LOCUS       sequence-380550-        6755 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Low copy vector backbone with G418 selectivity, encodes for TDH3(pr)
            driven expression of mNeon.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6755)
  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 6755)
  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: pRSII41K-TDH3pr-mNeon
FEATURES             Location/Qualifiers
     source          1..6755
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(98..117)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      911..1499
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1400..1419
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1653..1670
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    1787..1808
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(1823..1828,1829..1846,1847..1853)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1861..1877
                     /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     1866..1888
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     1885..1901
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1885..1901
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1920..1940
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1922..1940
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        1965..2609
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     2502..2525
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     terminator      3343..3590
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3359..3377)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(3608..3627)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3609..3627)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3637..3654)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3637..3653)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3646..3668)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3794..4249
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3881..3900)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4091..4112
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     gene            complement(4337..5693)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     terminator      complement(4337..4534)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(4540..5349)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     5263..5282
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(5350..5693)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     primer_bind     complement(5752..5771)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5871..5893
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5931..5949)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    5990..6493
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        6530..6634
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6635..6703,6704..6755,1..740)
                     /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"
ORIGIN
        1 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
       61 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
      121 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
      181 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
      241 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
      301 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
      361 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
      421 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
      481 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
      541 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
      601 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
      661 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
      721 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
      781 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
      841 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
      901 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
      961 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     1021 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     1081 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     1141 cagtggctgc tgccagtggc gataagtcgt gtcttaccag gttggactca agacgatagt
     1201 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     1261 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     1321 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     1381 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     1441 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     1501 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     1561 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     1621 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     1681 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
     1741 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
     1801 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
     1861 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg
     1921 caattaaccc tcactaaagg gaacaaaagc tggagctcag tttatcatta tcaatactcg
     1981 ccatttcaaa gaatacgtaa ataattaata gtagtgattt tcctaacttt atttagtcaa
     2041 aaaattagcc ttttaattct gctgtaaccc gtacatgccc aaaatagggg gcgggttaca
     2101 cagaatatat aacatcgtag gtgtctgggt gaacagttta ttcctggcat ccactaaata
     2161 taatggagcc cgctttttaa gctggcatcc agaaaaaaaa agaatcccag caccaaaata
     2221 ttgttttctt caccaaccat cagttcatag gtccattctc ttagcgcaac tacagagaac
     2281 aggggcacaa acaggcaaaa aacgggcaca acctcaatgg agtgatgcaa cctgcctgga
     2341 gtaaatgatg acacaaggca attgacccac gcatgtatct atctcatttt cttacacctt
     2401 ctattacctt ctgctctctc tgatttggaa aaagctgaaa aaaaaggttg aaaccagttc
     2461 cctgaaatta ttcccctact tgactaataa gtatataaag acggtaggta ttgattgtaa
     2521 ttctgtaaat ctatttctta aacttcttaa attctacttt tatagttagt ctttttttta
     2581 gttttaaaac accaagaact tagtttcgac ggatctagaa ctagtatggt ctcaaaaggg
     2641 gaggaggaca acatggcatc cttgccagca actcacgaat tacatatttt cggaagtatt
     2701 aatggtgttg actttgatat ggttggacaa ggcactggta atccaaacga tggatacgag
     2761 gaacttaatc ttaaatcaac aaaaggcgat ttgcaatttt ccccctggat tcttgtgccc
     2821 catattgggt acggctttca tcaatattta ccttaccctg atggaatgag tccatttcaa
     2881 gccgctatgg tagacggttc tggctaccaa gttcatagaa ccatgcagtt tgaagacggc
     2941 gcttccttaa cagtaaacta caggtacacg tacgagggtt cccatattaa gggagaagcc
     3001 caagttaagg gtactggttt tccggctgat ggtcccgtta tgactaatag cctgacagct
     3061 gctgactggt gtaggtctaa aaagacatat cctaacgata aaacaataat ttcaactttt
     3121 aagtggtctt ataccactgg caatggtaaa agatacagat ccactgcccg tactacgtat
     3181 acattcgcta aaccaatggc cgctaattat cttaaaaatc aacccatgta cgtgtttagg
     3241 aagactgaac taaaacatag taagaccgag ttgaacttta aggaatggca aaaagccttc
     3301 accgatgtca tgggaatgga tgaattatat aagtaactcg agtcatgtaa ttagttatgt
     3361 cacgcttaca ttcacgccct ccccccacat ccgctctaac cgaaaaggaa ggagttagac
     3421 aacctgaagt ctaggtccct atttattttt ttatagttat gttagtatta agaacgttat
     3481 ttatatttca aatttttctt ttttttctgt acagacgcgt gtacgcatgt aacattatac
     3541 tgaaaacctt gcttgagaag gttttgggac gctcgaaggc tttaatttgc ggccggtacc
     3601 caattcgccc tatagtgagt cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg
     3661 tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc
     3721 cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct
     3781 gaatggcgaa tggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     3841 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     3901 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
     3961 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
     4021 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
     4081 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     4141 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     4201 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaatttc ctgatgcggt
     4261 attttctcct tacgcatctg tgcggtattt cacaccgcat agacactgga tggcggcgtt
     4321 agtatcgaat cgacagcagt atagcgacca gcattcacat acgattgacg catgatatta
     4381 ctttctgcgc acttaacttc gcatctgggc agatgatgtc gaggcgaaaa aaaatataaa
     4441 tcacgctaac atttgattaa aatagaacaa ctacaatata aaaaaactat acaaatgaca
     4501 agttcttgaa aacaagaatc tttttattgt cagtactgat tagaaaaact catcgagcat
     4561 caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg
     4621 tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta
     4681 tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa
     4741 aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa
     4801 aagcttatgc atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa
     4861 atcactcgca tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac
     4921 gcgatcgctg ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac
     4981 tgccagcgca tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc
     5041 tgttttgccg gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg
     5101 cttgatggtc ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt
     5161 aacatcattg gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt
     5221 cccatacaat cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata
     5281 cccatataaa tcagcatcca tgttggaatt taatcgcggc ctcgaaacgt gagtcttttc
     5341 cttacccatg gttgtttatg ttcggatgtg atgtgagaac tgtatcctag caagatttta
     5401 aaaggaagta tatgaaagaa gaacctcagt ggcaaatcct aaccttttat atttctctac
     5461 aggggcgcgg cgtggggaca attcaacgcg tctgtgaggg gagcgtttcc ctgctcgcag
     5521 gtctgcagcg aggagccgta atttttgctt cgcgccgtgc ggccatcaaa atgtatggat
     5581 gcaaatgatt atacatgggg atgtatgggc taaatgtacg ggcgacagtc acatcatgcc
     5641 cctgagctgc gcacgtcaag actgtcaagg agggtattct gggcctccat gtcgctggcc
     5701 gggtgacccg gcggggacga ggcaagctaa ggtggtatgg tgcactctca gtacaatctg
     5761 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
     5821 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     5881 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat
     5941 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgg atcgcttgcc
     6001 tgtaacttac acgcgcctcg tatcttttaa tgatggaata atttgggaat ttactctgtg
     6061 tttatttatt tttatgtttt gtatttggat tttagaaagt aaataaagaa ggtagaagag
     6121 ttacggaatg aagaaaaaaa aataaacaaa ggtttaaaaa atttcaacaa aaagcgtact
     6181 ttacatatat atttattaga caagaaaagc agattaaata gatatacatt cgattaacga
     6241 taagtaaaat gtaaaatcac aggattttcg tgtgtggtct tctacacaga caagatgaaa
     6301 caattcggca ttaatacctg agagcaggaa gagcaagata aaaggtagta tttgttggcg
     6361 atccccctag agtcttttac atcttcggaa aacaaaaact attttttctt taatttcttt
     6421 ttttactttc tatttttaat ttatatattt atattaaaaa atttaaatta taattatttt
     6481 tatagcacgt gatgaaaagg acccaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     6541 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     6601 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     6661 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     6721 gaaagtaaaa gatgctgaag atcagttggg tgcac
//
