LOCUS       Exported                4910 bp ds-DNA     circular SYN 22-JUL-2020
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 4910)
  AUTHORS   Arthur
  TITLE     Direct Submission
  JOURNAL   Exported Wednesday, Jul 22, 2020 from SnapGene 5.1.4.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4910
                     /organism="recombinant plasmid"
                     /mol_type="other DNA"
     promoter        1..939
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 promoter
                     /note="inducible promoter, regulated by methanol"
     regulatory      941..950
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             947..951
                     /codon_start=1
                     /label=Start
                     /translation="M"
     CDS             953..970
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             977..997
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1004..1873
                     /codon_start=1
                     /label=K2P4.1
                     /translation="MTTAPQEPPARPLQAGSGAGPAPGRAMRSTTLLALLALVLLYLVS
                     GALVFRALEQPHEQQAQRELGEVREKFLRAHPCVSDQELGLLIKEVADALGGGADPETQ
                     STSQSSHSAWDLGSAFFFSGTIITTIGYGNVALRTDAGRLFCIFYALVGIPLFGILLAG
                     VGDRLGSSLRHGIGHIEAIFLKWHVPPELVRVLSAMLFLLIGCLLFVLTPTFVFCYMED
                     WSKLEAIYFVIVTLTTVGFGDYVAGADPRQDSPAYQPLVWFWILLGLAYFASVLTTIGN
                     WLRVVSRRT"
     CDS             1082..1084
                     /label=Isoform 1 Start
     CDS             1883..1903
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1910..2614
                     /codon_start=1
                     /label=mCherry_pichia_codon_optimized
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     CDS             2630..2653
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     terminator      2733..2979
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        2994..3405
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        3413..3460
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3479..3853
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     terminator      3919..4166
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     rep_origin      complement(4241..4829)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gatctaacat ccaaagacga aaggttgaat gaaacctttt tgccatccga catccacagg
       61 tccattctca cacataagtg ccaaacgcaa caggagggga tacactagca gcagaccgtt
      121 gcaaacgcag gacctccact cctcttctcc tcaacaccca cttttgccat cgaaaaacca
      181 gcccagttat tgggcttgat tggagctcgc tcattccaat tccttctatt aggctactaa
      241 caccatgact ttattagcct gtctatcctg gcccccctgg cgaggttcat gtttgtttat
      301 ttccgaatgc aacaagctcc gcattacacc cgaacatcac tccagatgag ggctttctga
      361 gtgtggggtc aaatagtttc atgttcccca aatggcccaa aactgacagt ttaaacgctg
      421 tcttggaacc taatatgaca aaagcgtgat ctcatccaag atgaactaag tttggttcgt
      481 tgaaatgcta acggccagtt ggtcaaaaag aaacttccaa aagtcggcat accgtttgtc
      541 ttgtttggta ttgattgacg aatgctcaaa aataatctca ttaatgctta gcgcagtctc
      601 tctatcgctt ctgaaccccg gtgcacctgt gccgaaacgc aaatggggaa acacccgctt
      661 tttggatgat tatgcattgt ctccacattg tatgcttcca agattctggt gggaatactg
      721 ctgatagcct aacgttcatg atcaaaattt aactgttcta acccctactt gacagcaata
      781 tataaacaga aggaagctgc cctgtcttaa accttttttt ttatcatcat tattagctta
      841 ctttcataat tgcgactggt tccaattgac aagcttttga ttttaacgac ttttaacgac
      901 aacttgagaa gatcaaaaaa caactaatta ttcgaagcta gccaccatgg ctcaccatca
      961 tcatcatcac ggttccgaga atctgtactt ccaaagcggt accatgacta ccgctcctca
     1021 agagccacca gcaagacccc ttcaagcagg ctcaggcgca ggtcccgctc ctggccgtgc
     1081 catgaggtca accactttgt tggctctgct tgctttagtg ctgctttacc tagtcagtgg
     1141 tgccctggtg tttagggcct tggaacaacc ccacgagcaa caagctcagc gtgagctggg
     1201 tgaggtcaga gaaaaatttt taagagcaca cccatgtgta tccgaccaag aacttggact
     1261 gctaatcaaa gaggtggccg acgcccttgg cggcggcgcc gaccctgaaa cccaatccac
     1321 ctctcagtca agtcacagtg cctgggatct gggaagtgcc ttctttttca gtggtacaat
     1381 tatcacgaca ataggctacg gaaacgtggc cttgaggacg gatgccggta gactgttttg
     1441 catattctac gcattagtag gaatccccct gtttggtatc ttgctagctg gtgtgggtga
     1501 tcgtctggga tcatcactta gacatggtat aggccatata gaggccatct tcctgaaatg
     1561 gcatgtacca ccagaattgg tcagggtttt atcagctatg ttatttttac ttatcggatg
     1621 cctattgttt gtactgaccc caacattcgt tttctgctac atggaagact ggtccaaact
     1681 tgaagctatt tattttgtga tagtgacctt aactacggtc ggctttggcg attacgtcgc
     1741 aggcgcagat ccacgtcaag attctccagc atatcaacca ctagtgtggt tctggatcct
     1801 actgggatta gcctacttcg cttctgtctt gactactata ggcaactggt tgagagtcgt
     1861 ttcccgtagg accggtacca gcgaaaacct gtattttcag ggtactagtg tctctaaggg
     1921 agaagaggac aatatggcta ttatcaagga gttcatgaga tttaaagtcc atatggaggg
     1981 ctctgttaat ggtcatgaat ttgaaattga aggtgaagga gaaggaagac cttatgaggg
     2041 aacccaaacg gctaagctga aagtaactaa aggcggccca cttcctttcg catgggatat
     2101 actgtctcca caattcatgt atggctctaa agcttatgta aagcaccccg ccgacattcc
     2161 cgattatcta aagttatcct ttcccgaggg atttaaatgg gaaagggtta tgaacttcga
     2221 ggatggcggc gtggtaactg ttacgcagga ctcctccctt caagatggtg aatttattta
     2281 caaagtgaag cttcgtggta ctaatttccc aagtgacggt cctgtgatgc agaagaagac
     2341 tatgggttgg gaagctagtt ccgagaggat gtaccccgaa gatggtgccc tgaagggtga
     2401 gattaagcaa cgtctaaaat taaaagacgg cggtcactat gacgctgaag taaagaccac
     2461 atataaggca aaaaagcccg tccagcttcc tggcgcctac aatgttaata tcaagttgga
     2521 catcacctcc cataacgaag attacactat cgtcgagcaa tacgagaggg cagagggaag
     2581 acattccaca ggtggtatgg acgagctata taaactcgag ggcgcttcct ggtcacatcc
     2641 acagtttgag aaatgagttt gtagccttag acatgactgt tcctcagttc aagttgggca
     2701 cttacgagaa gaccggtctt gctagattct aatcaagagg atgtcagaat gccatttgcc
     2761 tgagagatgc aggcttcatt tttgatactt ttttatttgt aacctatata gtataggatt
     2821 ttttttgtca ttttgtttct tctcgtacga gcttgctcct gatcagccta tctcgcagct
     2881 gatgaatatc ttgtggtagg ggtttgggaa aatcattcga gtttgatgtt tttcttggta
     2941 tttcccactc ctcttcagag tacagaagat taagtgagac cttcgtttgt gcggatcccc
     3001 cacacaccat agcttcaaaa tgtttctact ccttttttac tcttccagat tttctcggac
     3061 tccgcgcatc gccgtaccac ttcaaaacac ccaagcacag catactaaat tttccctctt
     3121 tcttcctcta gggtgtcgtt aattacccgt actaaaggtt tggaaaagaa aaaagagacc
     3181 gcctcgtttc tttttcttcg tcgaaaaagg caataaaaat ttttatcacg tttctttttc
     3241 ttgaaatttt tttttttagt ttttttctct ttcagtgacc tccattgata tttaagttaa
     3301 taaacggtct tcaatttctc aagtttcagt ttcatttttc ttgttctatt acaacttttt
     3361 ttacttcttg ttcattagaa agaaagcata gcaatctaat ctaaggggcg gtgttgacaa
     3421 ttaatcatcg gcatagtata tcggcatagt ataatacgac aaggtgagga actaaaccat
     3481 ggccaagttg accagtgccg ttccggtgct caccgcgcgc gacgtcgccg gagcggtcga
     3541 gttctggacc gaccggctcg ggttctcccg ggacttcgtg gaggacgact tcgccggtgt
     3601 ggtccgggac gacgtgaccc tgttcatcag cgcggtccag gaccaggtgg tgccggacaa
     3661 caccctggcc tgggtgtggg tgcgcggcct ggacgagctg tacgccgagt ggtcggaggt
     3721 cgtgtccacg aacttccggg acgcctccgg gccggccatg accgagatcg gcgagcagcc
     3781 gtgggggcgg gagttcgccc tgcgcgaccc ggccggcaac tgcgtgcact tcgtggccga
     3841 ggagcaggac tgacacgtcc gacggcggcc cacgggtccc aggcctcgga gatccgtccc
     3901 ccttttcctt tgtcgatatc atgtaattag ttatgtcacg cttacattca cgccctcccc
     3961 ccacatccgc tctaaccgaa aaggaaggag ttagacaacc tgaagtctag gtccctattt
     4021 atttttttat agttatgtta gtattaagaa cgttatttat atttcaaatt tttctttttt
     4081 ttctgtacag acgcgtgtac gcatgtaaca ttatactgaa aaccttgctt gagaaggttt
     4141 tgggacgctc gaaggcttta atttgcaagc tggagaccaa catgtgagca aaaggccagc
     4201 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     4261 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     4321 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     4381 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcaatgct
     4441 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     4501 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     4561 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     4621 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     4681 ggacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     4741 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     4801 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     4861 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatca
//
