LOCUS       sequence-7966-ex        7368 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 7368)
  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 7368)
  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-EGFP-ccdB
FEATURES             Location/Qualifiers
     source          1..7368
                     /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"
     CDS             join(2383..2385,2386..2388,2389..3099)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2383..2385,2386..2388,2389..3099)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2428..2449)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2689..2708)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3036..3057
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    3124..3248
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(3273..3278,3279..3296,3297..3303)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             3357..4016
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3423..3442)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             4361..4666
                     /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     4545..4564
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4707..4831)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     terminator      4838..5085
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4854..4872)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(5108..5126)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5147..5163)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5147..5163)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5160..5182)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5171..5187
                     /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(5195..5201,5202..5219,5220..5225))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(5378..5395)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5549..6137)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6308..7099,7100..7168))
                     /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     6931..6950
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7169..7273)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7341..7359
                     /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 ccccgggaca aaatggtgag caagggcgag
     2401 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
     2461 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
     2521 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc
     2581 tacggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag
     2641 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac
     2701 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
     2761 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
     2821 aacagccaca acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc
     2881 aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac
     2941 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc
     3001 gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     3061 gccgccggga tcactctcgg catggacgag ctgtacaagg gtggaccagg tggtggacat
     3121 caaacaagtt tgtacaaaaa agctgaacga gaaacgtaaa atgatataaa tatcaatata
     3181 ttaaattaga ttttgcataa aaaacagact acataatact gtaaaacaca acatatccag
     3241 tcactatggc ggccgcatta ggcaccccag gctttacact ttatgcttcc ggctcgtata
     3301 atgtgtggat tttgagttag gatccgtcga gattttcagg agctaaggaa gctaaaatgg
     3361 agaaaaaaat cactggatat accaccgttg atatatccca atggcatcgt aaagaacatt
     3421 ttgaggcatt tcagtcagtt gctcaatgta cctataacca gaccgttcag ctggatatta
     3481 cggccttttt aaagaccgta aagaaaaata agcacaagtt ttatccggcc tttattcaca
     3541 ttcttgcccg cctgatgaat gctcatccgg aattccgtat ggcaatgaaa gacggtgagc
     3601 tggtgatatg ggatagtgtt cacccttgtt acaccgtttt ccatgagcaa actgaaacgt
     3661 tttcatcgct ctggagtgaa taccacgacg atttccggca gtttctacac atatattcgc
     3721 aagatgtggc gtgttacggt gaaaacctgg cctatttccc taaagggttt attgagaata
     3781 tgtttttcgt ctcagccaat ccctgggtga gtttcaccag ttttgattta aacgtggcca
     3841 atatggacaa cttcttcgcc cccgttttca ccatgggcaa atattatacg caaggcgaca
     3901 aggtgctgat gccgctggcg attcaggttc atcatgccgt ctgtgatggc ttccatgtcg
     3961 gcagaatgct taatgaatta caacagtact gcgatgagtg gcagggcggg gcgtaaacgc
     4021 cgcgtggatc cggcttacta aaagccagat aacagtatgc gtatttgcgc gctgattttt
     4081 gcggtataag aatatatact gatatgtata cccgaagtat gtcaaaaaga ggtatgctat
     4141 gaagcagcgt attacagtga cagttgacag cgacagctat cagttgctca aggcatatat
     4201 gatgtcaata tctccggtct ggtaagcaca accatgcaga atgaagcccg tcgtctgcgt
     4261 gccgaacgct ggaaagcgga aaatcaggaa gggatggctg aggtcgcccg gtttattgaa
     4321 atgaacggct cttttgctga cgagaacagg ggctggtgaa atgcagttta aggtttacac
     4381 ctataaaaga gagagccgtt atcgtctgtt tgtggatgta cagagtgata ttattgacac
     4441 gcccgggcga cggatggtga tccccctggc cagtgcacgt ctgctgtcag ataaagtctc
     4501 ccgtgaactt tacccggtgg tgcatatcgg ggatgaaagc tggcgcatga tgaccaccga
     4561 tatggccagt gtgccggtct ccgttatcgg ggaagaagtg gctgatctca gccaccgcga
     4621 aaatgacatc aaaaacgcca ttaacctgat gttctgggga atataaatgt caggctccct
     4681 tatacacagc cagtctgcag gtcgaccata gtgactggat atgttgtgtt ttacagtatt
     4741 atgtagtctg ttttttatgc aaaatctaat ttaatatatt gatatttata tcattttacg
     4801 tttctcgttc agctttcttg tacaaagtgg ttcgatgtca tgtaattagt tatgtcacgc
     4861 ttacattcac gccctccccc cacatccgct ctaaccgaaa aggaaggagt tagacaacct
     4921 gaagtctagg tccctattta tttttttata gttatgttag tattaagaac gttatttata
     4981 tttcaaattt ttcttttttt tctgtacaga cgcgtgtacg catgtaacat tatactgaaa
     5041 accttgcttg agaaggtttt gggacgctcg aaggctttaa tttgcggccg gtacccagct
     5101 tttgttccct ttagtgaggg ttaattccga gcttggcgta atcatggtca tagctgtttc
     5161 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat aggagccgga agcataaagt
     5221 gtaaagcctg gggtgcctaa tgagtgaggt aactcacatt aattgcgttg cgctcactgc
     5281 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     5341 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     5401 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     5461 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     5521 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct cggcccccct gacgagcatc
     5581 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     5641 cgttcccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     5701 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcaatgctca cgctgtaggt
     5761 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     5821 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     5881 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     5941 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg
     6001 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     6061 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     6121 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     6181 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     6241 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     6301 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     6361 catccatagt tgcctgactg cccgtcgtgt agataactac gatacgggag ggcttaccat
     6421 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     6481 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     6541 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     6601 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     6661 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgaa
     6721 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     6781 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     6841 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     6901 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     6961 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     7021 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     7081 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     7141 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     7201 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     7261 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa accattatta
     7321 tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtc
//
