LOCUS       sequence-7907-ex        7594 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 7594)
  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 7594)
  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: pAG416GPD-ccdB-HA
FEATURES             Location/Qualifiers
     source          1..7594
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    70..573
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        610..714
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(715..783,784..1575)
                     /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(933..952)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1746..2334
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2235..2254
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2488..2505
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(2658..2663,2664..2681,2682..2688)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2696..2712
                     /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     2701..2723
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2720..2736
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2720..2736
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2755..2775
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2757..2775
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2804..3448
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     3342..3365
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     primer_bind     3454..3470
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3454..3470
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    3478..3602
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(3627..3632,3633..3650,3651..3657)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             3711..4370
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3777..3796)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             4715..5020
                     /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     4899..4918
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(5061..5185)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             5226..5315
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     terminator      5334..5581
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(5350..5368)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(5599..5618)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5600..5618)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5628..5645)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5628..5644)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5637..5659)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5789..6244
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5876..5895)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6086..6107
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(6375..7178)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(7179..7399)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(7425..7444)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7544..7566
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttaggacgg atcgcttgcc tgtaacttac acgcgcctcg tatcttttaa tgatggaata
      121 atttgggaat ttactctgtg tttatttatt tttatgtttt gtatttggat tttagaaagt
      181 aaataaagaa ggtagaagag ttacggaatg aagaaaaaaa aataaacaaa ggtttaaaaa
      241 atttcaacaa aaagcgtact ttacatatat atttattaga caagaaaagc agattaaata
      301 gatatacatt cgattaacga taagtaaaat gtaaaatcac aggattttcg tgtgtggtct
      361 tctacacaga caagatgaaa caattcggca ttaatacctg agagcaggaa gagcaagata
      421 aaaggtagta tttgttggcg atccccctag agtcttttac atcttcggaa aacaaaaact
      481 attttttctt taatttcttt ttttactttc tatttttaat ttatatattt atattaaaaa
      541 atttaaatta taattatttt tatagcacgt gatgaaaagg acccaggtgg cacttttcgg
      601 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
      661 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
      721 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
      781 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
      841 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
      901 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
      961 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     1021 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     1081 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     1141 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     1201 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     1261 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     1321 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     1381 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     1441 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     1501 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     1561 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     1621 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     1681 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     1741 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     1801 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     1861 ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac
     1921 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     1981 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     2041 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     2101 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     2161 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     2221 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     2281 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     2341 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     2401 cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc
     2461 gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc
     2521 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
     2581 aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag ttacctcact
     2641 cattaggcac cccaggcttt acactttatg cttccggctc ctatgttgtg tggaattgtg
     2701 agcggataac aatttcacac aggaaacagc tatgaccatg attacgccaa gcgcgcaatt
     2761 aaccctcact aaagggaaca aaagctggag ctcttcgagt ttatcattat caatactcgc
     2821 catttcaaag aatacgtaaa taattaatag tagtgatttt cctaacttta tttagtcaaa
     2881 aaattagcct tttaattctg ctgtaacccg tacatgccca aaataggggg cgggttacac
     2941 agaatatata acaccgaagg tgcctgggtg aacaggttta ttcctggcat ccactaaata
     3001 taatggagcc cgctttttaa gctggcatcc agaaaaaaaa agaatcccag caccaaaata
     3061 ttgttttctt caccaaccat cagttcatag gtccattctc ttagcgcaac tacagagaac
     3121 aggggcacaa acaggcaaaa aacgggcaca acctcaatgg agtgatgcaa cctgcctgga
     3181 gtaaatgatg acacaaggca attgacccac gcatgtatct atctcatttt cttacacctt
     3241 ctattacctt ctgctctctc tgatttggaa aaagctgaaa aaaaaggttg aaaccagttc
     3301 cctgaaatta ttcccctact tgactaataa gtatataaag acggtaggta ttgattgtaa
     3361 ttctgtaaat ctatttctta aacttcttaa attctacttt tatagttagt ctttttttta
     3421 gttttaaaac accagaactt agtttcgacg gattctagaa ctagtggatc ccccatcaca
     3481 agtttgtaca aaaaagctga acgagaaacg taaaatgata taaatatcaa tatattaaat
     3541 tagattttgc ataaaaaaca gactacataa tactgtaaaa cacaacatat ccagtcacta
     3601 tggcggccgc attaggcacc ccaggcttta cactttatgc ttccggctcg tataatgtgt
     3661 ggattttgag ttaggatccg tcgagatttt caggagctaa ggaagctaaa atggagaaaa
     3721 aaatcactgg atataccacc gttgatatat cccaatggca tcgtaaagaa cattttgagg
     3781 catttcagtc agttgctcaa tgtacctata accagaccgt tcagctggat attacggcct
     3841 ttttaaagac cgtaaagaaa aataagcaca agttttatcc ggcctttatt cacattcttg
     3901 cccgcctgat gaatgctcat ccggaattcc gtatggcaat gaaagacggt gagctggtga
     3961 tatgggatag tgttcaccct tgttacaccg ttttccatga gcaaactgaa acgttttcat
     4021 cgctctggag tgaataccac gacgatttcc ggcagtttct acacatatat tcgcaagatg
     4081 tggcgtgtta cggtgaaaac ctggcctatt tccctaaagg gtttattgag aatatgtttt
     4141 tcgtctcagc caatccctgg gtgagtttca ccagttttga tttaaacgtg gccaatatgg
     4201 acaacttctt cgcccccgtt ttcaccatgg gcaaatatta tacgcaaggc gacaaggtgc
     4261 tgatgccgct ggcgattcag gttcatcatg ccgtctgtga tggcttccat gtcggcagaa
     4321 tgcttaatga attacaacag tactgcgatg agtggcaggg cggggcgtaa acgccgcgtg
     4381 gatccggctt actaaaagcc agataacagt atgcgtattt gcgcgctgat ttttgcggta
     4441 taagaatata tactgatatg tatacccgaa gtatgtcaaa aagaggtatg ctatgaagca
     4501 gcgtattaca gtgacagttg acagcgacag ctatcagttg ctcaaggcat atatgatgtc
     4561 aatatctccg gtctggtaag cacaaccatg cagaatgaag cccgtcgtct gcgtgccgaa
     4621 cgctggaaag cggaaaatca ggaagggatg gctgaggtcg cccggtttat tgaaatgaac
     4681 ggctcttttg ctgacgagaa caggggctgg tgaaatgcag tttaaggttt acacctataa
     4741 aagagagagc cgttatcgtc tgtttgtgga tgtacagagt gatattattg acacgcccgg
     4801 gcgacggatg gtgatccccc tggccagtgc acgtctgctg tcagataaag tctcccgtga
     4861 actttacccg gtggtgcata tcggggatga aagctggcgc atgatgacca ccgatatggc
     4921 cagtgtgccg gtctccgtta tcggggaaga agtggctgat ctcagccacc gcgaaaatga
     4981 catcaaaaac gccattaacc tgatgttctg gggaatataa atgtcaggct cccttataca
     5041 cagccagtct gcaggtcgac catagtgact ggatatgttg tgttttacag tattatgtag
     5101 tctgtttttt atgcaaaatc taatttaata tattgatatt tatatcattt tacgtttctc
     5161 gttcagcttt cttgtacaaa gtggtgatgg gctgcaggaa ttcgatatca agcttaggtg
     5221 gaatgtaccc atacgatgtt cctgactatg cgggctatcc ctatgacgtc ccggactatg
     5281 caggatccta tccatatgac gttccagatt acgctgctca gtgctaactc gagtcatgta
     5341 attagttatg tcacgcttac attcacgccc tccccccaca tccgctctaa ccgaaaagga
     5401 aggagttaga caacctgaag tctaggtccc tatttatttt tttatagtta tgttagtatt
     5461 aagaacgtta tttatatttc aaatttttct tttttttctg tacagacgcg tgtacgcatg
     5521 taacattata ctgaaaacct tgcttgagaa ggttttggga cgctcgaagg ctttaatttg
     5581 cggccggtac ccaattcgcc ctatagtgag tcgtattacg cgcgctcact ggccgtcgtt
     5641 ttacaacgtc gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat
     5701 ccccctttcg ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag
     5761 ttgcgcagcc tgaatggcga atggcgcgac gcgccctgta gcggcgcatt aagcgcggcg
     5821 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     5881 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     5941 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     6001 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     6061 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     6121 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     6181 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca
     6241 atttcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatagggt
     6301 aataactgat ataattaaat tgaagctcta atttgtgagt ttagtataca tgcatttact
     6361 tataatacag ttttttagtt ttgctggccg catcttctca aatatgcttc ccagcctgct
     6421 tttctgtaac gttcaccctc taccttagca tcccttccct ttgcaaatag tcctcttcca
     6481 acaataataa tgtcagatcc tgtagagacc acatcatcca cggttctata ctgttgaccc
     6541 aatgcgtctc ccttgtcatc taaacccaca ccgggtgtca taatcaacca atcgtaacct
     6601 tcatctcttc cacccatgtc tctttgagca ataaagccga taacaaaatc tttgtcgctc
     6661 ttcgcaatgt caacagtacc cttagtatat tctccagtag atagggagcc cttgcatgac
     6721 aattctgcta acatcaaaag gcctctaggt tcctttgtta cttcttctgc cgcctgcttc
     6781 aaaccgctaa caatacctgg gcccaccaca ccgtgtgcat tcgtaatgtc tgcccattct
     6841 gctattctgt atacacccgc agagtactgc aatttgactg tattaccaat gtcagcaaat
     6901 tttctgtctt cgaagagtaa aaaattgtac ttggcggata atgcctttag cggcttaact
     6961 gtgccctcca tggaaaaatc agtcaagata tccacatgtg tttttagtaa acaaattttg
     7021 ggacctaatg cttcaactaa ctccagtaat tccttggtgg tacgaacatc caatgaagca
     7081 cacaagtttg tttgcttttc gtgcatgata ttaaatagct tggcagcaac aggactagga
     7141 tgagtagcag cacgttcctt atatgtagct ttcgacatga tttatcttcg tttcctgcag
     7201 gtttttgttc tgtgcagttg ggttaagaat actgggcaat ttcatgtttc ttcaacacta
     7261 catatgcgta tatataccaa tctaagtctg tgctccttcc ttcgttcttc cttctgttcg
     7321 gagattaccg aatcaaaaaa atttcaaaga aaccgaaatc aaaaaaaaga ataaaaaaaa
     7381 aatgatgaat tgaattgaaa agctgtggta tggtgcactc tcagtacaat ctgctctgat
     7441 gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     7501 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     7561 cagaggtttt caccgtcatc accgaaacgc gcga
//
