LOCUS       Exported                6275 bp ds-DNA     circular SYN 20-DEC-2019
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6275)
  AUTHORS   Doudna Lab
  TITLE     Direct Submission
  JOURNAL   Exported Friday, Dec 20, 2019 from SnapGene 4.3.11
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..6275
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          3599..3901
                     /organism="Staphylococcus simulans"
                     /host="bovine"
                     /strain="19"
                     /mol_type="genomic DNA"
                     /country="Finland: Helsinki"
                     /collection_date=2012
                     /note="color: #ffffff"
                     /db_xref="taxon:1286"
     misc_feature    233..375
                     /label=bom
                     /note="basis of mobility region from pBR322"
                     /note="color: #a6acb3"
     CDS             complement(477..668)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /note="color: #993366"
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     promoter        2229..2247
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     RBS             2336..2358
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
                     /note="color: #a6acb3"
     CDS             2366..2407
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /note="color: #cc99b2"
                     /translation="MKSSHHHHHHHHHH"
     CDS             2417..3517
                     /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."
                     /note="color: #cc99b2"
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     CDS             3572..3592
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /note="color: #cc99b2
                     Cleavage site after base 3589"
                     /translation="ENLYFQS"
     CDS             3599..3901
                     /codon_start=1
                     /label=truncated AcrIIA14
                     /note="color: #993366"
                     /translation="MKSVKYISNMSKQEKGYRVYVNVVNEDTDKGFLFPSVPKEVIEND
                     KIDELFNFEHHKPYVQKAKSRYDKNGIGYKIVQLDEGFQKFIELNKEKMKENLDY"
     misc_feature    3902..3906
                     /label=Islander Island
                     /note="color: #a6acb3"
     misc_feature    3902..3906
                     /label=Comparator Island
                     /note="color: #00ff00"
     gene            3902..3906
                     /locus_tag="AAX06_RS01960"
                     /old_locus_tag="AAX06_01960"
                     /label=AAX06_RS01960
                     /note="color: #a6acb3; direction: RIGHT"
     misc_feature    3902..3906
                     /label=PHASTER prophage region
                     /note="color: #a6acb3"
     terminator      4044..4091
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
                     /note="color: #ffffff"
     promoter        complement(4457..4485)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
                     /note="This reverse directional feature has 3 segments:
                        1: 4457 .. 4462 / #ffffff / -10
                        2: 4463 .. 4479 / #ffffff
                        3: 4480 .. 4485 / #ffffff / -35"
     promoter        4598..4702
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: RIGHT"
     CDS             4703..5563
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /note="This feature has 2 segments:
                        1: 4703 .. 4771 / #ccffcc / signal sequence
                        2: 4772 .. 5563 / #ccffcc
                     Cleavage site after base 4771"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      join(5734..6275,1..47)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
                     /note="color: #ffff00"
ORIGIN
        1 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
       61 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
      121 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
      181 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcctgatgc
      241 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catatatggt gcactctcag
      301 tacaatctgc tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac
      361 tgggtcatgg ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
      421 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
      481 aggttttcac cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg
      541 tcgtgaagcg attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc
      601 agaagcgtta atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt
      661 ttggtcactg atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg
      721 aaacgagaga ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa
      781 cgttgtgagg gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag
      841 ggtcaatgcc agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat
      901 cctgcgatgc agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt
      961 tacgaaacac ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag
     1021 cagcagtcgc ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa
     1081 ccccgccagc ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggccag
     1141 gacccaacgc tgcccgagat gcgccgcgtg cggctgctgg agatggcgga cgcgatggat
     1201 atgttctgcc aagggttggt ttgcgcattc acagttctcc gcaagaattg attggctcca
     1261 attcttggag tggtgaatcc gttagcgagg tgccgccggc ttccattcag gtcgaggtgg
     1321 cccggctcca tgcaccgcga cgcaacgcgg ggaggcagac aaggtatagg gcggcgccta
     1381 caatccatgc caacccgttc catgtgctcg ccgaggcggc ataaatcgcc gtgacgatca
     1441 gcggtccagt gatcgaagtt aggctggtaa gagccgcgag cgatccttga agctgtccct
     1501 gatggtcgtc atctacctgc ctggacagca tggcctgcaa cgcgggcatc ccgatgccgc
     1561 cggaagcgag aagaatcata atggggaagg ccatccagcc tcgcgtcgcg aacgccagca
     1621 agacgtagcc cagcgcgtcg gccgccatgc cggcgataat ggcctgcttc tcgccgaaac
     1681 gtttggtggc gggaccagtg acgaaggctt gagcgagggc gtgcaagatt ccgaataccg
     1741 caagcgacag gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc
     1801 agagcgctgc cggcacctgt cctacgagtt gcatgataaa gaagacagtc ataagtgcgg
     1861 cgacgatagt catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg
     1921 gcatcggtcg acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt
     1981 tgaggccgtt gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca
     2041 gtcccccggc cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga
     2101 agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac
     2161 ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc
     2221 cgcgaaatta atacgactca ctatagggag accacaacgg tttccctcta gtgccggctc
     2281 cggagagctc tttaattaag cggccgccct gcaggactcg agttctagaa ataattttgt
     2341 ttaactttaa gaaggagata tacatatgaa atcttctcac catcaccatc accatcacca
     2401 tcaccatggt tcttctatga aaatcgaaga aggtaaactg gtaatctgga ttaacggcga
     2461 taaaggctat aacggtctcg ctgaagtcgg taagaaattc gagaaagata ccggaattaa
     2521 agtcaccgtt gagcatccgg ataaactgga agagaaattc ccacaggttg cggcaactgg
     2581 cgatggccct gacattatct tctgggcaca cgaccgcttt ggtggctacg ctcaatctgg
     2641 cctgttggct gaaatcaccc cggacaaagc gttccaggac aagctgtatc cgtttacctg
     2701 ggatgccgta cgttacaacg gcaagctgat tgcttacccg atcgctgttg aagcgttatc
     2761 gctgatttat aacaaagatc tgctgccgaa cccgccaaaa acctgggaag agatcccggc
     2821 gctggataaa gaactgaaag cgaaaggtaa gagcgcgctg atgttcaacc tgcaagaacc
     2881 gtacttcacc tggccgctga ttgctgctga cgggggttat gcgttcaagt atgaaaacgg
     2941 caagtacgac attaaagacg tgggcgtgga taacgctggc gcgaaagcgg gtctgacctt
     3001 cctggttgac ctgattaaaa acaaacacat gaatgcagac accgattact ccatcgcaga
     3061 agctgccttt aataaaggcg aaacagcgat gaccatcaac ggcccgtggg catggtccaa
     3121 catcgacacc agcaaagtga attatggtgt aacggtactg ccgaccttca agggtcaacc
     3181 atccaaaccg ttcgttggcg tgctgagcgc aggtattaac gccgccagtc cgaacaaaga
     3241 gctggcaaaa gagttcctcg aaaactatct gctgactgat gaaggtctgg aagcggttaa
     3301 taaagacaaa ccgctgggtg ccgtagcgct gaagtcttac gaggaagagt tggcgaaaga
     3361 tccacgtatt gccgccacta tggaaaacgc ccagaaaggt gaaatcatgc cgaacatccc
     3421 gcagatgtcc gctttctggt atgccgtgcg tactgcggtg atcaacgccg ccagcggtcg
     3481 tcagactgtc gatgaagccc tgaaagacgc gcagactaat tcgagctcga acaacaacaa
     3541 caataacaat aacaacaacc tcgggatcga ggaaaacctg tacttccaat ccaatgcaat
     3601 gaagagtgta aaatatataa gtaatatgag taaacaagaa aaaggttata gagtttatgt
     3661 gaatgtcgtg aacgaagata ctgacaaagg gtttttgttc cctagcgtcc ctaaagaagt
     3721 tatcgagaat gacaaaattg acgaattatt taactttgaa catcataaac catatgttca
     3781 aaaagctaaa tcaagatacg ataaaaatgg cattggatat aaaattgttc aattagatga
     3841 aggttttcaa aaattcatcg aattgaataa agaaaaaatg aaagagaatt tggattatta
     3901 ataataacat tggaagtgga taacggatcc gcgatcgcgg cgcgccacct ggtggccggc
     3961 cggtaccacg cgtgcgcgct gatccggctg ctaacaaagc ccgaaaggaa gctgagttgg
     4021 ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa cgggtcttga
     4081 ggggtttttt gctgaaagga ggaactatat ccggatatcc acaggacggg tgtggtcgcc
     4141 atgatcgcgt agtcgatagt ggctccaagt agcgaagcga gcaggactgg gcggcggcca
     4201 aagcggtcgg acagtgctcc gagaacgggt gcgcatagaa attgcatcaa cgcatatagc
     4261 gctagcagca cgccatagtg actggcgatg ctgtcggaat ggacgatatc ccgcaagagg
     4321 cccggcagta ccggcataac caagcctatg cctacagcat ccagggtgac ggtgccgagg
     4381 atgacgatga gcgcattgtt agatttcata cacggtgcct gactgcgtta gcaatttaac
     4441 tgtgataaac taccgcatta aagcttatcg atgataagct gtcaaacatg agaattcttg
     4501 aagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt
     4561 ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     4621 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     4681 ataacattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     4741 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     4801 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     4861 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     4921 gctatgtggc gcggtattat cccgtgttga cgccgggcaa gagcaactcg gtcgccgcat
     4981 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     5041 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     5101 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     5161 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     5221 cgacgagcgt gacaccacga tgcctgcagc aatggcaaca acgttgcgca aactattaac
     5281 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     5341 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     5401 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     5461 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     5521 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     5581 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     5641 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     5701 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     5761 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     5821 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     5881 ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     5941 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     6001 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     6061 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     6121 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     6181 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     6241 ttatagtcct gtcgggtttc gccacctctg acttg
//
