LOCUS       sequence-380542-        6928 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  High copy vector backbone with G418 selectivity, contains 
            TDH3(pr)-MCS-CYC1(3'UTR).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6928)
  AUTHORS   Smith JR, Sislak CD, Fernandez Mendoza P, Carmichael L, Lewis AG, 
            Chen A, Jiang GZ, Gibney PA
  TITLE     Auxotrophy-Independent Plasmid Shuttle Vectors for Applications in 
            Diverse Yeasts
  JOURNAL   Appl Microbiol. 2024; 4(1):453-469. doi: 
            10.3390/applmicrobiol4010031
REFERENCE   2  (bases 1 to 6928)
  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: "Appl 
            Microbiol."; date: "2024"; volume: " 4(1):453-469. doi"; pages: " 
            10.3390/applmicrobiol401003"
COMMENT     Sequence Label: pRSII42K-TDH3pr
FEATURES             Location/Qualifiers
     source          1..6928
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      683..880
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     rep_origin      complement(968..1423)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1105..1126)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1317..1336
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1549..1571
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1563..1580
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1564..1580
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1590..1609
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1590..1608
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      complement(1627..1874)
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     1840..1858
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     1876..1892
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1877..1893
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1927..1943)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1927..1943)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(1948..2592)
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     complement(2032..2055)
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     primer_bind     complement(2617..2637)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(2617..2635)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(2656..2672)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2656..2672)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2669..2691)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2680..2696
                     /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(2704..2710,2711..2728,2729..2734))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2749..2770
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2887..2904)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3058..3646)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3138..3157)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3817..4608,4609..4677))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQRFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4440..4459
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4678..4782)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4809..6155
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     primer_bind     6196..6214
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6252..6274)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     6374..6393
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     gene            join(6452..6928,1..880)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        6452..6795
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             join(6796..6928,1..677)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPICNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGDFFPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(6863..6882)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
ORIGIN
        1 ccgatgcgcc agagttgttt ctgaaacatg gcaaaggtag cgttgccaat gatgttacag
       61 atgagatggt cagactaaac tggctgacgg aatttatgcc tcttccgacc atcaagcatt
      121 ttatccgtac tcctgatgat gcatggttac tcaccactgc gatccccggc aaaacagcat
      181 tccaggtatt agaagaatat cctgattcag gtgaaaatat tgttgatgcg ctggcagtgt
      241 tcctgcgccg gttgcattcg attcctattt gtaattgtcc ttttaacagc gatcgcgtat
      301 ttcgtctcgc tcaggcgcaa tcacgaatga ataacggttt ggttgatgcg agtgattttg
      361 atgacgagcg taatggctgg cctgttgaac aagtctggaa agaaatgcat aagcttttgc
      421 cattctcacc ggattcagtc gtcactcatg gtgatttctc acttgataac cttatttttg
      481 acgaggggaa attaataggt tgtattgatg ttggacgagt cggaatcgca gaccgatacc
      541 aggatcttgc catcctatgg aactgcctcg gtgatttttt tccttcatta cagaaacggc
      601 tttttcaaaa atatggtatt gataatcctg atatgaataa attgcagttt catttgatgc
      661 ttgatgagtt tttctaatca gtactgacaa taaaaagatt cttgttttca agaacttgtc
      721 atttgtatag tttttttata ttgtagttgt tctattttaa tcaaatgtta gcgtgattta
      781 tatttttttt cgcctcgaca tcatctgccc agatgcgaag ttaagtgcgc agaaagtaat
      841 atcatgcgtc aatcgtatgt gaatgctggt cgctatactg ctgtcgattc gatactaacg
      901 ccgccatcca gtgtctatgc ggtgtgaaat accgcacaga tgcgtaagga gaaaataccg
      961 catcaggaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
     1021 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
     1081 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
     1141 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
     1201 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
     1261 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
     1321 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
     1381 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtccattcg ccattcaggc
     1441 tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga
     1501 aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac
     1561 gttgtaaaac gacggccagt gagcgcgcgt aatacgactc actatagggc gaattgggta
     1621 ccggccgcaa attaaagcct tcgagcgtcc caaaaccttc tcaagcaagg ttttcagtat
     1681 aatgttacat gcgtacacgc gtctgtacag aaaaaaaaga aaaatttgaa atataaataa
     1741 cgttcttaat actaacataa ctataaaaaa ataaataggg acctagactt caggttgtct
     1801 aactccttcc ttttcggtta gagcggatgt ggggggaggg cgtgaatgta agcgtgacat
     1861 aactaattac atgactcgag gtcgacggta tcgataagct tgatatcgaa ttcctgcagc
     1921 ccgggggatc cactagttct agatccgtcg aaactaagtt cttggtgttt taaaactaaa
     1981 aaaaagacta actataaaag tagaatttaa gaagtttaag aaatagattt acagaattac
     2041 aatcaatacc taccgtcttt atatacttat tagtcaagta ggggaataat ttcagggaac
     2101 tggtttcaac cttttttttc agctttttcc aaatcagaga gagcagaagg taatagaagg
     2161 tgtaagaaaa tgagatagat acatgcgtgg gtcaattgcc ttgtgtcatc atttactcca
     2221 ggcaggttgc atcactccat tgaggttgtg cccgtttttt gcctgtttgt gcccctgttc
     2281 tctgtagttg cgctaagaga atggacctat gaactgatgg ttggtgaaga aaacaatatt
     2341 ttggtgctgg gattcttttt ttttctggat gccagcttaa aaagcgggct ccattatatt
     2401 tagtggatgc caggaataaa ctgttcaccc agacacctac gatgttatat attctgtgta
     2461 acccgccccc tattttgggc atgtacgggt tacagcagaa ttaaaaggct aattttttga
     2521 ctaaataaag ttaggaaaat cactactatt aattatttac gtattctttg aaatggcgag
     2581 tattgataat gataaactga gctccagctt ttgttccctt tagtgagggt taattgcgcg
     2641 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     2701 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     2761 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     2821 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     2881 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     2941 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     3001 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     3061 ccataggctc cgcccccctg acgagcatca caaaaatgga cgctcaagtc agaggtggcg
     3121 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3181 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     3241 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     3301 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     3361 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     3421 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     3481 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     3541 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     3601 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     3661 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     3721 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     3781 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     3841 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     3901 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     3961 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     4021 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     4081 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     4141 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggctccc aacgatcaag
     4201 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     4261 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     4321 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     4381 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     4441 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     4501 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     4561 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     4621 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaacgttgaa tactcatact
     4681 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     4741 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     4801 gccacctgaa cgaagcatct gtgcttcatt ttgtagaaca aaaatgcaac gcgagagcgc
     4861 taatttttca aacaaagaat ctgagctgca tttttacaga acagaaatgc aacgcgaaag
     4921 cgctatttta ccaacgaaga atctgtgctt catttttgta aaacaaaaat gcaacgcgag
     4981 agcgctaatt tttcaaacaa agaatctgag ctgcattttt acagaacaga aatgcaacgc
     5041 gagagcgcta ttttaccaac aaagaatcta tacttctttt ttgttctaca aaaatgcatc
     5101 ccgagagcgc tatttttcta acaaagcatc ttagattact ttttttctcc tttgtgcgct
     5161 ctataatgca gtctcttgat aactttttgc actgtaggtc cgttaaggtt agaagaaggc
     5221 tactttggtg tctattttct cttccataaa aaaagcctga ctccacttcc cgcgtttact
     5281 gattactagc gaagctgcgg gtgcattttt tcaagataaa ggcatccccg attatattct
     5341 ataccgatgt ggattgcgca tactttgtga acagaaagtg atagcgttga tgattcttca
     5401 ttggtcagaa aattatgaac ggtttcttct attttgtctc tatatactac gtataggaaa
     5461 tgtttacatt ttcgtattgt tttcgattca ctctatgaat agttcttact acaatttttt
     5521 tgtctaaaga gtaatactag agataaacat aaaaaatgta gaggtcgagt ttagatgcaa
     5581 gttcaaggag cgaaaggtgg atgggtaggt tatataggga tatagcacag agatatatag
     5641 caaagagata cttttgagca atgtttgtgg aagcggtatt cgcaatattt tagtagctcg
     5701 ttacagtccg gtgcgttttt ggttttttga aagtgcgtct tcagagcgct tttggttttc
     5761 aaaagcgctc tgaagttcct atactttcta gctagagaat aggaacttcg gaataggaac
     5821 ttcaaagcgt ttccgaaaac gagcgcttcc gaaaatgcaa cgcgagctgc gcacatacag
     5881 ctcactgttc acgtcgcacc tatatctgcg tgttgcctgt atatatatat acgtgagaag
     5941 aacggcatag tgcgtgttta tgcttaaatg cgtacttata tgcgtctatt tatgtaggat
     6001 gaaaggtagt ctagtacctc ctgtgatatt atcccattcc atgcggggta tcgtatgctt
     6061 ccttcagcac taccctttag ctgttctata tgctgccact cctcaattgg attagtctca
     6121 tccttcaatg ctatcatttc ctttgatatt ggatcatact aagaaaccat tattatcatg
     6181 acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat
     6241 gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg
     6301 gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc
     6361 tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccatac caccttagct
     6421 tgcctcgtcc ccgccgggtc acccggccag cgacatggag gcccagaata ccctccttga
     6481 cagtcttgac gtgcgcagct caggggcatg atgtgactgt cgcccgtaca tttagcccat
     6541 acatccccat gtataatcat ttgcatccat acattttgat ggccgcacgg cgcgaagcaa
     6601 aaattacggc tcctcgctgc agacctgcga gcagggaaac gctcccctca cagacgcgtt
     6661 gaattgtccc cacgccgcgc ccctgtagag aaatataaaa ggttaggatt tgccactgag
     6721 gttcttcttt catatacttc cttttaaaat cttgctagga tacagttctc acatcacatc
     6781 cgaacataaa caaccatggg taaggaaaag actcacgttt cgaggccgcg attaaattcc
     6841 aacatggatg ctgatttata tgggtataaa tgggctcgcg ataatgtcgg gcaatcaggt
     6901 gcgacaatct atcgattgta tgggaagc
//
