LOCUS       sequence-64348-e        6862 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6862)
  TITLE     Yeast Secretion Vectors
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6862)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pRS316-4M5.3
FEATURES             Location/Qualifiers
     source          1..6862
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    69..571
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        608..712
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(713..781,782..1573)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(931..950)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1744..2332
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2486..2503
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(2656..2661,2662..2679,2680..2686)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2694..2710
                     /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     2699..2721
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2718..2734
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2718..2734
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        2755..2773
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2827..2845
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     2877..2893
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     2878..2894
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2922..3586
                     /gene="GAL1 and GAL10 genes of S. cerevisiae"
                     /label=GAL1,10 promoter
                     /note="divergent inducible promoter, regulated by Gal4"
     protein_bind    3138..3255
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     3550..3573
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             3721..3744
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(4878..4897)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4879..4897)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4904..4921)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4904..4920)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4913..4935)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5065..5520
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5152..5171)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5362..5383
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5649..6452)
                     /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(6453..6673)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(6693..6712)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6812..6834
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgga tcgcttgcct gtaacttaca cgcgcctcgt atcttttaat gatggaataa
      121 tttgggaatt tactctgtgt ttatttattt ttatgttttg tatttggatt ttagaaagta
      181 aataaagaag gtagaagagt tacggaatga agaaaaaaaa ataaacaaag gtttaaaaaa
      241 tttcaacaaa aagcgtactt tacatatata tttattagac aagaaaagca gattaaatag
      301 atatacattc gattaacgat aagtaaaatg taaaatcaca ggattttcgt gtgtggtctt
      361 ctacacagac aagatgaaac aattcggcat taatacctga gagcaggaag agcaagataa
      421 aaggtagtat ttgttggcga tccccctaga gtcttttaca tcttcggaaa acaaaaacta
      481 ttttttcttt aatttctttt ttactttcta tttttaattt atatatttat attaaaaaat
      541 ttaaattata attattttta tagcacgtga tgaaaaggac ccaggtggca cttttcgggg
      601 aaatgtgcgc gaaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
      661 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
      721 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
      781 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
      841 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
      901 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
      961 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     1021 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     1081 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     1141 gaaggagcta accgcttttt ttcacaacat gggggatcat gtaactcgcc ttgatcgttg
     1201 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     1261 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     1321 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     1381 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     1441 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     1501 cagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     1561 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     1621 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     1681 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     1741 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     1801 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     1861 cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca
     1921 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     1981 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     2041 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     2101 gacctacacc gaactgagat acctacagcg tgagcattga gaaagcgcca cgcttcccga
     2161 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     2221 ggagcttcca ggggggaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     2281 acttgagcgt cgatttttgt gatgctcgtc aggggggccg agcctatgga aaaacgccag
     2341 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     2401 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     2461 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
     2521 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag
     2581 gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt acctcactca
     2641 ttaggcaccc caggctttac actttatgct tccggctcct atgttgtgtg gaattgtgag
     2701 cggataacaa tttcacacag gaaacagcta tgaccatgat tacgccaagc tcggaattaa
     2761 ccctcactaa agggaacaaa agctgtttcc acggaacagc tatgaccatg attacgccaa
     2821 gctcgaaatt aaccctcact aaagggaaca aaagctggta ccgggccccc cctcgatcga
     2881 ggtcgacggt atcgataagc ttgatatcga attccttgaa ttttcaaaaa ttcttacttt
     2941 ttttttggat ggacgcaaag aagtttaata atcatattac atggcattac caccatatac
     3001 atatccatat acatatccat atctaatctt acttatatgt tgtggaaatg taaagagccc
     3061 cattatctta gcctaaaaaa accttctctt tggaactttc agtaatacgc ttaactgctc
     3121 attgctatat tgaagtacgg attagaagcc gccgagcggg tgacagccct ccgaaggaag
     3181 actctcctcc gtgcgtcctc gtcttcaccg gtcgcgttcc tgaaacgcag atgtgcctcg
     3241 cgccgcactg ctccgaacaa taaagattct acaatactag cttttatggt tatgaagagg
     3301 aaaaattggc agtaacctgg ccccacaaac cttcaaatga acgaatcaaa ttaacaacca
     3361 taggatgata atgcgattag ttttttagcc ttatttctgg ggtaattaat cagcgaagcg
     3421 atgatttttg atctattaac agatatataa atgcaaaaac tgcataacca ctttaactaa
     3481 tactttcaac attttcggtt tgtattactt cttattcaaa tgtaataaaa gtatcaacaa
     3541 aaaattgtta atatacctct atactttaac gtcaaggaga aaaaaccccg gatcatgaag
     3601 gttttgattg tcttgttggc tatcttcgct gctttgccct tggctctagc tcaaccggtt
     3661 atttctacta ccgtcggttc cgctgcagaa ggctctttgg acaagagaga agctcggccg
     3721 gactacaagg acgacgatga caaggctagc gacgtcgtta tgactcaaac accactatca
     3781 cttcctgtta gtctaggtga tcaagcctcc atctcttgca gatctagtca gagcctcgta
     3841 cacagtaatg gaaacaccta tttacgttgg tacctgcaga agccaggcca gtctccaaag
     3901 gtcctgatct acaaagtttc caaccgagtt tctggggtcc cagacaggtt cagtggcagt
     3961 ggatcaggga cagatttcac actcaagatc aacagagtgg aggctgagga tctgggagtt
     4021 tatttctgct ctcaaagtac acatgttccg tggacgttcg gtggaggcac caagcttgaa
     4081 attaagtcct ctgctgatga tgctaagaag gatgctgcta agaaggatga tgctaagaaa
     4141 gatgatgcta agaaagatgg tggcgtcaaa ctggatgaga ctggaggagg cttggtgcaa
     4201 cctggggggg ccatgaaact ctcctgtgtt acctctggat tcacttttgg tcactactgg
     4261 atgaactggg tccgccagtc tccagagaaa ggactggagt gggtagcaca atttagaaac
     4321 aaaccttata attatgaaac atattattca gattctgtga aaggcagatt caccatctca
     4381 agagatgatt ccaaaagtag tgtctatctg caaatgaaca acttaagagt tgaagacacg
     4441 ggtatctatt actgtacggg tgcttcctat ggtatggaat acttgggtca aggaacctca
     4501 gtcaccgtct cctgataaca aaagcttatt tctgaagaag acttgtaata gctcgagatc
     4561 tgataacaac agtgtagatg taacaaaatc gactttgttc ccactgtact tttagctcgt
     4621 acaaaataca atatactttt catttctccg taaacaacat gttttcccat gtaatatcct
     4681 tttctatttt tcgttccgtt accaacttta cacatacttt atatagctat tcacttctat
     4741 acactaaaaa actaagacaa ttttaatttt gctgcctgcc atatttcaat ttgttataaa
     4801 ttcctataat ttatcctatt agtagctaaa aaaagatgaa tgtgaatcga atcctaagag
     4861 aattgagctc caattcgccc tatagtgagt cgtattacaa ttcactggcc gtcgttttac
     4921 aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc
     4981 ccttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc
     5041 gcagcctgaa tggcgaatgg cgcgacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     5101 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     5161 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     5221 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     5281 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     5341 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     5401 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     5461 atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg tttacaattt
     5521 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca gggtaataac
     5581 tgatataatt aaattgaagc tctaatttgt gagtttagta tacatgcatt tacttataat
     5641 acagtttttt agttttgctg gccgcatctt ctcaaatatg cttcccagcc tgcttttctg
     5701 taacgttcac cctctacctt agcatccctt ccctttgcaa atagtcctct tccaacaata
     5761 ataatgtcag atcctgtaga gaccacatca tccacggttc tatactgttg acccaatgcg
     5821 tctcccttgt catctaaacc cacaccgggt gtcataatca accaatcgta accttcatct
     5881 cttccaccca tgtctctttg agcaataaag ccgataacaa aatctttgtc gctcttcgca
     5941 atgtcaacag tacccttagt atattctcca gtagataggg agcccttgca tgacaattct
     6001 gctaacatca aaaggcctct aggttccttt gttacttctt ctgccgcctg cttcaaaccg
     6061 ctaacaatac ctgggcccac cacaccgtgt gcattcgtaa tgtctgccca ttctgctatt
     6121 ctgtatacac ccgcagagta ctgcaatttg actgtattac caatgtcagc aaattttctg
     6181 tcttcgaaga gtaaaaaatt gtacttggcg gataatgcct ttagcggctt aactgtgccc
     6241 tccatggaaa aatcagtcaa gatatccaca tgtgttttta gtaaacaaat tttgggacct
     6301 aatgcttcaa ctaactccag taattccttg gtggtacgaa catccaatga agcacacaag
     6361 tttgtttgct tttcgtgcat gatattaaat agcttggcag caacaggact aggatgagta
     6421 gcagcacgtt ccttatatgt agctttcgac atgatttatc ttcgtttcct gcaggttttt
     6481 gttctgtgca gttgggttaa gaatactggg caatttcatg tttcttcaac actacatatg
     6541 cgtatatata ccaatctaag tctgtgctcc ttccttcgtt cttccttctg ttcggagatt
     6601 accgaatcaa aaaaatttca aagaaaccga aatcaaaaaa aagaataaaa aaaaaatgat
     6661 gaattgaatt gaaaagcgtg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     6721 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     6781 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     6841 ccgtcatcac cgaaacgcgc ga
//
