LOCUS       sequence-7991-ex        7567 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 7567)
  AUTHORS   Alberti S, Gitler AD, Lindquist S
  TITLE     A suite of Gateway cloning vectors for high-throughput genetic 
            analysis in Saccharomyces cerevisiae.
  JOURNAL   Yeast. 2007 Oct . 24(10):913-9.
  PUBMED    17583893
REFERENCE   2  (bases 1 to 7567)
  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: "Yeast. 2007
            Oct . 24(10):913-9."
COMMENT     Sequence Label: pAG304GPD-EYFP-ccdB
FEATURES             Location/Qualifiers
     source          1..7567
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        185..465
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     CDS             466..1140
                     /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"
     rep_origin      complement(1237..1692)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1374..1395)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1586..1605
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1822..1844
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1836..1853
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1837..1853
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1860..1879
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1860..1878
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        1903..2547
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     2441..2464
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     primer_bind     2553..2569
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2553..2569
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             join(2582..2584,2585..2587,2588..3298)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2627..2648)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2888..2907)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3235..3256
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    3323..3447
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(3472..3477,3478..3495,3496..3502)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             3556..4215
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3622..3641)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             4560..4865
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     4744..4763
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4906..5030)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     terminator      5037..5284
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(5053..5071)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(5307..5325)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5346..5362)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5346..5362)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5359..5381)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5370..5386
                     /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(5394..5400,5401..5418,5419..5424))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(5577..5594)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5748..6336)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6507..7298,7299..7367))
                     /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     7130..7149
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7368..7472)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7540..7558
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accaaacgac attactatat atataatata ggaagcattt aatagacagc atcgtaatat
      241 atgtgtactt tgcagttatg acgccagatg gcagtagtgg aagatattct ttattgaaaa
      301 atagcttgtc accttacgta caatcttgat ccggagcttt tctttttttg ccgattaaga
      361 attaattcgg tcgaaaaaag aaaaggagag ggccaagagg gagggcattg gtgactattg
      421 agcacgtgag tatacgtgat taagcacaca aaggcagctt ggagtatgtc tgttattaat
      481 ttcacaggta gttctggtcc attggtgaaa gtttgcggct tgcagagcac agaggccgca
      541 gaatgtgctc tagattccga tgctgacttg ctgggtatta tatgtgtgcc caatagaaag
      601 agaacaattg acccggttat tgcaaggaaa atttcaagtc ttgtaaaagc atataaaaat
      661 agttcaggca ctccgaaata cttggttggc gtgtttcgta atcaacctaa ggaggatgtt
      721 ttggctctgg tcaatgatta cggcattgat atcgtccaac tgcatggaga tgagtcgtgg
      781 caagaatacc aagagttcct cggtttgcca gttattaaaa gactcgtatt tccaaaagac
      841 tgcaacatac tactcagtgc agcttcacag aaacctcatt cgtttattcc cttgtttgat
      901 tcagaagcag gtgggacagg tgaacttttg gattggaact cgatttctga ctgggttgga
      961 aggcaagaga gccccgaaag cttacatttt atgttagctg gtggactgac gccagaaaat
     1021 gttggtgatg cgcttagatt aaatggcgtt attggtgttg atgtaagcgg aggtgtggag
     1081 acaaatggtg taaaagactc taacaaaata gcaaatttcg tcaaaaatgc taagaaatag
     1141 gttattactg agtagtattt atttaagtat tgtttgtgca cttgcctgcg gtgtgaaata
     1201 ccgcacagat gcgtaaggag aaaataccgc atcaggaaat tgtaaacgtt aatattttgt
     1261 taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag gccgaaatcg
     1321 gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt gttccagttt
     1381 ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga aaaaccgtct
     1441 atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg gggtcgaggt
     1501 gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct tgacggggaa
     1561 agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc gctagggcgc
     1621 tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc
     1681 tacagggcgc gtcgcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg
     1741 tgcgggcctc ttcgctatta cgccagctgg cgaagggggg atgtgctgca aggcgattaa
     1801 gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgaattgt
     1861 aatacgactc actatagggc gaattggagc tcttcgagtt tatcattatc aatactcgcc
     1921 atttcaaaga atacgtaaat aattaatagt agtgattttc ctaactttat ttagtcaaaa
     1981 aattagcctt ttaattctgc tgtaacccgt acatgcccaa aatagggggc gggttacaca
     2041 gaatatataa caccgaaggt gcctgggtga acaggtttat tcctggcatc cactaaatat
     2101 aatggagccc gctttttaag ctggcatcca gaaaaaaaaa gaatcccagc accaaaatat
     2161 tgttttcttc accaaccatc agttcatagg tccattctct tagcgcaact acagagaaca
     2221 ggggcacaaa caggcaaaaa acgggcacaa cctcaatgga gtgatgcaac ctgcctggag
     2281 taaatgatga cacaaggcaa ttgacccacg catgtatcta tctcattttc ttacaccttc
     2341 tattaccttc tgctctctct gatttggaaa aagctgaaaa aaaaggttga aaccagttcc
     2401 ctgaaattat tcccctactt gactaataag tatataaaga cggtaggtat tgattgtaat
     2461 tctgtaaatc tatttcttaa acttcttaaa ttctactttt atagttagtc ttttttttag
     2521 ttttaaaaca ccagaactta gtttcgacgg attctagaac tagtggatcc cccgggacaa
     2581 aatggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
     2641 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta
     2701 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
     2761 cctcgtgacc accttcggct acggcctgca gtgcttcgcc cgctaccccg accacatgaa
     2821 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt
     2881 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
     2941 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
     3001 caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa
     3061 cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc
     3121 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
     3181 ctacctgagc taccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt
     3241 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaaggg
     3301 tggaccaggt ggtggacatc aaacaagttt gtacaaaaaa gctgaacgag aaacgtaaaa
     3361 tgatataaat atcaatatat taaattagat tttgcataaa aaacagacta cataatactg
     3421 taaaacacaa catatccagt cactatggcg gccgcattag gcaccccagg ctttacactt
     3481 tatgcttccg gctcgtataa tgtgtggatt ttgagttagg atccgtcgag attttcagga
     3541 gctaaggaag ctaaaatgga gaaaaaaatc actggatata ccaccgttga tatatcccaa
     3601 tggcatcgta aagaacattt tgaggcattt cagtcagttg ctcaatgtac ctataaccag
     3661 accgttcagc tggatattac ggccttttta aagaccgtaa agaaaaataa gcacaagttt
     3721 tatccggcct ttattcacat tcttgcccgc ctgatgaatg ctcatccgga attccgtatg
     3781 gcaatgaaag acggtgagct ggtgatatgg gatagtgttc acccttgtta caccgttttc
     3841 catgagcaaa ctgaaacgtt ttcatcgctc tggagtgaat accacgacga tttccggcag
     3901 tttctacaca tatattcgca agatgtggcg tgttacggtg aaaacctggc ctatttccct
     3961 aaagggttta ttgagaatat gtttttcgtc tcagccaatc cctgggtgag tttcaccagt
     4021 tttgatttaa acgtggccaa tatggacaac ttcttcgccc ccgttttcac catgggcaaa
     4081 tattatacgc aaggcgacaa ggtgctgatg ccgctggcga ttcaggttca tcatgccgtc
     4141 tgtgatggct tccatgtcgg cagaatgctt aatgaattac aacagtactg cgatgagtgg
     4201 cagggcgggg cgtaaacgcc gcgtggatcc ggcttactaa aagccagata acagtatgcg
     4261 tatttgcgcg ctgatttttg cggtataaga atatatactg atatgtatac ccgaagtatg
     4321 tcaaaaagag gtatgctatg aagcagcgta ttacagtgac agttgacagc gacagctatc
     4381 agttgctcaa ggcatatatg atgtcaatat ctccggtctg gtaagcacaa ccatgcagaa
     4441 tgaagcccgt cgtctgcgtg ccgaacgctg gaaagcggaa aatcaggaag ggatggctga
     4501 ggtcgcccgg tttattgaaa tgaacggctc ttttgctgac gagaacaggg gctggtgaaa
     4561 tgcagtttaa ggtttacacc tataaaagag agagccgtta tcgtctgttt gtggatgtac
     4621 agagtgatat tattgacacg cccgggcgac ggatggtgat ccccctggcc agtgcacgtc
     4681 tgctgtcaga taaagtctcc cgtgaacttt acccggtggt gcatatcggg gatgaaagct
     4741 ggcgcatgat gaccaccgat atggccagtg tgccggtctc cgttatcggg gaagaagtgg
     4801 ctgatctcag ccaccgcgaa aatgacatca aaaacgccat taacctgatg ttctggggaa
     4861 tataaatgtc aggctccctt atacacagcc agtctgcagg tcgaccatag tgactggata
     4921 tgttgtgttt tacagtatta tgtagtctgt tttttatgca aaatctaatt taatatattg
     4981 atatttatat cattttacgt ttctcgttca gctttcttgt acaaagtggt tcgatgtcat
     5041 gtaattagtt atgtcacgct tacattcacg ccctcccccc acatccgctc taaccgaaaa
     5101 ggaaggagtt agacaacctg aagtctaggt ccctatttat ttttttatag ttatgttagt
     5161 attaagaacg ttatttatat ttcaaatttt tctttttttt ctgtacagac gcgtgtacgc
     5221 atgtaacatt atactgaaaa ccttgcttga gaaggttttg ggacgctcga aggctttaat
     5281 ttgcggccgg tacccagctt ttgttccctt tagtgagggt taattccgag cttggcgtaa
     5341 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     5401 ggagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgaggta actcacatta
     5461 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     5521 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     5581 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     5641 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     5701 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     5761 ggcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     5821 ggactataaa gataccaggc gttcccccct ggaagctccc tcgtgcgctc tcctgttccg
     5881 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     5941 caatgctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     6001 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     6061 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     6121 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     6181 actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     6241 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     6301 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     6361 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     6421 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     6481 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     6541 gcgatctgtc tatttcgttc atccatagtt gcctgactgc ccgtcgtgta gataactacg
     6601 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     6661 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     6721 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     6781 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     6841 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     6901 tgatccccca tgttgtgaaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     6961 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     7021 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     7081 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     7141 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     7201 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     7261 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     7321 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     7381 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     7441 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     7501 gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc
     7561 tttcgtc
//
