LOCUS       Exported                8930 bp ds-DNA     circular SYN 21-MAR-2019
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8930)
  AUTHORS   Trial User
  TITLE     Direct Submission
  JOURNAL   Exported Apr 12, 2019 from SnapGene 4.2.6
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8930
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        671..689
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             778..800
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             820..849
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             859..1959
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     CDS             2014..2034
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2041..5001
                     /codon_start=1
                     /label=DpbCasX
                     /translation="MEKRINKIRKKLSADNATKPVSRSGPMKTLLVRVMTDDLKKRLEK
                     RRKKPEVMPQVISNNAANNLRMLLDDYTKMKEAILQVYWQEFKDDHVGLMCKFAQPASK
                     KIDQNKLKPEMDEKGNLTTAGFACSQCGQPLFVYKLEQVSEKGKAYTNYFGRCNVAEHE
                     KLILLAQLKPEKDSDEAVTYSLGKFGQRALDFYSIHVTKESTHPVKPLAQIAGNRYASG
                     PVGKALSDACMGTIASFLSKYQDIIIEHQKVVKGNQKRLESLRELAGKENLEYPSVTLP
                     PQPHTKEGVDAYNEVIARVRMWVNLNLWQKLKLSRDDAKPLLRLKGFPSFPVVERRENE
                     VDWWNTINEVKKLIDAKRDMGRVFWSGVTAEKRNTILEGYNYLPNENDHKKREGSLENP
                     KKPAKRQFGDLLLYLEKKYAGDWGKVFDEAWERIDKKIAGLTSHIEREEARNAEDAQSK
                     AVLTDWLRAKASFVLERLKEMDEKEFYACEIQLQKWYGDLRGNPFAVEAENRVVDISGF
                     SIGSDGHSIQYRNLLAWKYLENGKREFYLLMNYGKKGRIRFTDGTDIKKSGKWQGLLYG
                     GGKAKVIDLTFDPDDEQLIILPLAFGTRQGREFIWNDLLSLETGLIKLANGRVIEKTIY
                     NKKIGRDEPALFVALTFERREVVDPSNIKPVNLIGVARGENIPAVIALTDPEGCPLPEF
                     KDSSGGPTDILRIGEGYKEKQRAIQAAKEVEQRRAGGYSRKFASKSRNLADDMVRNSAR
                     DLFYHAVTHDAVLVFANLSRGFGRQGKRTFMTERQYTKMEDWLTAKLAYEGLTSKTYLS
                     KTLAQYTSKTCSNCGFTITTADYDGMLVRLKKTSDGWATTLNNKELKAEGQITYYNRYK
                     RQTVEKELSAELDRLSEESGNNDISKWTKGRRDEALFLLKKRFSHRPVQEQFVCLDCGH
                     EVHAAEQAALNIARSWLFLNSNSTEFKSYKSGKQPFVGAWQAFYKRRLKEVWKPNA"
     misc_feature    4054..4056
                     /label=D672A
     misc_feature    4345..4347
                     /label=E769A
     misc_feature    4843..4845
                     /label=D935A
     terminator      5141..5188
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(5554..5582)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     promoter        5695..5799
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5800..6660
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      6831..7419
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    7605..7747
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(7849..8040)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 gccggaagcg agaagaatca taatggggaa ggccatccag cctcgcgtcg cgaacgccag
       61 caagacgtag cccagcgcgt cggccgccat gccggcgata atggcctgct tctcgccgaa
      121 acgtttggtg gcgggaccag tgacgaaggc ttgagcgagg gcgtgcaaga ttccgaatac
      181 cgcaagcgac aggccgatca tcgtcgcgct ccagcgaaag cggtcctcgc cgaaaatgac
      241 ccagagcgct gccggcacct gtcctacgag ttgcatgata aagaagacag tcataagtgc
      301 ggcgacgata gtcatgcccc gcgcccaccg gaaggagctg actgggttga aggctctcaa
      361 gggcatcggt cgacgctctc ccttatgcga ctcctgcatt aggaagcagc ccagtagtag
      421 gttgaggccg ttgagcaccg ccgccgcaag gaatggtgca tgcaaggaga tggcgcccaa
      481 cagtcccccg gccacggggc ctgccaccat acccacgccg aaacaagcgc tcatgagccc
      541 gaagtggcga gcccgatctt ccccatcggt gatgtcggcg atataggcgc cagcaaccgc
      601 acctgtggcg ccggtgatgc cggccacgat gcgtccggcg tagaggatcg agatctcgat
      661 cccgcgaaat taatacgact cactataggg agaccacaac ggtttccctc tagtgccggc
      721 tccggagagc tctttaatta agcggccgcc ctgcaggact cgagttctag aaataatttt
      781 gtttaacttt aagaaggaga tatacatatg aaatcttctc accatcacca tcaccatcac
      841 catcaccatg gttcttctat gaaaatcgaa gaaggtaaac tggtaatctg gattaacggc
      901 gataaaggct ataacggtct cgctgaagtc ggtaagaaat tcgagaaaga taccggaatt
      961 aaagtcaccg ttgagcatcc ggataaactg gaagagaaat tcccacaggt tgcggcaact
     1021 ggcgatggcc ctgacattat cttctgggca cacgaccgct ttggtggcta cgctcaatct
     1081 ggcctgttgg ctgaaatcac cccggacaaa gcgttccagg acaagctgta tccgtttacc
     1141 tgggatgccg tacgttacaa cggcaagctg attgcttacc cgatcgctgt tgaagcgtta
     1201 tcgctgattt ataacaaaga tctgctgccg aacccgccaa aaacctggga agagatcccg
     1261 gcgctggata aagaactgaa agcgaaaggt aagagcgcgc tgatgttcaa cctgcaagaa
     1321 ccgtacttca cctggccgct gattgctgct gacgggggtt atgcgttcaa gtatgaaaac
     1381 ggcaagtacg acattaaaga cgtgggcgtg gataacgctg gcgcgaaagc gggtctgacc
     1441 ttcctggttg acctgattaa aaacaaacac atgaatgcag acaccgatta ctccatcgca
     1501 gaagctgcct ttaataaagg cgaaacagcg atgaccatca acggcccgtg ggcatggtcc
     1561 aacatcgaca ccagcaaagt gaattatggt gtaacggtac tgccgacctt caagggtcaa
     1621 ccatccaaac cgttcgttgg cgtgctgagc gcaggtatta acgccgccag tccgaacaaa
     1681 gagctggcaa aagagttcct cgaaaactat ctgctgactg atgaaggtct ggaagcggtt
     1741 aataaagaca aaccgctggg tgccgtagcg ctgaagtctt acgaggaaga gttggcgaaa
     1801 gatccacgta ttgccgccac tatggaaaac gcccagaaag gtgaaatcat gccgaacatc
     1861 ccgcagatgt ccgctttctg gtatgccgtg cgtactgcgg tgatcaacgc cgccagcggt
     1921 cgtcagactg tcgatgaagc cctgaaagac gcgcagacta attcgagctc gaacaacaac
     1981 aacaataaca ataacaacaa cctcgggatc gaggaaaacc tgtacttcca atccaatgca
     2041 atggagaagc gtattaacaa gattcgtaag aagttatcgg ctgataatgc cacaaaacct
     2101 gtaagccgct caggacctat gaaaacgctt ttagttcgtg ttatgacaga tgatctgaaa
     2161 aaacgtcttg agaagcgtcg taaaaagccg gaggtcatgc ctcaagtcat ttcgaataat
     2221 gccgccaata atttacgtat gcttcttgat gactatacaa agatgaaaga agcaatcctg
     2281 caagtctact ggcaagagtt caaagatgat catgtaggtt tgatgtgtaa gtttgcacag
     2341 cccgctagta aaaagatcga ccaaaacaag ttaaagccag agatggacga gaaagggaat
     2401 cttaccactg ccgggttcgc ttgctctcaa tgcggccaac cgctgttcgt atataagctg
     2461 gagcaagttt cagaaaaggg taaggcatac acaaattatt tcggacgttg caacgtcgct
     2521 gagcacgaga agcttattct tttagcccag ctgaaaccag agaaggactc tgatgaagcg
     2581 gttacctact ccttgggaaa gttcggacag cgcgctcttg acttttattc catccacgtg
     2641 acaaaggaaa gtacacaccc cgtcaaacca ctggcccaaa ttgcagggaa ccgctacgcg
     2701 agcgggccag tgggaaaagc attgtcagac gcttgtatgg gcacgattgc ttcctttctt
     2761 agtaaatacc aggatattat cattgaacat caaaaggtag ttaaggggaa ccagaagcgt
     2821 ctggaaagct tacgtgaact tgcagggaaa gagaaccttg agtacccatc ggtcacattg
     2881 cctccgcagc cgcacacaaa ggagggtgtc gatgcttata acgaggtgat tgcacgcgta
     2941 cgcatgtggg taaatttgaa tctgtggcag aagttgaaac tgtcgcgtga tgacgccaaa
     3001 cctcttttac gcctgaaggg cttcccctca tttccagtcg tagagcgccg tgaaaatgaa
     3061 gtagactggt ggaatactat caacgaagta aaaaagctga ttgacgcgaa gcgcgacatg
     3121 ggacgcgtgt tttggagcgg cgttactgct gaaaagcgca acaccatcct tgaaggatac
     3181 aactatttac ccaacgagaa cgatcataaa aagcgcgagg gcagtttaga gaatccgaag
     3241 aagccggcca aacgtcagtt tggcgacctt ttattatacc tggaaaagaa atatgcgggt
     3301 gactggggca aagtgttcga tgaagcctgg gagcgcattg acaagaaaat cgctggtctg
     3361 acgtcgcaca ttgagcgcga ggaagctcgt aacgcagaag acgcccaatc caaagcggtc
     3421 ctgacagact ggctgcgcgc taaggcgtcc tttgttcttg agcgtttaaa agagatggac
     3481 gaaaaggaat tctatgcctg tgagattcaa ttacaaaagt ggtatggtga cttacgcggg
     3541 aacccctttg ctgtagaggc tgagaatcgt gtagtggaca tttcaggttt cagcatcgga
     3601 tcggatggac attctatcca atatcgtaac cttcttgcat ggaaatattt ggagaatggt
     3661 aagcgcgagt tctatctgct tatgaactac gggaaaaaag gacgtattcg cttcaccgat
     3721 ggtacagaca ttaaaaaaag tggcaaatgg caagggttgt tatatggggg tggcaaagca
     3781 aaggttatcg atttaacgtt tgatcccgat gatgagcagc tgattatctt gcctttagct
     3841 ttcggaaccc gccagggccg tgaatttatc tggaacgact tgctttctct tgagacagga
     3901 cttatcaaac ttgcgaatgg acgcgtgatc gaaaagacta tttacaataa aaagatcggt
     3961 cgcgatgagc cggccctgtt tgtcgcctta acattcgagc gtcgcgaagt tgttgatccg
     4021 tcaaatatca agccggtaaa cttaatcggg gtcgcccgtg gtgaaaatat cccggcagta
     4081 atcgctctta cagacccaga aggctgcccc ttacccgagt tcaaggactc atccggcggt
     4141 ccaacagaca tccttcgcat tggcgaagga tataaagaga aacagcgtgc aatccaggca
     4201 gcaaaagaag tggaacaacg tcgcgccgga ggctacagcc gcaaatttgc gtcaaaatcg
     4261 cgtaacttgg cagatgatat ggtccgcaac agcgcccgtg atttgtttta ccacgcggtg
     4321 acacatgacg ctgttttagt ttttgccaac ttgtcacgtg ggttcggacg tcagggaaaa
     4381 cgcaccttca tgactgagcg ccaatacaca aaaatggaag actggttgac ggctaaactg
     4441 gcttacgagg ggcttactag taaaacgtat ttgtcgaaaa ctcttgctca gtacacctct
     4501 aaaacttgtt ctaactgcgg gttcacgatt accacagcgg actacgatgg catgctggta
     4561 cgccttaaaa agacgagcga cgggtgggca acgacattga acaataaaga acttaaggcc
     4621 gagggacaaa tcacctacta taatcgttat aagcgccaga cggtagaaaa agagttaagc
     4681 gccgaacttg accgcctgag cgaggagagt gggaacaacg acatttcaaa atggactaag
     4741 gggcgccgtg acgaggcttt gtttttgctg aagaaacgtt tctcccatcg tcccgtacaa
     4801 gagcagttcg tttgcttaga ttgtggccat gaagttcatg ctgccgaaca ggctgcttta
     4861 aatatcgcgc gttcctggtt gtttcttaat agtaactcaa ctgagttcaa atcatataag
     4921 tcagggaaac agccgtttgt gggagcatgg caggcttttt ataaacgccg cctgaaggag
     4981 gtctggaagc cgaatgcttg ataacattgg aagtggataa cggatccgcg atcgcggcgc
     5041 gccacctggt ggccggccgg taccacgcgt gcgcgctgat ccggctgcta acaaagcccg
     5101 aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc
     5161 ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg gatatccaca
     5221 ggacgggtgt ggtcgccatg atcgcgtagt cgatagtggc tccaagtagc gaagcgagca
     5281 ggactgggcg gcggccaaag cggtcggaca gtgctccgag aacgggtgcg catagaaatt
     5341 gcatcaacgc atatagcgct agcagcacgc catagtgact ggcgatgctg tcggaatgga
     5401 cgatatcccg caagaggccc ggcagtaccg gcataaccaa gcctatgcct acagcatcca
     5461 gggtgacggt gccgaggatg acgatgagcg cattgttaga tttcatacac ggtgcctgac
     5521 tgcgttagca atttaactgt gataaactac cgcattaaag cttatcgatg ataagctgtc
     5581 aaacatgaga attcttgaag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     5641 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     5701 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     5761 ccctgataaa tgcttcaata acattgaaaa aggaagagta tgagtattca acatttccgt
     5821 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     5881 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     5941 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     6001 agcactttta aagttctgct atgtggcgcg gtattatccc gtgttgacgc cgggcaagag
     6061 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     6121 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     6181 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     6241 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     6301 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgcagcaat ggcaacaacg
     6361 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     6421 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     6481 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     6541 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     6601 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     6661 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     6721 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     6781 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     6841 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     6901 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     6961 cagataccaa atactgtcct tctagtgtag ccgtagttag gccaccactt caagaactct
     7021 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     7081 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     7141 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     7201 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     7261 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     7321 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     7381 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     7441 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     7501 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     7561 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcctgat gcggtatttt
     7621 ctccttacgc atctgtgcgg tatttcacac cgcatatatg gtgcactctc agtacaatct
     7681 gctctgatgc cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat
     7741 ggctgcgccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     7801 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     7861 accgtcatca ccgaaacgcg cgaggcagct gcggtaaagc tcatcagcgt ggtcgtgaag
     7921 cgattcacag atgtctgcct gttcatccgc gtccagctcg ttgagtttct ccagaagcgt
     7981 taatgtctgg cttctgataa agcgggccat gttaagggcg gttttttcct gtttggtcac
     8041 tgatgcctcc gtgtaagggg gatttctgtt catgggggta atgataccga tgaaacgaga
     8101 gaggatgctc acgatacggg ttactgatga tgaacatgcc cggttactgg aacgttgtga
     8161 gggtaaacaa ctggcggtat ggatgcggcg ggaccagaga aaaatcactc agggtcaatg
     8221 ccagcgcttc gttaatacag atgtaggtgt tccacagggt agccagcagc atcctgcgat
     8281 gcagatccgg aacataatgg tgcagggcgc tgacttccgc gtttccagac tttacgaaac
     8341 acggaaaccg aagaccattc atgttgttgc tcaggtcgca gacgttttgc agcagcagtc
     8401 gcttcacgtt cgctcgcgta tcggtgattc attctgctaa ccagtaaggc aaccccgcca
     8461 gcctagccgg gtcctcaacg acaggagcac gatcatgcgc acccgtggcc aggacccaac
     8521 gctgcccgag atgcgccgcg tgcggctgct ggagatggcg gacgcgatgg atatgttctg
     8581 ccaagggttg gtttgcgcat tcacagttct ccgcaagaat tgattggctc caattcttgg
     8641 agtggtgaat ccgttagcga ggtgccgccg gcttccattc aggtcgaggt ggcccggctc
     8701 catgcaccgc gacgcaacgc ggggaggcag acaaggtata gggcggcgcc tacaatccat
     8761 gccaacccgt tccatgtgct cgccgaggcg gcataaatcg ccgtgacgat cagcggtcca
     8821 gtgatcgaag ttaggctggt aagagccgcg agcgatcctt gaagctgtcc ctgatggtcg
     8881 tcatctacct gcctggacag catggcctgc aacgcgggca tcccgatgcc
//
