LOCUS       sequence-7821-ex        7757 bp    DNA     circular SYN 15-APR-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7757)
  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 7757)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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-ECFP
FEATURES             Location/Qualifiers
     source          1..7757
                     /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        315..502
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             503..1162
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALLARGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHFL
                     ESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      complement(1423..1878)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1560..1581)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1772..1791
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2008..2030
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2022..2039
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2023..2039
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2046..2065
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2046..2064
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2089..2733
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     2627..2650
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     primer_bind     2739..2755
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2739..2755
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    2763..2887
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2912..2917,2918..2935,2936..2942)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2996..3655
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3062..3081)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             4000..4305
                     /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     4184..4203
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4346..4470)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             join(4502..4504,4505..4507,4508..5218)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4547..4568)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4808..4827)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5155..5176
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      5237..5484
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(5253..5271)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(5507..5525)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5546..5562)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5546..5562)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5559..5581)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5570..5586
                     /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(5594..5600,5601..5618,5619..5624))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(5777..5794)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5948..6536)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6707..7498,7499..7567))
                     /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     7330..7349
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7568..7672)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7730..7748
                     /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 accataattc cgttttaaga gcttggtgag cgctaggagt cactgccagg tatcgtttga
      241 acacggcatt agtcagggaa gtcataacac agtcctttcc cgcaattttc tttttctatt
      301 actcttggcc tcctctagta cactctatat ttttttatgc ctcggtaatg attttcattt
      361 ttttttttcc acctagcgga tgactctttt tttttcttag cgattggcat tatcacataa
      421 tgaattatac attatataaa gtaatgtgat ttcttcgaag aatatactaa aaaatgagca
      481 ggcaagataa acgaaggcaa agatgacaga gcagaaagcc ctagtaaagc gtattacaaa
      541 tgaaaccaag attcagattg cgatctcttt aaagggtggt cccctagcga tagagcactc
      601 gatcttccca gaaaaagagg cagaagcagt agcagaacag gccacacaat cgcaagtgat
      661 taacgtccac acaggtatag ggtttctgga ccatatgata catgctctgg ccaagcattc
      721 cggctggtcg ctaatcgttg agtgcattgg tgacttacac atagacgacc atcacaccac
      781 tgaagactgc gggattgctc tcggtcaagc ttttaaagag gccctactgg cgcgtggagt
      841 aaaaaggttt ggatcaggat ttgcgccttt ggatgaggca ctttccagag cggtggtaga
      901 tctttcgaac aggccgtacg cagttgtcga acttggtttg caaagggaga aagtaggaga
      961 tctctcttgc gagatgatcc cgcattttct tgaaagcttt gcagaggcta gcagaattac
     1021 cctccacgtt gattgtctgc gaggcaagaa tgatcatcac cgtagtgaga gtgcgttcaa
     1081 ggctcttgcg gttgccataa gagaagccac ctcgcccaat ggtaccaacg atgttccctc
     1141 caccaaaggt gttcttatgt agtgacaccg attatttaaa gctgcagcat acgatatata
     1201 tacatgtgta tatatgtata cctatgaatg tcagtaagta tgtatacgaa cagtatgata
     1261 ctgaagatga caaggtaatg catcattcta tacgtgtcat tctgaacgag gcgcgctttc
     1321 cttttttctt tttgcttttt cttttttttt ctcttgaact cgacggatca tatgcggtgt
     1381 gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggaaattgta aacgttaata
     1441 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     1501 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     1561 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     1621 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     1681 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     1741 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     1801 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     1861 cgccgctaca gggcgcgtcg cgccattcgc cattcaggct gcgcaactgt tgggaagggc
     1921 gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa ggggggatgt gctgcaaggc
     1981 gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg
     2041 aattgtaata cgactcacta tagggcgaat tggagctctt cgagtttatc attatcaata
     2101 ctcgccattt caaagaatac gtaaataatt aatagtagtg attttcctaa ctttatttag
     2161 tcaaaaaatt agccttttaa ttctgctgta acccgtacat gcccaaaata gggggcgggt
     2221 tacacagaat atataacacc gaaggtgcct gggtgaacag gtttattcct ggcatccact
     2281 aaatataatg gagcccgctt tttaagctgg catccagaaa aaaaaagaat cccagcacca
     2341 aaatattgtt ttcttcacca accatcagtt cataggtcca ttctcttagc gcaactacag
     2401 agaacagggg cacaaacagg caaaaaacgg gcacaacctc aatggagtga tgcaacctgc
     2461 ctggagtaaa tgatgacaca aggcaattga cccacgcatg tatctatctc attttcttac
     2521 accttctatt accttctgct ctctctgatt tggaaaaagc tgaaaaaaaa ggttgaaacc
     2581 agttccctga aattattccc ctacttgact aataagtata taaagacggt aggtattgat
     2641 tgtaattctg taaatctatt tcttaaactt cttaaattct acttttatag ttagtctttt
     2701 ttttagtttt aaaacaccag aacttagttt cgacggattc tagaactagt ggatccccca
     2761 tcacaagttt gtacaaaaaa gctgaacgag aaacgtaaaa tgatataaat atcaatatat
     2821 taaattagat tttgcataaa aaacagacta cataatactg taaaacacaa catatccagt
     2881 cactatggcg gccgcattag gcaccccagg ctttacactt tatgcttccg gctcgtataa
     2941 tgtgtggatt ttgagttagg atccgtcgag attttcagga gctaaggaag ctaaaatgga
     3001 gaaaaaaatc actggatata ccaccgttga tatatcccaa tggcatcgta aagaacattt
     3061 tgaggcattt cagtcagttg ctcaatgtac ctataaccag accgttcagc tggatattac
     3121 ggccttttta aagaccgtaa agaaaaataa gcacaagttt tatccggcct ttattcacat
     3181 tcttgcccgc ctgatgaatg ctcatccgga attccgtatg gcaatgaaag acggtgagct
     3241 ggtgatatgg gatagtgttc acccttgtta caccgttttc catgagcaaa ctgaaacgtt
     3301 ttcatcgctc tggagtgaat accacgacga tttccggcag tttctacaca tatattcgca
     3361 agatgtggcg tgttacggtg aaaacctggc ctatttccct aaagggttta ttgagaatat
     3421 gtttttcgtc tcagccaatc cctgggtgag tttcaccagt tttgatttaa acgtggccaa
     3481 tatggacaac ttcttcgccc ccgttttcac catgggcaaa tattatacgc aaggcgacaa
     3541 ggtgctgatg ccgctggcga ttcaggttca tcatgccgtc tgtgatggct tccatgtcgg
     3601 cagaatgctt aatgaattac aacagtactg cgatgagtgg cagggcgggg cgtaaacgcc
     3661 gcgtggatcc ggcttactaa aagccagata acagtatgcg tatttgcgcg ctgatttttg
     3721 cggtataaga atatatactg atatgtatac ccgaagtatg tcaaaaagag gtatgctatg
     3781 aagcagcgta ttacagtgac agttgacagc gacagctatc agttgctcaa ggcatatatg
     3841 atgtcaatat ctccggtctg gtaagcacaa ccatgcagaa tgaagcccgt cgtctgcgtg
     3901 ccgaacgctg gaaagcggaa aatcaggaag ggatggctga ggtcgcccgg tttattgaaa
     3961 tgaacggctc ttttgctgac gagaacaggg gctggtgaaa tgcagtttaa ggtttacacc
     4021 tataaaagag agagccgtta tcgtctgttt gtggatgtac agagtgatat tattgacacg
     4081 cccgggcgac ggatggtgat ccccctggcc agtgcacgtc tgctgtcaga taaagtctcc
     4141 cgtgaacttt acccggtggt gcatatcggg gatgaaagct ggcgcatgat gaccaccgat
     4201 atggccagtg tgccggtctc cgttatcggg gaagaagtgg ctgatctcag ccaccgcgaa
     4261 aatgacatca aaaacgccat taacctgatg ttctggggaa tataaatgtc aggctccctt
     4321 atacacagcc agtctgcagg tcgaccatag tgactggata tgttgtgttt tacagtatta
     4381 tgtagtctgt tttttatgca aaatctaatt taatatattg atatttatat cattttacgt
     4441 ttctcgttca gctttcttgt acaaagtggt gatgggctgc aggaattcga tatcaagctt
     4501 aatggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
     4561 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta
     4621 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
     4681 cctcgtgacc accctgacct ggggcgtgca gtgcttcagc cgctaccccg accacatgaa
     4741 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt
     4801 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
     4861 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
     4921 caagctggag tacaactaca tcagccacaa cgtctatatc accgccgaca agcagaagaa
     4981 cggcatcaag gccaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc
     5041 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
     5101 ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt
     5161 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagct
     5221 gtacaagtaa ctcgagtcat gtaattagtt atgtcacgct tacattcacg ccctcccccc
     5281 acatccgctc taaccgaaaa ggaaggagtt agacaacctg aagtctaggt ccctatttat
     5341 ttttttatag ttatgttagt attaagaacg ttatttatat ttcaaatttt tctttttttt
     5401 ctgtacagac gcgtgtacgc atgtaacatt atactgaaaa ccttgcttga gaaggttttg
     5461 ggacgctcga aggctttaat ttgcggccgg tacccagctt ttgttccctt tagtgagggt
     5521 taattccgag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     5581 tcacaattcc acacaacata ggagccggaa gcataaagtg taaagcctgg ggtgcctaat
     5641 gagtgaggta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     5701 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     5761 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     5821 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     5881 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     5941 tggcgttttt ccataggctc ggcccccctg acgagcatca caaaaatcga cgctcaagtc
     6001 agaggtggcg aaacccgaca ggactataaa gataccaggc gttcccccct ggaagctccc
     6061 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     6121 cgggaagcgt ggcgctttct caatgctcac gctgtaggta tctcagttcg gtgtaggtcg
     6181 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     6241 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     6301 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     6361 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     6421 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     6481 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     6541 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     6601 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     6661 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     6721 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactgc
     6781 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     6841 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     6901 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     6961 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     7021 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     7081 aacgatcaag gcgagttaca tgatccccca tgttgtgaaa aaaagcggtt agctccttcg
     7141 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     7201 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     7261 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     7321 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     7381 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     7441 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     7501 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     7561 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     7621 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     7681 cccgaaaagt gccacctgac gtcttattat catgacatta acctataaaa ataggcgtat
     7741 cacgaggccc tttcgtc
//
