LOCUS       Exported                8223 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    pAG426GAL-ccdB-HA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8223)
  TITLE     S cerevisiae Advanced Gateway Destination Vectors
REFERENCE   2  (bases 1 to 8223)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8223
                     /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      69..1411
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    401..448
                     /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        1438..1542
                     /gene="bla"
                     /label=AmpR promoter
     CDS             1543..2403
                     /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(1761..1780)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2574..3162
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3063..3082
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3316..3333
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3486..3516
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3524..3540
                     /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     3529..3551
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3548..3564
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3548..3564
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3583..3603
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3585..3603
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3627..4068
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    3627..3744
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     4039..4062
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     4083..4099
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4083..4099
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    4107..4231
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        4256..4286
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             4340..4999
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(4406..4425)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             5344..5649
                     /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     5528..5547
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(5690..5814)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             5855..5944
                     /codon_start=1
                     /product="three tandem HA epitope tags"
                     /label=3xHA
                     /translation="YPYDVPDYAGYPYDVPDYAGSYPYDVPDYA"
     terminator      5963..6210
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(5979..5997)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(6228..6247)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6229..6247)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6257..6274)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6257..6273)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6266..6288)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      6418..6873
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6505..6524)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6715..6736
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(7004..7807)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(7808..8023)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(8054..8073)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8173..8195
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagtatga tccaatatca aaggaaatga tagcattgaa ggatgagact aatccaattg
      121 aggagtggca gcatatagaa cagctaaagg gtagtgctga aggaagcata cgataccccg
      181 catggaatgg gataatatca caggaggtac tagactacct ttcatcctac ataaatagac
      241 gcatataagt acgcatttaa gcataaacac gcactatgcc gttcttctca tgtatatata
      301 tatacaggca acacgcagat ataggtgcga cgtgaacagt gagctgtatg tgcgcagctc
      361 gcgttgcatt ttcggaagcg ctcgttttcg gaaacgcttt gaagttccta ttccgaagtt
      421 cctattctct agaaagtata ggaacttcag agcgcttttg aaaaccaaaa gcgctctgaa
      481 gacgcacttt caaaaaacca aaaacgcacc ggactgtaac gagctactaa aatattgcga
      541 ataccgcttc cacaaacatt gctcaaaagt atctctttgc tatatatctc tgtgctatat
      601 ccctatataa cctacccatc cacctttcgc tccttgaact tgcatctaaa ctcgacctct
      661 acatttttta tgtttatctc tagtattact ctttagacaa aaaaattgta gtaagaacta
      721 ttcatagagt gaatcgaaaa caatacgaaa atgtaaacat ttcctatacg tagtatatag
      781 agacaaaata gaagaaaccg ttcataattt tctgaccaat gaagaatcat caacgctatc
      841 actttctgtt cacaaagtat gcgcaatcca catcggtata gaatataatc ggggatgcct
      901 ttatcttgaa aaaatgcacc cgcagcttcg ctagtaatca gtaaacgcgg gaagtggagt
      961 caggcttttt ttatggaaga gaaaatagac accaaagtag ccttcttcta accttaacgg
     1021 acctacagtg caaaaagtta tcaagagact gcattataga gcgcacaaag gagaaaaaaa
     1081 gtaatctaag atgctttgtt agaaaaatag cgctctcggg atgcattttt gtagaacaaa
     1141 aaagaagtat agattctttg ttggtaaaat agcgctctcg cgttgcattt ctgttctgta
     1201 aaaatgcagc tcagattctt tgtttgaaaa attagcgctc tcgcgttgca tttttgtttt
     1261 acaaaaatga agcacagatt cttcgttggt aaaatagcgc tttcgcgttg catttctgtt
     1321 ctgtaaaaat gcagctcaga ttctttgttt gaaaaattag cgctctcgcg ttgcattttt
     1381 gttctacaaa atgaagcaca gatgcttcgt tcaggtggca cttttcgggg aaatgtgcgc
     1441 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     1501 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     1561 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     1621 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     1681 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     1741 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     1801 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     1861 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     1921 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     1981 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     2041 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     2101 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     2161 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     2221 tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca
     2281 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     2341 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     2401 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     2461 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     2521 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     2581 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     2641 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     2701 gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac
     2761 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     2821 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     2881 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     2941 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     3001 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     3061 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     3121 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     3181 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     3241 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     3301 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa
     3361 ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac
     3421 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt acctcactca ttaggcaccc
     3481 caggctttac actttatgct tccggctcct atgttgtgtg gaattgtgag cggataacaa
     3541 tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa ccctcactaa
     3601 agggaacaaa agctggagct ctagtacgga ttagaagccg ccgagcgggc gacagccctc
     3661 cgacggaaga ctctcctccg tgcgtcctcg tcttcaccgg tcgcgttcct gaaacgcaga
     3721 tgtgcctcgc gccgcactgc tccgaacaat aaagattcta caatactagc ttttatggtt
     3781 atgaagagga aaaattggca gtaacctggc cccacaaacc ttcaaattaa cgaatcaaat
     3841 taacaaccat aggatgataa tgcgattagt tttttagcct tatttctggg gtaattaatc
     3901 agcgaagcga tgatttttga tctattaaca gatatataaa tggaaaagct gcataaccac
     3961 tttaactaat actttcaaca ttttcagttt gtattacttc ttattcaaat gtcataaaag
     4021 tatcaacaaa aaattgttaa tatacctcta tactttaacg tcaaggagaa aaaaccccgg
     4081 attctagaac tagtggatcc cccatcacaa gtttgtacaa aaaagctgaa cgagaaacgt
     4141 aaaatgatat aaatatcaat atattaaatt agattttgca taaaaaacag actacataat
     4201 actgtaaaac acaacatatc cagtcactat ggcggccgca ttaggcaccc caggctttac
     4261 actttatgct tccggctcgt ataatgtgtg gattttgagt taggatccgt cgagattttc
     4321 aggagctaag gaagctaaaa tggagaaaaa aatcactgga tataccaccg ttgatatatc
     4381 ccaatggcat cgtaaagaac attttgaggc atttcagtca gttgctcaat gtacctataa
     4441 ccagaccgtt cagctggata ttacggcctt tttaaagacc gtaaagaaaa ataagcacaa
     4501 gttttatccg gcctttattc acattcttgc ccgcctgatg aatgctcatc cggaattccg
     4561 tatggcaatg aaagacggtg agctggtgat atgggatagt gttcaccctt gttacaccgt
     4621 tttccatgag caaactgaaa cgttttcatc gctctggagt gaataccacg acgatttccg
     4681 gcagtttcta cacatatatt cgcaagatgt ggcgtgttac ggtgaaaacc tggcctattt
     4741 ccctaaaggg tttattgaga atatgttttt cgtctcagcc aatccctggg tgagtttcac
     4801 cagttttgat ttaaacgtgg ccaatatgga caacttcttc gcccccgttt tcaccatggg
     4861 caaatattat acgcaaggcg acaaggtgct gatgccgctg gcgattcagg ttcatcatgc
     4921 cgtctgtgat ggcttccatg tcggcagaat gcttaatgaa ttacaacagt actgcgatga
     4981 gtggcagggc ggggcgtaaa cgccgcgtgg atccggctta ctaaaagcca gataacagta
     5041 tgcgtatttg cgcgctgatt tttgcggtat aagaatatat actgatatgt atacccgaag
     5101 tatgtcaaaa agaggtatgc tatgaagcag cgtattacag tgacagttga cagcgacagc
     5161 tatcagttgc tcaaggcata tatgatgtca atatctccgg tctggtaagc acaaccatgc
     5221 agaatgaagc ccgtcgtctg cgtgccgaac gctggaaagc ggaaaatcag gaagggatgg
     5281 ctgaggtcgc ccggtttatt gaaatgaacg gctcttttgc tgacgagaac aggggctggt
     5341 gaaatgcagt ttaaggttta cacctataaa agagagagcc gttatcgtct gtttgtggat
     5401 gtacagagtg atattattga cacgcccggg cgacggatgg tgatccccct ggccagtgca
     5461 cgtctgctgt cagataaagt ctcccgtgaa ctttacccgg tggtgcatat cggggatgaa
     5521 agctggcgca tgatgaccac cgatatggcc agtgtgccgg tctccgttat cggggaagaa
     5581 gtggctgatc tcagccaccg cgaaaatgac atcaaaaacg ccattaacct gatgttctgg
     5641 ggaatataaa tgtcaggctc ccttatacac agccagtctg caggtcgacc atagtgactg
     5701 gatatgttgt gttttacagt attatgtagt ctgtttttta tgcaaaatct aatttaatat
     5761 attgatattt atatcatttt acgtttctcg ttcagctttc ttgtacaaag tggtgatggg
     5821 ctgcaggaat tcgatatcaa gcttaggtgg aatgtaccca tacgatgttc ctgactatgc
     5881 gggctatccc tatgacgtcc cggactatgc aggatcctat ccatatgacg ttccagatta
     5941 cgctgctcag tgctaactcg agtcatgtaa ttagttatgt cacgcttaca ttcacgccct
     6001 ccccccacat ccgctctaac cgaaaaggaa ggagttagac aacctgaagt ctaggtccct
     6061 atttattttt ttatagttat gttagtatta agaacgttat ttatatttca aatttttctt
     6121 ttttttctgt acagacgcgt gtacgcatgt aacattatac tgaaaacctt gcttgagaag
     6181 gttttgggac gctcgaaggc tttaatttgc ggccggtacc caattcgccc tatagtgagt
     6241 cgtattacgc gcgctcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg
     6301 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag
     6361 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggcgcgacg
     6421 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     6481 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     6541 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     6601 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     6661 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     6721 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     6781 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     6841 cgaattttaa caaaatatta acgtttacaa tttcctgatg cggtattttc tccttacgca
     6901 tctgtgcggt atttcacacc gcatagggta ataactgata taattaaatt gaagctctaa
     6961 tttgtgagtt tagtatacat gcatttactt ataatacagt tttttagttt tgctggccgc
     7021 atcttctcaa atatgcttcc cagcctgctt ttctgtaacg ttcaccctct accttagcat
     7081 cccttccctt tgcaaatagt cctcttccaa caataataat gtcagatcct gtagagacca
     7141 catcatccac ggttctatac tgttgaccca atgcgtctcc cttgtcatct aaacccacac
     7201 cgggtgtcat aatcaaccaa tcgtaacctt catctcttcc acccatgtct ctttgagcaa
     7261 taaagccgat aacaaaatct ttgtcgctct tcgcaatgtc aacagtaccc ttagtatatt
     7321 ctccagtaga tagggagccc ttgcatgaca attctgctaa catcaaaagg cctctaggtt
     7381 cctttgttac ttcttctgcc gcctgcttca aaccgctaac aatacctggg cccaccacac
     7441 cgtgtgcatt cgtaatgtct gcccattctg ctattctgta tacacccgca gagtactgca
     7501 atttgactgt attaccaatg tcagcaaatt ttctgtcttc gaagagtaaa aaattgtact
     7561 tggcggataa tgcctttagc ggcttaactg tgccctccat ggaaaaatca gtcaagatat
     7621 ccacatgtgt ttttagtaaa caaattttgg gacctaatgc ttcaactaac tccagtaatt
     7681 ccttggtggt acgaacatcc aatgaagcac acaagtttgt ttgcttttcg tgcatgatat
     7741 taaatagctt ggcagcaaca ggactaggat gagtagcagc acgttcctta tatgtagctt
     7801 tcgacatgat ttatcttcgt ttcctgcagg tttttgttct gtgcagttgg gttaagaata
     7861 ctgggcaatt tcatgtttct tcaacactac atatgcgtat atataccaat ctaagtctgt
     7921 gctccttcct tcgttcttcc ttctgttcgg agattaccga atcaaaaaaa tttcaaagaa
     7981 accgaaatca aaaaaaagaa taaaaaaaaa atgatgaatt gaattgaaaa gctgtggtat
     8041 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc
     8101 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     8161 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     8221 cga
//
