501 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     4561 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     4621 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     4681 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     4741 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     4801 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     4861 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     4921 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     4981 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     5041 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     5101 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     5161 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     5221 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     5281 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     5341 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     5401 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     5461 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     5521 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     5581 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     5641 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     5701 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     5761 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     5821 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa
     5881 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     5941 cgagtcagtg agcgaggaag cggaa
//
