LOCUS       sequence-7820-ex        7558 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 7558)
  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 7558)
  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: pAG303GAL-ccdB-ECFP
FEATURES             Location/Qualifiers
     source          1..7558
                     /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        2084..2525
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    2084..2201
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     2496..2519
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     2540..2556
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2540..2556
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    2564..2688
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2713..2718,2719..2736,2737..2743)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2797..3456
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2863..2882)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             3801..4106
                     /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     3985..4004
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4147..4271)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             join(4303..4305,4306..4308,4309..5019)
                     /codon_start=1
                     /product="enhanced CFP"
                     /label=ECFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4348..4369)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4609..4628)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4956..4977
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      5038..5285
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(5054..5072)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(5308..5326)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5347..5363)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5347..5363)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5360..5382)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5371..5387
                     /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(5395..5401,5402..5419,5420..5425))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(5578..5595)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5749..6337)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6508..7299,7300..7368))
                     /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     7131..7150
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7369..7473)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7531..7549
                     /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 tggagctcta gtacggatta gaagccgccg
     2101 agcgggcgac agccctccga cggaagactc tcctccgtgc gtcctcgtct tcaccggtcg
     2161 cgttcctgaa acgcagatgt gcctcgcgcc gcactgctcc gaacaataaa gattctacaa
     2221 tactagcttt tatggttatg aagaggaaaa attggcagta acctggcccc acaaaccttc
     2281 aaattaacga atcaaattaa caaccatagg atgataatgc gattagtttt ttagccttat
     2341 ttctggggta attaatcagc gaagcgatga tttttgatct attaacagat atataaatgg
     2401 aaaagctgca taaccacttt aactaatact ttcaacattt tcagtttgta ttacttctta
     2461 ttcaaatgtc ataaaagtat caacaaaaaa ttgttaatat acctctatac tttaacgtca
     2521 aggagaaaaa accccggatt ctagaactag tggatccccc atcacaagtt tgtacaaaaa
     2581 agctgaacga gaaacgtaaa atgatataaa tatcaatata ttaaattaga ttttgcataa
     2641 aaaacagact acataatact gtaaaacaca acatatccag tcactatggc ggccgcatta
     2701 ggcaccccag gctttacact ttatgcttcc ggctcgtata atgtgtggat tttgagttag
     2761 gatccgtcga gattttcagg agctaaggaa gctaaaatgg agaaaaaaat cactggatat
     2821 accaccgttg atatatccca atggcatcgt aaagaacatt ttgaggcatt tcagtcagtt
     2881 gctcaatgta cctataacca gaccgttcag ctggatatta cggccttttt aaagaccgta
     2941 aagaaaaata agcacaagtt ttatccggcc tttattcaca ttcttgcccg cctgatgaat
     3001 gctcatccgg aattccgtat ggcaatgaaa gacggtgagc tggtgatatg ggatagtgtt
     3061 cacccttgtt acaccgtttt ccatgagcaa actgaaacgt tttcatcgct ctggagtgaa
     3121 taccacgacg atttccggca gtttctacac atatattcgc aagatgtggc gtgttacggt
     3181 gaaaacctgg cctatttccc taaagggttt attgagaata tgtttttcgt ctcagccaat
     3241 ccctgggtga gtttcaccag ttttgattta aacgtggcca atatggacaa cttcttcgcc
     3301 cccgttttca ccatgggcaa atattatacg caaggcgaca aggtgctgat gccgctggcg
     3361 attcaggttc atcatgccgt ctgtgatggc ttccatgtcg gcagaatgct taatgaatta
     3421 caacagtact gcgatgagtg gcagggcggg gcgtaaacgc cgcgtggatc cggcttacta
     3481 aaagccagat aacagtatgc gtatttgcgc gctgattttt gcggtataag aatatatact
     3541 gatatgtata cccgaagtat gtcaaaaaga ggtatgctat gaagcagcgt attacagtga
     3601 cagttgacag cgacagctat cagttgctca aggcatatat gatgtcaata tctccggtct
     3661 ggtaagcaca accatgcaga atgaagcccg tcgtctgcgt gccgaacgct ggaaagcgga
     3721 aaatcaggaa gggatggctg aggtcgcccg gtttattgaa atgaacggct cttttgctga
     3781 cgagaacagg ggctggtgaa atgcagttta aggtttacac ctataaaaga gagagccgtt
     3841 atcgtctgtt tgtggatgta cagagtgata ttattgacac gcccgggcga cggatggtga
     3901 tccccctggc cagtgcacgt ctgctgtcag ataaagtctc ccgtgaactt tacccggtgg
     3961 tgcatatcgg ggatgaaagc tggcgcatga tgaccaccga tatggccagt gtgccggtct
     4021 ccgttatcgg ggaagaagtg gctgatctca gccaccgcga aaatgacatc aaaaacgcca
     4081 ttaacctgat gttctgggga atataaatgt caggctccct tatacacagc cagtctgcag
     4141 gtcgaccata gtgactggat atgttgtgtt ttacagtatt atgtagtctg ttttttatgc
     4201 aaaatctaat ttaatatatt gatatttata tcattttacg tttctcgttc agctttcttg
     4261 tacaaagtgg tgatgggctg caggaattcg atatcaagct taatggtgag caagggcgag
     4321 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
     4381 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
     4441 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc
     4501 tggggcgtgc agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag
     4561 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac
     4621 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
     4681 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
     4741 atcagccaca acgtctatat caccgccgac aagcagaaga acggcatcaa ggccaacttc
     4801 aagatccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac
     4861 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc
     4921 gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     4981 gccgccggga tcactctcgg catggacgag ctgtacaagc tgtacaagta actcgagtca
     5041 tgtaattagt tatgtcacgc ttacattcac gccctccccc cacatccgct ctaaccgaaa
     5101 aggaaggagt tagacaacct gaagtctagg tccctattta tttttttata gttatgttag
     5161 tattaagaac gttatttata tttcaaattt ttcttttttt tctgtacaga cgcgtgtacg
     5221 catgtaacat tatactgaaa accttgcttg agaaggtttt gggacgctcg aaggctttaa
     5281 tttgcggccg gtacccagct tttgttccct ttagtgaggg ttaattccga gcttggcgta
     5341 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     5401 aggagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgaggt aactcacatt
     5461 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     5521 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     5581 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
     5641 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
     5701 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     5761 cggcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     5821 aggactataa agataccagg cgttcccccc tggaagctcc ctcgtgcgct ctcctgttcc
     5881 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     5941 tcaatgctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     6001 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     6061 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     6121 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     6181 cactagaagg acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     6241 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     6301 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     6361 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     6421 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     6481 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     6541 agcgatctgt ctatttcgtt catccatagt tgcctgactg cccgtcgtgt agataactac
     6601 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
     6661 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
     6721 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
     6781 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
     6841 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
     6901 atgatccccc atgttgtgaa aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
     6961 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
     7021 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
     7081 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
     7141 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
     7201 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
     7261 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
     7321 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
     7381 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
     7441 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
     7501 cgtcttatta tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtc
//
