LOCUS       sequence-8060-ex        7544 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 7544)
  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 7544)
  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: pAG303GAL-Cerulean-ccdB
FEATURES             Location/Qualifiers
     source          1..7544
                     /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"
     CDS             join(2569..2571,2572..2574,2575..3285)
                     /codon_start=1
                     /product="monomeric optimized variant of ECFP (Kremers et 
                     al., 2006)"
                     /label=SCFP3A
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISDNVYITADKQKNGIK
                     ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2614..2635)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2875..2894)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3222..3243
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    3310..3434
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(3459..3464,3465..3482,3483..3489)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             3543..4202
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3609..3628)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             4547..4852
                     /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     4731..4750
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4893..5017)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     terminator      5024..5271
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(5040..5058)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     promoter        complement(5294..5312)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5333..5349)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5333..5349)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5346..5368)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5357..5373
                     /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(5381..5387,5388..5405,5406..5411))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     primer_bind     complement(5564..5581)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5735..6323)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6494..7285,7286..7354))
                     /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     7117..7136
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7355..7459)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     7517..7535
                     /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 gggacaaaat ggtgagcaag
     2581 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
     2641 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
     2701 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
     2761 ctgacctggg gcgtgcagtg cttcgcccgc taccccgacc acatgaagca gcacgacttc
     2821 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
     2881 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
     2941 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
     3001 aactacatca gcgacaacgt ctatatcacc gccgacaagc agaagaacgg catcaaggcc
     3061 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
     3121 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
     3181 cagtccaagc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
     3241 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagggtgg accaggtggt
     3301 ggacatcaaa caagtttgta caaaaaagct gaacgagaaa cgtaaaatga tataaatatc
     3361 aatatattaa attagatttt gcataaaaaa cagactacat aatactgtaa aacacaacat
     3421 atccagtcac tatggcggcc gcattaggca ccccaggctt tacactttat gcttccggct
     3481 cgtataatgt gtggattttg agttaggatc cgtcgagatt ttcaggagct aaggaagcta
     3541 aaatggagaa aaaaatcact ggatatacca ccgttgatat atcccaatgg catcgtaaag
     3601 aacattttga ggcatttcag tcagttgctc aatgtaccta taaccagacc gttcagctgg
     3661 atattacggc ctttttaaag accgtaaaga aaaataagca caagttttat ccggccttta
     3721 ttcacattct tgcccgcctg atgaatgctc atccggaatt ccgtatggca atgaaagacg
     3781 gtgagctggt gatatgggat agtgttcacc cttgttacac cgttttccat gagcaaactg
     3841 aaacgttttc atcgctctgg agtgaatacc acgacgattt ccggcagttt ctacacatat
     3901 attcgcaaga tgtggcgtgt tacggtgaaa acctggccta tttccctaaa gggtttattg
     3961 agaatatgtt tttcgtctca gccaatccct gggtgagttt caccagtttt gatttaaacg
     4021 tggccaatat ggacaacttc ttcgcccccg ttttcaccat gggcaaatat tatacgcaag
     4081 gcgacaaggt gctgatgccg ctggcgattc aggttcatca tgccgtctgt gatggcttcc
     4141 atgtcggcag aatgcttaat gaattacaac agtactgcga tgagtggcag ggcggggcgt
     4201 aaacgccgcg tggatccggc ttactaaaag ccagataaca gtatgcgtat ttgcgcgctg
     4261 atttttgcgg tataagaata tatactgata tgtatacccg aagtatgtca aaaagaggta
     4321 tgctatgaag cagcgtatta cagtgacagt tgacagcgac agctatcagt tgctcaaggc
     4381 atatatgatg tcaatatctc cggtctggta agcacaacca tgcagaatga agcccgtcgt
     4441 ctgcgtgccg aacgctggaa agcggaaaat caggaaggga tggctgaggt cgcccggttt
     4501 attgaaatga acggctcttt tgctgacgag aacaggggct ggtgaaatgc agtttaaggt
     4561 ttacacctat aaaagagaga gccgttatcg tctgtttgtg gatgtacaga gtgatattat
     4621 tgacacgccc gggcgacgga tggtgatccc cctggccagt gcacgtctgc tgtcagataa
     4681 agtctcccgt gaactttacc cggtggtgca tatcggggat gaaagctggc gcatgatgac
     4741 caccgatatg gccagtgtgc cggtctccgt tatcggggaa gaagtggctg atctcagcca
     4801 ccgcgaaaat gacatcaaaa acgccattaa cctgatgttc tggggaatat aaatgtcagg
     4861 ctcccttata cacagccagt ctgcaggtcg accatagtga ctggatatgt tgtgttttac
     4921 agtattatgt agtctgtttt ttatgcaaaa tctaatttaa tatattgata tttatatcat
     4981 tttacgtttc tcgttcagct ttcttgtaca aagtggttcg atgtcatgta attagttatg
     5041 tcacgcttac attcacgccc tccccccaca tccgctctaa ccgaaaagga aggagttaga
     5101 caacctgaag tctaggtccc tatttatttt tttatagtta tgttagtatt aagaacgtta
     5161 tttatatttc aaatttttct tttttttctg tacagacgcg tgtacgcatg taacattata
     5221 ctgaaaacct tgcttgagaa ggttttggga cgctcgaagg ctttaatttg cggccggtac
     5281 ccagcttttg ttccctttag tgagggttaa ttccgagctt ggcgtaatca tggtcatagc
     5341 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatagga gccggaagca
     5401 taaagtgtaa agcctggggt gcctaatgag tgaggtaact cacattaatt gcgttgcgct
     5461 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     5521 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     5581 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     5641 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     5701 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctcggc ccccctgacg
     5761 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     5821 accaggcgtt cccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     5881 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcaa tgctcacgct
     5941 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     6001 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     6061 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     6121 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag
     6181 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     6241 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     6301 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     6361 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     6421 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     6481 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     6541 ttcgttcatc catagttgcc tgactgcccg tcgtgtagat aactacgata cgggagggct
     6601 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     6661 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     6721 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     6781 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     6841 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     6901 tgtgaaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     6961 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     7021 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     7081 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     7141 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     7201 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     7261 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     7321 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     7381 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     7441 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc ttattatcat
     7501 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtc
//
