LOCUS       sequence-382816-        7506 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 7506)
  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 7506)
  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: pAG303GPD-ccdB-EGFP
FEATURES             Location/Qualifiers
     source          1..7506
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(133..795)
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     promoter        complement(796..983)
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     primer_bind     complement(1130..1149)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1249..1271
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1309..1327)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1395..1499
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1500..1568,1569..2360)
                     /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(1718..1737)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2531..3119
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3020..3039
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3273..3290
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3407..3428
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3443..3448,3449..3466,3467..3473)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3481..3497
                     /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     3486..3508
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3505..3521
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3505..3521
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        3542..3560
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             complement(join(3593..4306,4307..4309,4310..4312))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(3596..4306,4307..4309,4310..4312))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3638..3659)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     3987..4006
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4246..4267
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     protein_bind    4344..4468
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             complement(4509..4814)
                     /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     complement(4611..4630)
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(5156..5815)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     5730..5749
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(join(5869..5875,5876..5893,5894..5899))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    complement(5924..6048)
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(6056..6072)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6056..6072)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        complement(6078..6721)
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     complement(6161..6184)
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     primer_bind     complement(6741..6760)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6742..6760)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6767..6784)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6767..6783)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6776..6798)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6924..7379
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7011..7030)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7221..7242
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 cgtatagaat gatgcattac cttgtcatct tcagtatcat actgttcgta tacatactta
       61 ctgacattca taggtataca tatatacaca tgtatatata tcgtatgctg cagctttaaa
      121 taatcggtgt cactacataa gaacaccttt ggtggaggga acatcgttgg taccattggg
      181 cgaggtggct tctcttatgg caaccgcaag agccttgaac gcactctcac tacggtgatg
      241 atcattcttg cctcgcagac aatcaacgtg gagggtaatt ctgctagcct ctgcaaagct
      301 ttcaagaaaa tgcgggatca tctcgcaaga gagatctcct actttctccc tttgcaaacc
      361 aagttcgaca actgcgtacg gcctgttcga aagatctacc accgctctgg aaagtgcctc
      421 atccaaaggc gcaaatcctg atccaaacct ttttactcca cgcacggccc ctagggcctc
      481 tttaaaagct tgaccgagag caatcccgca gtcttcagtg gtgtgatggt cgtctatgtg
      541 taagtcacca atgcactcaa cgattagcga ccagccggaa tgcttggcca gagcatgtat
      601 catatggtcc agaaacccta tacctgtgtg gacgttaatc acttgcgatt gtgtggcctg
      661 ttctgctact gcttctgcct ctttttctgg gaagatcgag tgctctatcg ctaggggacc
      721 accctttaaa gagatcgcaa tctgaatctt ggtttcattt gtaatacgct ttactagggc
      781 tttctgctct gtcatctttg ccttcgttta tcttgcctgc tcatttttta gtatattctt
      841 cgaagaaatc acattacttt atataatgta taattcatta tgtgataatg ccaatcgcta
      901 agaaaaaaaa agagtcatcc gctaggtgga aaaaaaaaaa tgaaaatcat taccgaggca
      961 taaaaaaata tagagtgtac tagaggaggc caagagtaat agaaaaagaa aattgcggga
     1021 aaggactgtg ttatgacttc cctgactaat gccgtgttca aacgatacct ggcagtgact
     1081 cctagcgctc accaagctct taaaacggga atttatggtg cactctcagt acaatctgct
     1141 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     1201 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     1261 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     1321 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     1381 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     1441 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     1501 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     1561 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     1621 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     1681 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     1741 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     1801 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     1861 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     1921 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     1981 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     2041 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     2101 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     2161 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     2221 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     2281 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     2341 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     2401 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     2461 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     2521 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     2581 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     2641 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     2701 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     2761 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     2821 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     2881 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     2941 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     3001 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     3061 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     3121 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     3181 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     3241 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     3301 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
     3361 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
     3421 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
     3481 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctcg
     3541 aaattaaccc tcactaaagg gaacaaaagc tggtaccggg ccccccctcg agttacttgt
     3601 acagctcgtc catgccgaga gtgatcccgg cggcggtcac gaactccagc aggaccatgt
     3661 gatcgcgctt ctcgttgggg tctttgctca gggcggactg ggtgctcagg tagtggttgt
     3721 cgggcagcag cacggggccg tcgccgatgg gggtgttctg ctggtagtgg tcggcgagct
     3781 gcacgctgcc gtcctcgatg ttgtggcgga tcttgaagtt caccttgatg ccgttcttct
     3841 gcttgtcggc catgatatag acgttgtggc tgttgtagtt gtactccagc ttgtgcccca
     3901 ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc gatgcggttc accagggtgt
     3961 cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc gtcgtccttg aagaagatgg
     4021 tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa gaagtcgtgc tgcttcatgt
     4081 ggtcggggta gcggctgaag cactgcacgc cgtaggtcag ggtggtcacg agggtgggcc
     4141 agggcacggg cagcttgccg gtggtgcaga tgaacttcag ggtcagcttg ccgtaggtgg
     4201 catcgccctc gccctcgccg gacacgctga acttgtggcc gtttacgtcg ccgtccagct
     4261 cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc cttgctcacc attaagcttg
     4321 atatcgaatt cctgcagccc atcaccactt tgtacaagaa agctgaacga gaaacgtaaa
     4381 atgatataaa tatcaatata ttaaattaga ttttgcataa aaaacagact acataatact
     4441 gtaaaacaca acatatccag tcactatggt cgacctgcag actggctgtg tataagggag
     4501 cctgacattt atattcccca gaacatcagg ttaatggcgt ttttgatgtc attttcgcgg
     4561 tggctgagat cagccacttc ttccccgata acggagaccg gcacactggc catatcggtg
     4621 gtcatcatgc gccagctttc atccccgata tgcaccaccg ggtaaagttc acgggagact
     4681 ttatctgaca gcagacgtgc actggccagg gggatcacca tccgtcgccc gggcgtgtca
     4741 ataatatcac tctgtacatc cacaaacaga cgataacggc tctctctttt ataggtgtaa
     4801 accttaaact gcatttcacc agcccctgtt ctcgtcagca aaagagccgt tcatttcaat
     4861 aaaccgggcg acctcagcca tcccttcctg attttccgct ttccagcgtt cggcacgcag
     4921 acgacgggct tcattctgca tggttgtgct taccagaccg gagatattga catcatatat
     4981 gccttgagca actgatagct gtcgctgtca actgtcactg taatacgctg cttcatagca
     5041 tacctctttt tgacatactt cgggtataca tatcagtata tattcttata ccgcaaaaat
     5101 cagcgcgcaa atacgcatac tgttatctgg cttttagtaa gccggatcca cgcgtttacg
     5161 ccccgccctg ccactcatcg cagtactgtt gtaattcatt aagcattctg ccgacatgga
     5221 agccatcaca aacggcatga tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt
     5281 gcgtataata tttgcccatg gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt
     5341 ttaaatcaaa actggtgaaa ctcacccagg gattggctga gacgaaaaac atattctcaa
     5401 taaacccttt agggaaatag gccaggtttt caccgtaaca cgccacatct tgcgaatata
     5461 tgtgtagaaa ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa aacgtttcag
     5521 tttgctcatg gaaaacggtg taacaagggt gaacactatc ccatatcacc agctcaccgt
     5581 ctttcattgc catacggaat tccggatgag cattcatcag gcgggcaaga atgtgaataa
     5641 aggccggata aaacttgtgc ttatttttct ttacggtctt taaaaaggcc gtaatatcca
     5701 gctgaacggt ctggttatag gtacattgag caactgactg aaatgcctca aaatgttctt
     5761 tacgatgcca ttgggatata tcaacggtgg tatatccagt gatttttttc tccattttag
     5821 cttccttagc tcctgaaaat ctcgacggat cctaactcaa aatccacaca ttatacgagc
     5881 cggaagcata aagtgtaaag cctggggtgc ctaatgcggc cgccatagtg actggatatg
     5941 ttgtgtttta cagtattatg tagtctgttt tttatgcaaa atctaattta atatattgat
     6001 atttatatca ttttacgttt ctcgttcagc ttttttgtac aaacttgtga tgggggatcc
     6061 actagttcta gaatccgtcg aaactaagtt ctggtgtttt aaaactaaaa aaaagactaa
     6121 ctataaaagt agaatttaag aagtttaaga aatagattta cagaattaca atcaatacct
     6181 accgtcttta tatacttatt agtcaagtag gggaataatt tcagggaact ggtttcaacc
     6241 ttttttttca gctttttcca aatcagagag agcagaaggt aatagaaggt gtaagaaaat
     6301 gagatagata catgcgtggg tcaattgcct tgtgtcatca tttactccag gcaggttgca
     6361 tcactccatt gaggttgtgc ccgttttttg cctgtttgtg cccctgttct ctgtagttgc
     6421 gctaagagaa tggacctatg aactgatggt tggtgaagaa aacaatattt tggtgctggg
     6481 attctttttt tttctggatg ccagcttaaa aagcgggctc cattatattt agtggatgcc
     6541 aggaataaac tgttcaccca gacacctacg atgttatata ttctgtgtaa cccgccccct
     6601 attttgggca tgtacgggtt acagcagaat taaaaggcta attttttgac taaataaagt
     6661 taggaaaatc actactatta attatttacg tattctttga aatggcgagt attgataatg
     6721 ataaactgag ctccaattcg ccctatagtg agtcgtatta caattcactg gccgtcgttt
     6781 tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc
     6841 cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt
     6901 tgcgcagcct gaatggcgaa tggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     6961 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
     7021 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     7081 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
     7141 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     7201 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     7261 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     7321 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaatttc
     7381 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat agatccgtcg
     7441 agttcaagag aaaaaaaaag aaaaagcaaa aagaaaaaag gaaagcgcgc ctcgttcaga
     7501 atgaca
//
