LOCUS       Exported                8197 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pAG423GAL-ccdB
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8197)
  TITLE     S cerevisiae Advanced Gateway Destination Vectors
REFERENCE   2  (bases 1 to 8197)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8197
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     rep_origin      68..1410
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    400..447
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        1437..1541
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1542..2402
                     /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(1760..1779)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2573..3161
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3062..3081
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3315..3332
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3485..3515
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3523..3539
                     /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     3528..3550
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3547..3563
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3547..3563
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3582..3602
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3584..3602
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3626..4067
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    3626..3743
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     4038..4061
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     4082..4098
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4082..4098
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    4106..4230
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        4255..4285
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             4339..4998
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(4405..4424)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             5343..5648
                     /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     5527..5546
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(5689..5813)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(5846..5862)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5847..5863)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      5865..6112
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(5881..5899)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(6130..6149)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6131..6149)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6159..6176)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6159..6175)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6168..6190)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6320..6775
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6407..6426)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6617..6638
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(7035..7694)
                     /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"
     promoter        complement(7695..7881)
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     primer_bind     complement(8028..8047)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8147..8169
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagatgat ccaatatcaa aggaaatgat agcattgaag gatgagacta atccaattga
      121 ggagtggcag catatagaac agctaaaggg tagtgctgaa ggaagcatac gataccccgc
      181 atggaatggg ataatatcac aggaggtact agactacctt tcatcctaca taaatagacg
      241 catataagta cgcatttaag cataaacacg cactatgccg ttcttctcat gtatatatat
      301 atacaggcaa cacgcagata taggtgcgac gtgaacagtg agctgtatgt gcgcagctcg
      361 cgttgcattt tcggaagcgc tcgttttcgg aaacgctttg aagttcctat tccgaagttc
      421 ctattctcta gaaagtatag gaacttcaga gcgcttttga aaaccaaaag cgctctgaag
      481 acgcactttc aaaaaaccaa aaacgcaccg gactgtaacg agctactaaa atattgcgaa
      541 taccgcttcc acaaacattg ctcaaaagta tctctttgct atatatctct gtgctatatc
      601 cctatataac ctacccatcc acctttcgct ccttgaactt gcatctaaac tcgacctcta
      661 cattttttat gtttatctct agtattactc tttagacaaa aaaattgtag taagaactat
      721 tcatagagtg aatcgaaaac aatacgaaaa tgtaaacatt tcctatacgt agtatataga
      781 gacaaaatag aagaaaccgt tcataatttt ctgaccaatg aagaatcatc aacgctatca
      841 ctttctgttc acaaagtatg cgcaatccac atcggtatag aatataatcg gggatgcctt
      901 tatcttgaaa aaatgcaccc gcagcttcgc tagtaatcag taaacgcggg aagtggagtc
      961 aggctttttt tatggaagag aaaatagaca ccaaagtagc cttcttctaa ccttaacgga
     1021 cctacagtgc aaaaagttat caagagactg cattatagag cgcacaaagg agaaaaaaag
     1081 taatctaaga tgctttgtta gaaaaatagc gctctcggga tgcatttttg tagaacaaaa
     1141 aagaagtata gattctttgt tggtaaaata gcgctctcgc gttgcatttc tgttctgtaa
     1201 aaatgcagct cagattcttt gtttgaaaaa ttagcgctct cgcgttgcat ttttgtttta
     1261 caaaaatgaa gcacagattc ttcgttggta aaatagcgct ttcgcgttgc atttctgttc
     1321 tgtaaaaatg cagctcagat tctttgtttg aaaaattagc gctctcgcgt tgcatttttg
     1381 ttctacaaaa tgaagcacag atgcttcgtt caggtggcac ttttcgggga aatgtgcgcg
     1441 gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     1501 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     1561 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     1621 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     1681 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     1741 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag
     1801 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     1861 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     1921 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     1981 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     2041 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa
     2101 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     2161 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     2221 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     2281 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     2341 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     2401 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     2461 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     2521 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     2581 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     2641 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     2701 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     2761 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     2821 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     2881 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     2941 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     3001 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     3061 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     3121 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     3181 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     3241 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     3301 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac
     3361 cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg tttcccgact
     3421 ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta cctcactcat taggcacccc
     3481 aggctttaca ctttatgctt ccggctccta tgttgtgtgg aattgtgagc ggataacaat
     3541 ttcacacagg aaacagctat gaccatgatt acgccaagcg cgcaattaac cctcactaaa
     3601 gggaacaaaa gctggagctc tagtacggat tagaagccgc cgagcgggcg acagccctcc
     3661 gacggaagac tctcctccgt gcgtcctcgt cttcaccggt cgcgttcctg aaacgcagat
     3721 gtgcctcgcg ccgcactgct ccgaacaata aagattctac aatactagct tttatggtta
     3781 tgaagaggaa aaattggcag taacctggcc ccacaaacct tcaaattaac gaatcaaatt
     3841 aacaaccata ggatgataat gcgattagtt ttttagcctt atttctgggg taattaatca
     3901 gcgaagcgat gatttttgat ctattaacag atatataaat ggaaaagctg cataaccact
     3961 ttaactaata ctttcaacat tttcagtttg tattacttct tattcaaatg tcataaaagt
     4021 atcaacaaaa aattgttaat atacctctat actttaacgt caaggagaaa aaaccccgga
     4081 ttctagaact agtggatccc ccatcacaag tttgtacaaa aaagctgaac gagaaacgta
     4141 aaatgatata aatatcaata tattaaatta gattttgcat aaaaaacaga ctacataata
     4201 ctgtaaaaca caacatatcc agtcactatg gcggccgcat taggcacccc aggctttaca
     4261 ctttatgctt ccggctcgta taatgtgtgg attttgagtt aggatccgtc gagattttca
     4321 ggagctaagg aagctaaaat ggagaaaaaa atcactggat ataccaccgt tgatatatcc
     4381 caatggcatc gtaaagaaca ttttgaggca tttcagtcag ttgctcaatg tacctataac
     4441 cagaccgttc agctggatat tacggccttt ttaaagaccg taaagaaaaa taagcacaag
     4501 ttttatccgg cctttattca cattcttgcc cgcctgatga atgctcatcc ggaattccgt
     4561 atggcaatga aagacggtga gctggtgata tgggatagtg ttcacccttg ttacaccgtt
     4621 ttccatgagc aaactgaaac gttttcatcg ctctggagtg aataccacga cgatttccgg
     4681 cagtttctac acatatattc gcaagatgtg gcgtgttacg gtgaaaacct ggcctatttc
     4741 cctaaagggt ttattgagaa tatgtttttc gtctcagcca atccctgggt gagtttcacc
     4801 agttttgatt taaacgtggc caatatggac aacttcttcg cccccgtttt caccatgggc
     4861 aaatattata cgcaaggcga caaggtgctg atgccgctgg cgattcaggt tcatcatgcc
     4921 gtctgtgatg gcttccatgt cggcagaatg cttaatgaat tacaacagta ctgcgatgag
     4981 tggcagggcg gggcgtaaac gccgcgtgga tccggcttac taaaagccag ataacagtat
     5041 gcgtatttgc gcgctgattt ttgcggtata agaatatata ctgatatgta tacccgaagt
     5101 atgtcaaaaa gaggtatgct atgaagcagc gtattacagt gacagttgac agcgacagct
     5161 atcagttgct caaggcatat atgatgtcaa tatctccggt ctggtaagca caaccatgca
     5221 gaatgaagcc cgtcgtctgc gtgccgaacg ctggaaagcg gaaaatcagg aagggatggc
     5281 tgaggtcgcc cggtttattg aaatgaacgg ctcttttgct gacgagaaca ggggctggtg
     5341 aaatgcagtt taaggtttac acctataaaa gagagagccg ttatcgtctg tttgtggatg
     5401 tacagagtga tattattgac acgcccgggc gacggatggt gatccccctg gccagtgcac
     5461 gtctgctgtc agataaagtc tcccgtgaac tttacccggt ggtgcatatc ggggatgaaa
     5521 gctggcgcat gatgaccacc gatatggcca gtgtgccggt ctccgttatc ggggaagaag
     5581 tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc cattaacctg atgttctggg
     5641 gaatataaat gtcaggctcc cttatacaca gccagtctgc aggtcgacca tagtgactgg
     5701 atatgttgtg ttttacagta ttatgtagtc tgttttttat gcaaaatcta atttaatata
     5761 ttgatattta tatcatttta cgtttctcgt tcagctttct tgtacaaagt ggtgatgggc
     5821 tgcaggaatt cgatatcaag cttatcgata ccgtcgacct cgagtcatgt aattagttat
     5881 gtcacgctta cattcacgcc ctccccccac atccgctcta accgaaaagg aaggagttag
     5941 acaacctgaa gtctaggtcc ctatttattt ttttatagtt atgttagtat taagaacgtt
     6001 atttatattt caaatttttc ttttttttct gtacagacgc gtgtacgcat gtaacattat
     6061 actgaaaacc ttgcttgaga aggttttggg acgctcgaag gctttaattt gcggccggta
     6121 cccaattcgc cctatagtga gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt
     6181 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     6241 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     6301 ctgaatggcg aatggcgcga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     6361 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     6421 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     6481 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     6541 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     6601 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     6661 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     6721 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aatttcctga
     6781 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatagat ccgtcgagtt
     6841 caagagaaaa aaaaagaaaa agcaaaaaga aaaaaggaaa gcgcgcctcg ttcagaatga
     6901 cacgtataga atgatgcatt accttgtcat cttcagtatc atactgttcg tatacatact
     6961 tactgacatt cataggtata catatataca catgtatata tatcgtatgc tgcagcttta
     7021 aataatcggt gtcactacat aagaacacct ttggtggagg gaacatcgtt ggtaccattg
     7081 ggcgaggtgg cttctcttat ggcaaccgca agagccttga acgcactctc actacggtga
     7141 tgatcattct tgcctcgcag acaatcaacg tggagggtaa ttctgctagc ctctgcaaag
     7201 ctttcaagaa aatgcgggat catctcgcaa gagagatctc ctactttctc cctttgcaaa
     7261 ccaagttcga caactgcgta cggcctgttc gaaagatcta ccaccgctct ggaaagtgcc
     7321 tcatccaaag gcgcaaatcc tgatccaaac ctttttactc cacgcgccag tagggcctct
     7381 ttaaaagctt gaccgagagc aatcccgcag tcttcagtgg tgtgatggtc gtctatgtgt
     7441 aagtcaccaa tgcactcaac gattagcgac cagccggaat gcttggccag agcatgtatc
     7501 atatggtcca gaaaccctat acctgtgtgg acgttaatca cttgcgattg tgtggcctgt
     7561 tctgctactg cttctgcctc tttttctggg aagatcgagt gctctatcgc taggggacca
     7621 ccctttaaag agatcgcaat ctgaatcttg gtttcatttg taatacgctt tactagggct
     7681 ttctgctctg tcatctttgc cttcgtttat cttgcctgct cattttttag tatattcttc
     7741 gaagaaatca cattacttta tataatgtat aattcattat gtgataatgc caatcgctaa
     7801 gaaaaaaaaa gagtcatccg ctaggggaaa aaaaaaaatg aaaatcatta ccgaggcata
     7861 aaaaaatata gagtgtacta gaggaggcca agagtaatag aaaaagaaaa ttgcgggaaa
     7921 ggactgtgtt atgacttccc tgactaatgc cgtgttcaaa cgatacctgg cagtgactcc
     7981 tagcgctcac caagctctta aaacgggaat ttatggtgca ctctcagtac aatctgctct
     8041 gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     8101 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     8161 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcga
//
