LOCUS       sequence-7894-ex        6764 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 6764)
  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 6764)
  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: pAG304GAL-ccdB-HA
FEATURES             Location/Qualifiers
     source          1..6764
                     /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        1898..2339
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    1898..2015
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     2310..2333
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     2354..2370
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2354..2370
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    2378..2502
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2527..2532,2533..2550,2551..2557)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2611..3270
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2677..2696)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3615..3920
                     /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     3799..3818
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(3961..4085)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             4126..4215
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     terminator      4234..4481
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4250..4268)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(4504..4522)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4543..4559)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4543..4559)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4556..4578)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4567..4583
                     /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(4591..4597,4598..4615,4616..4621))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(4774..4791)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4945..5533)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(5704..6495,6496..6564))
                     /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     6327..6346
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6565..6669)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6737..6755
                     /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 tctagtacgg attagaagcc gccgagcggg
     1921 cgacagccct ccgacggaag actctcctcc gtgcgtcctc gtcttcaccg gtcgcgttcc
     1981 tgaaacgcag atgtgcctcg cgccgcactg ctccgaacaa taaagattct acaatactag
     2041 cttttatggt tatgaagagg aaaaattggc agtaacctgg ccccacaaac cttcaaatta
     2101 acgaatcaaa ttaacaacca taggatgata atgcgattag ttttttagcc ttatttctgg
     2161 ggtaattaat cagcgaagcg atgatttttg atctattaac agatatataa atggaaaagc
     2221 tgcataacca ctttaactaa tactttcaac attttcagtt tgtattactt cttattcaaa
     2281 tgtcataaaa gtatcaacaa aaaattgtta atatacctct atactttaac gtcaaggaga
     2341 aaaaaccccg gattctagaa ctagtggatc ccccatcaca agtttgtaca aaaaagctga
     2401 acgagaaacg taaaatgata taaatatcaa tatattaaat tagattttgc ataaaaaaca
     2461 gactacataa tactgtaaaa cacaacatat ccagtcacta tggcggccgc attaggcacc
     2521 ccaggcttta cactttatgc ttccggctcg tataatgtgt ggattttgag ttaggatccg
     2581 tcgagatttt caggagctaa ggaagctaaa atggagaaaa aaatcactgg atataccacc
     2641 gttgatatat cccaatggca tcgtaaagaa cattttgagg catttcagtc agttgctcaa
     2701 tgtacctata accagaccgt tcagctggat attacggcct ttttaaagac cgtaaagaaa
     2761 aataagcaca agttttatcc ggcctttatt cacattcttg cccgcctgat gaatgctcat
     2821 ccggaattcc gtatggcaat gaaagacggt gagctggtga tatgggatag tgttcaccct
     2881 tgttacaccg ttttccatga gcaaactgaa acgttttcat cgctctggag tgaataccac
     2941 gacgatttcc ggcagtttct acacatatat tcgcaagatg tggcgtgtta cggtgaaaac
     3001 ctggcctatt tccctaaagg gtttattgag aatatgtttt tcgtctcagc caatccctgg
     3061 gtgagtttca ccagttttga tttaaacgtg gccaatatgg acaacttctt cgcccccgtt
     3121 ttcaccatgg gcaaatatta tacgcaaggc gacaaggtgc tgatgccgct ggcgattcag
     3181 gttcatcatg ccgtctgtga tggcttccat gtcggcagaa tgcttaatga attacaacag
     3241 tactgcgatg agtggcaggg cggggcgtaa acgccgcgtg gatccggctt actaaaagcc
     3301 agataacagt atgcgtattt gcgcgctgat ttttgcggta taagaatata tactgatatg
     3361 tatacccgaa gtatgtcaaa aagaggtatg ctatgaagca gcgtattaca gtgacagttg
     3421 acagcgacag ctatcagttg ctcaaggcat atatgatgtc aatatctccg gtctggtaag
     3481 cacaaccatg cagaatgaag cccgtcgtct gcgtgccgaa cgctggaaag cggaaaatca
     3541 ggaagggatg gctgaggtcg cccggtttat tgaaatgaac ggctcttttg ctgacgagaa
     3601 caggggctgg tgaaatgcag tttaaggttt acacctataa aagagagagc cgttatcgtc
     3661 tgtttgtgga tgtacagagt gatattattg acacgcccgg gcgacggatg gtgatccccc
     3721 tggccagtgc acgtctgctg tcagataaag tctcccgtga actttacccg gtggtgcata
     3781 tcggggatga aagctggcgc atgatgacca ccgatatggc cagtgtgccg gtctccgtta
     3841 tcggggaaga agtggctgat ctcagccacc gcgaaaatga catcaaaaac gccattaacc
     3901 tgatgttctg gggaatataa atgtcaggct cccttataca cagccagtct gcaggtcgac
     3961 catagtgact ggatatgttg tgttttacag tattatgtag tctgtttttt atgcaaaatc
     4021 taatttaata tattgatatt tatatcattt tacgtttctc gttcagcttt cttgtacaaa
     4081 gtggtgatgg gctgcaggaa ttcgatatca agcttaggtg gaatgtaccc atacgatgtt
     4141 cctgactatg cgggctatcc ctatgacgtc ccggactatg caggatccta tccatatgac
     4201 gttccagatt acgctgctca gtgctaactc gagtcatgta attagttatg tcacgcttac
     4261 attcacgccc tccccccaca tccgctctaa ccgaaaagga aggagttaga caacctgaag
     4321 tctaggtccc tatttatttt tttatagtta tgttagtatt aagaacgtta tttatatttc
     4381 aaatttttct tttttttctg tacagacgcg tgtacgcatg taacattata ctgaaaacct
     4441 tgcttgagaa ggttttggga cgctcgaagg ctttaatttg cggccggtac ccagcttttg
     4501 ttccctttag tgagggttaa ttccgagctt ggcgtaatca tggtcatagc tgtttcctgt
     4561 gtgaaattgt tatccgctca caattccaca caacatagga gccggaagca taaagtgtaa
     4621 agcctggggt gcctaatgag tgaggtaact cacattaatt gcgttgcgct cactgcccgc
     4681 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4741 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4801 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4861 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4921 taaaaaggcc gcgttgctgg cgtttttcca taggctcggc ccccctgacg agcatcacaa
     4981 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     5041 cccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     5101 gtccgccttt ctcccttcgg gaagcgtggc gctttctcaa tgctcacgct gtaggtatct
     5161 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     5221 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     5281 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     5341 tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat
     5401 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     5461 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     5521 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     5581 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5641 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5701 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5761 catagttgcc tgactgcccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5821 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5881 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5941 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     6001 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     6061 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgaaaaaa
     6121 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     6181 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     6241 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     6301 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     6361 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6421 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6481 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6541 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6601 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6661 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     6721 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtc
//
