LOCUS       sequence-8463-ex        5465 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 5465)
  AUTHORS   Nakashima N, Noguchi E, Nishimoto T
  TITLE     Saccharomyces cerevisiae putative G protein, Gtr1p, which forms 
            complexes with itself and a novel protein designated as Gtr2p, 
            negatively regulates the Ran/Gsp1p G protein cycle through Gtr2p.
  JOURNAL   Genetics. 1999 Jul . 152(3):853-67.
  PUBMED    10388807
REFERENCE   2  (bases 1 to 5465)
  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: "Genetics. 
            1999 Jul . 152(3):853-67."
COMMENT     Sequence Label: pAS404
FEATURES             Location/Qualifiers
     source          1..5465
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        96..200
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(201..269,270..1061)
                     /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(419..438)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1232..1820
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1721..1740
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1974..1991
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(2144..2149,2150..2167,2168..2174)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2182..2198
                     /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     2187..2209
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2206..2222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2206..2222
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2241..2261
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2243..2261
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2291..2995
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             3023..3463
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="DNA binding domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 DNA binding domain
                     /translation="MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTK
                     RSPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNK
                     DAVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVS"
     CDS             3479..3505
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3485..3511
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     primer_bind     complement(3580..3599)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3581..3599)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3609..3626)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3609..3625)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3618..3640)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3770..4225
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3857..3876)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4067..4088
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4324..4998)
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     promoter        complement(4999..5279)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(5296..5315)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5415..5437
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
      241 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
      301 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
      361 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
      421 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
      481 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
      541 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
      601 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
      661 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
      721 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
      781 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
      841 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
      901 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
      961 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     1021 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     1081 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     1141 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     1201 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     1261 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     1321 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     1381 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     1441 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     1501 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     1561 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     1621 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     1681 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     1741 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     1801 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     1861 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     1921 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     1981 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     2041 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     2101 acgcaattaa tgtgagttac ctcactcatt aggcacccca ggctttacac tttatgcttc
     2161 cggctcctat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     2221 accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctggagctcg
     2281 agatctagat atccttttgt tgtttccggg tgtacaatat ggacttcctc ttttctggca
     2341 accaaaccca tacatcggga ttcctataat accttcgttg gtctccctaa catgtaggtg
     2401 gcggagggga gatatacaat agaacagata ccagacaaga cataatgggc taaacaagac
     2461 tacaccaatt acactgcctc attgatggtg gtacataacg aactaatact gtagccctag
     2521 acttgatagc catcatcata tcgaagtttc actacccttt ttccatttgc catctattga
     2581 agtaataata ggcgcatgca acttcttttc tttttttttc ttttctctct cccccgttgt
     2641 tgtctcacca tatccgcaat gacaaaaaaa atgatggaag acactaaagg aaaaaattaa
     2701 cgacaaagac agcaccaaca gatgtcgttg ttccagagct gatgaggggt atcttcgaac
     2761 acacgaaact ttttccttcc ttcattcacg cacactactc tctaatgagc aacggtatac
     2821 ggccttcctt ccagttactt gaatttgaaa taaaaaaagt ttgccgcttt gctatcaagt
     2881 ataaatagac ctgcaattat taatcttttg tttcctcgtc attgttctcg ttccctttct
     2941 tccttgtttc tttttctgca caatatttca agctatacca agcatacaat caactccaag
     3001 cttgaagcaa gcctcctgaa agatgaagct actgtcttct atcgaacaag catgcgatat
     3061 ttgccgactt aaaaagctca agtgctccaa agaaaaaccg aagtgcgcca agtgtctgaa
     3121 gaacaactgg gagtgtcgct actctcccaa aaccaaaagg tctccgctga ctagggcaca
     3181 tctgacagaa gtggaatcaa ggctagaaag actggaacag ctatttctac tgatttttcc
     3241 tcgagaagac cttgacatga ttttgaaaat ggattcttta caggatataa aagcattgtt
     3301 aacaggatta tttgtacaag ataatgtgaa taaagatgcc gtcacagata gattggcttc
     3361 agtggagact gatatgcctc taacattgag acagcataga ataagtgcga catcatcatc
     3421 ggaagagagt agtaacaaag gtcaaagaca gttgactgta tcgccggaat tcatggctta
     3481 cccatacgat gttccagatt acgctagctt gggtggtcat atggccatgg aggccccggg
     3541 gatccgtcga cctcgagggg gggcccggta cccaattcgc cctatagtga gtcgtattac
     3601 gcgcgctcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
     3661 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
     3721 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgcga cgcgccctgt
     3781 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     3841 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     3901 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     3961 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     4021 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     4081 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     4141 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     4201 aacaaaatat taacgtttac aatttcctga tgcggtattt tctccttacg catctgtgcg
     4261 gtatttcaca ccgcataggc aagtgcacaa acaatactta aataaatact actcagtaat
     4321 aacctatttc ttagcatttt tgacgaaatt tgctattttg ttagagtctt ttacaccatt
     4381 tgtctccaca cctccgctta catcaacacc aataacgcca tttaatctaa gcgcatcacc
     4441 aacattttct ggcgtcagtc caccagctaa cataaaatgt aagctttcgg ggctctcttg
     4501 ccttccaacc cagtcagaaa tcgagttcca atccaaaagt tcacctgtcc cacctgcttc
     4561 tgaatcaaac aagggaataa acgaatgagg tttctgtgaa gctgcactga gtagtatgtt
     4621 gcagtctttt ggaaatacga gtcttttaat aactggcaaa ccgaggaact cttggtattc
     4681 ttgccacgac tcatctccat gcagttggac gatatcaatg ccgtaatcat tgaccagagc
     4741 caaaacatcc tccttaggtt gattacgaaa cacgccaacc aagtatttcg gagtgcctga
     4801 actattttta tatgctttta caagacttga aattttcctt gcaataaccg ggtcaattgt
     4861 tctctttcta ttgggcacac atataatacc cagcaagtca gcatcggaat ctagagcaca
     4921 ttctgcggcc tctgtgctct gcaagccgca aactttcacc aatggaccag aactacctgt
     4981 gaaattaata acagacatac tccaagctgc ctttgtgtgc ttaatcacgt atactcacgt
     5041 gctcaatagt caccaatgcc ctccctcttg gccctctcct tttctttttt cgaccgaatt
     5101 aattcttaat cggcaaaaaa agaaaagctc cggatcaaga ttgtacgtaa ggtgacaagc
     5161 tatttttcaa taaagaatat cttccactac tgccatctgg cgtcataact gcaaagtaca
     5221 catatattac gatgctgtct attaaatgct tcctatatta tatatatagt aatgtcgttt
     5281 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     5341 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     5401 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     5461 cgcga
//
