LOCUS       62056_11225        7588 bp DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7588)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7588
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        771..1476
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             1522..1542
                     /codon_start=1
                     /label=SV40 NLS
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /translation="PKKKRKV"
     CDS             1558..1899
                     /codon_start=1
                     /label=GAL4 activation domain
                     /note="activation domain of the GAL4 transcriptional
                     activator"
                     /translation="ANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN
                     VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD
                     EDTPPNPKKE"
     promoter        1905..1923
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1942..1968
                     /codon_start=1
                     /label=HA
                     /note="HA (human influenza hemagglutinin) epitope tag"
                     /translation="YPYDVPDYA"
     terminator      2487..2674
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae
                     alcohol dehydrogenase 1 (ADH1) gene"
     CDS             complement(2794..3885)
                     /codon_start=1
                     /label=LEU2
                     /note="3-isopropylmalate dehydrogenase, required for
                     leucine biosynthesis"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(3886..4291)
                     /label=LEU2 promoter
     primer_bind     complement(4333..4349)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(4357..4373)
                     /label=lac operator
                     /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(4381..4411)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(4426..4447)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     protein_bind    4502..4535
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(4886..5474)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5648..6505)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6506..6610)
                     /label=AmpR promoter
     misc_feature    7079..7582
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an
                     autonomously replicating sequence"
ORIGIN
        1 tgcatgcctg caggtcgaga tccgggatcg aagaaatgat ggtaaatgaa ataggaaatc
       61 aaggagcatg aaggcaaaag acaaatataa gggtcgaacg aaaaataaag tgaaaagtgt
      121 tgatatgatg tatttggctt tgcggcgccg aaaaaacgag tttacgcaat tgcacaatca
      181 tgctgactct gtggcggacc cgcgctcttg ccggcccggc gataacgctg ggcgtgaggc
      241 tgtgcccggc ggagtttttt gcgcctgcat tttccaaggt ttaccctgcg ctaaggggcg
      301 agattggaga agcaataaga atgccggttg gggttgcgat gatgacgacc acgacaactg
      361 gtgtcattat ttaagttgcc gaaagaacct gagtgcattt gcaacatgag tatactagaa
      421 gaatgagcca agacttgcga gacgcgagtt tgccggtggt gcgaacaata gagcgaccat
      481 gaccttgaag gtgagacgcg cataaccgct agagtacttt gaagaggaaa cagcaatagg
      541 gttgctacca gtataaatag acaggtacat acaacactgg aaatggttgt ctgtttgagt
      601 acgctttcaa ttcatttggg tgtgcacttt attatgttac aatatggaag ggaactttac
      661 acttctccta tgcacatata ttaattaaag tccaatgcta gtagagaagg ggggtaacac
      721 ccctccgcgc tcttttccga tttttttcta aaccgtggaa tatttcggat atccttttgt
      781 tgtttccggg tgtacaatat ggacttcctc ttttctggca accaaaccca tacatcggga
      841 ttcctataat accttcgttg gtctccctaa catgtaggtg gcggagggga gatatacaat
      901 agaacagata ccagacaaga cataatgggc taaacaagac tacaccaatt acactgcctc
      961 attgatggtg gtacataacg aactaatact gtagccctag acttgatagc catcatcata
     1021 tcgaagtttc actacccttt ttccatttgc catctattga agtaataata ggcgcatgca
     1081 acttcttttc tttttttttc ttttctctct cccccgttgt tgtctcacca tatccgcaat
     1141 gacaaaaaaa atgatggaag acactaaagg aaaaaattaa cgacaaagac agcaccaaca
     1201 gatgtcgttg ttccagagct gatgaggggt atctcgaagc acacgaaact ttttccttcc
     1261 ttcattcacg cacactactc tctaatgagc aacggtatac ggccttcctt ccagttactt
     1321 gaatttgaaa taaaaaaaag tttgctgtct tgctatcaag tataaataga cctgcaatta
     1381 ttaatctttt gtttcctcgt cattgttctc gttccctttc ttccttgttt ctttttctgc
     1441 acaatatttc aagctatacc aagcatacaa tcaactccaa gctttgcaaa gatggataaa
     1501 gcggaattaa ttcccgagcc tccaaaaaag aagagaaagg tcgaattggg taccgccgcc
     1561 aattttaatc aaagtgggaa tattgctgat agctcattgt ccttcacttt cactaacagt
     1621 agcaacggtc cgaacctcat aacaactcaa acaaattctc aagcgctttc acaaccaatt
     1681 gcctcctcta acgttcatga taacttcatg aataatgaaa tcacggctag taaaattgat
     1741 gatggtaata attcaaaacc actgtcacct ggttggacgg accaaactgc gtataacgcg
     1801 tttggaatca ctacagggat gtttaatacc actacaatgg atgatgtata taactatcta
     1861 ttcgatgatg aagatacccc accaaaccca aaaaaagaga tctttaatac gactcactat
     1921 agggcgagcg ccgccatgga gtacccatac gacgtaccag attacgctca tatggccatg
     1981 gaggccagtg aattccaccc aagcagtggt atcaacgcag agtggccatt atggcccggg
     2041 aaaaaacatg tcggccgcct cggcctctag agggtgggca tcgatacggg atccatcgag
     2101 ctcgagctgc agatgaatcg tagatactga aaaaccccgc aagttcactt caactgtgca
     2161 tcgtgcacca tctcaatttc tttcatttat acatcgtttt gccttctttt atgtaactat
     2221 actcctctaa gtttcaatct tggccatgta acctctgatc tatagaattt tttaaatgac
     2281 tagaattaat gcccatcttt tttttggacc taaattcttc atgaaaatat attacgaggg
     2341 cttattcaga agctttggac ttcttcgcca gaggtttggt caagtctcca atcaaggttg
     2401 tcggcttgtc taccttgcca gaaatttacg aaaagatgga aaagggtcaa atcgttggta
     2461 gatacgttgt tgacacttct aaataagcga atttcttatg atttatgatt tttattatta
     2521 aataagttat aaaaaaaata agtgtataca aattttaaag tgactcttag gttttaaaac
     2581 gaaaattctt attcttgagt aactctttcc tgtaggtcag gttgctttct caggtatagc
     2641 atgaggtcgc tcttattgac cacacctcta ccggccggtc gaaattcccc taccctatga
     2701 acatattcca ttttgtaatt tcgtgtcgtt tctattatga atttcattta taaagtttat
     2761 gtacaaatat cataaaaaaa gagaatcttt ttaagcaagg attttcttaa cttcttcggc
     2821 gacagcatca ccgacttcgg tggtactgtt ggaaccacct aaatcaccag ttctgatacc
     2881 tgcatccaaa acctttttaa ctgcatcttc aatggcctta ccttcttcag gcaagttcaa
     2941 tgacaatttc aacatcattg cagcagacaa gatagtggcg atagggttga ccttattctt
     3001 tggcaaatct ggagcagaac cgtggcatgg ttcgtacaaa ccaaatgcgg tgttcttgtc
     3061 tggcaaagag gccaaggacg cagatggcaa caaacccaag gaacctggga taacggaggc
     3121 ttcatcggag atgatatcac caaacatgtt gctggtgatt ataataccat ttaggtgggt
     3181 tgggttctta actaggatca tggcggcaga atcaatcaat tgatgttgaa ccttcaatgt
     3241 aggaaattcg ttcttgatgg tttcctccac agtttttctc cataatcttg aagaggccaa
     3301 aacattagct ttatccaagg accaaatagg caatggtggc tcatgttgta gggccatgaa
     3361 agcggccatt cttgtgattc tttgcacttc tggaacggtg tattgttcac tatcccaagc
     3421 gacaccatca ccatcgtctt cctttctctt accaaagtaa atacctccca ctaattctct
     3481 gacaacaacg aagtcagtac ctttagcaaa ttgtggcttg attggagata agtctaaaag
     3541 agagtcggat gcaaagttac atggtcttaa gttggcgtac aattgaagtt ctttacggat
     3601 ttttagtaaa ccttgttcag gtctaacact acctgtaccc catttaggac cacccacagc
     3661 acctaacaaa acggcatcaa ccttcttgga ggcttccagc gcctcatctg gaagtgggac
     3721 acctgtagcg tcgatagcag caccaccaat taaatgattt tcgaaatcga acttgacatt
     3781 ggaacgaaca tcagaaatag ctttaagaac cttaatggct tcggctgtga tttcttgacc
     3841 aacgtggtca cctggcaaaa cgacgatctt cttaggggca gacattagaa tggtatatcc
     3901 ttgaaatata tatatatatt gctgaaatgt aaaaggtaag aaaagttaga aagtaagacg
     3961 attgctaacc acctattgga aaaaacaata ggtccttaaa taatattgtc aacttcaagt
     4021 attgtgatgc aagcatttag tcatgaacgc ttctctattc tatatgaaaa gccggttccg
     4081 gcgctctcac ctttcctttt tctcccaatt tttcagttga aaaaggtata tgcgtcaggc
     4141 gacctctgaa attaacaaaa aatttccagt catcgaattt gattctgtgc gatagcgccc
     4201 ctgtgtgttc tcgttatgtt gaggaaaaaa ataatggttg ctaagagatt cgaactcttg
     4261 catcttacga tacctgagta ttcccacagt tggggatctc gactctagct agaggatcaa
     4321 ttcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     4381 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     4441 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     4501 gataacttcg tataatgtat gctatacgaa gttattaggt ctgaagagga gtttacgtcc
     4561 agccaagcta gcttggctgc aggtcgagcg gccgcgatcc ggaaccctta atataacttc
     4621 gtataatgta tgctatacga agttatcagc tgcattaatg aatcggccaa cgcgcgggga
     4681 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     4741 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     4801 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4861 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4921 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4981 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     5041 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     5101 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     5161 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     5221 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     5281 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     5341 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     5401 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     5461 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     5521 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     5581 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     5641 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     5701 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     5761 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     5821 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     5881 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     5941 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     6001 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     6061 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     6121 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     6181 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     6241 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     6301 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     6361 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     6421 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     6481 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     6541 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     6601 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca
     6661 tgacattaac ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg
     6721 atgacggtga aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag
     6781 cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg
     6841 gctggcttaa ctatgcggca tcagagcaga ttgtactgag agtgcaccat aacgcattta
     6901 agcataaaca cgcactatgc cgttcttctc atgtatatat atatacaggc aacacgcaga
     6961 tataggtgcg acgtgaacag tgagctgtat gtgcgcagct cgcgttgcat tttcggaagc
     7021 gctcgttttc ggaaacgctt tgaagttcct attccgaagt tcctattcgg tccttttcat
     7081 cacgtgctat aaaaataatt ataatttaaa ttttttaata taaatatata aattaaaaat
     7141 agaaagtaaa aaaagaaatt aaagaaaaaa tagtttttgt tttccgaaga tgtaaaagac
     7201 tctaggggga tcgccaacaa atactacctt ttatcttgct cttcctgctc tcaggtatta
     7261 atgccgaatt gtttcatctt gtctgtgtag aagaccacac acgaaaatcc tgtgatttta
     7321 cattttactt atcgttaatc gaatgtatat ctatttaatc tgcttttctt gtctaataaa
     7381 tatatatgta aagtacgctt tttgttgaaa ttttttaaac ctttgtttat ttttttttct
     7441 tcattccgta actcttctac cttctttatt tactttctaa aatccaaata caaaacataa
     7501 aaataaataa acacagagta aattcccaaa ttattccatc attaaaagat acgaggcgcg
     7561 tgtaagttac aggcaagcga tccgtcct
//
