LOCUS       Exported                7149 bp DNA     circular SYN 22-JUN-2017
DEFINITION  .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7149)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 27, 2021 from SnapGene Viewer 5.3.1
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7149
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        81..99
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
                     /note="color: #ffffff; direction: RIGHT"
     protein_bind    100..124
                     /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)."
                     /note="color: #31849b"
     RBS             139..161
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
                     /note="color: #a6acb3"
     CDS             169..2160
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP-DSep2
                     /note="mammalian codon-optimized"
                     /note="This feature has 5 segments:
                        1:  169 ..  171 / #339966
                        2:  172 ..  174 / #339966 / 1a
                        3:  175 ..  885 / #339966 / mEGFP
                        4:  886 ..  900 / #ffff00 / linker
                        5:  901 .. 2160 / #00ff00 / DSep2"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYKGTGSGMSVEVDFVDKKEVHLRTLKQSGHVGFDSLPDQLVNKS
                     VQNGFVFNVMCIGETGLGKSTLMDTLFNTSFESTPSPHTLPSVKLKAHTYELQESNVRL
                     KLTICDTVGYGDQINKDDSFKAVVDYIDAQFENYLQEELKIKRSLVTCHDSRIHICLYF
                     ICPTGHGLKSLDLVCMKKLDSKVNIIPVIAKADTISKVELQRFKAKIIQELNANGVHIY
                     QFPTDDETVAETNTSMNSHIPFAVVGSTEFIKVGNKLIRARQYPWGTVQVENETHCDFV
                     KLREMLIRTNMEDMREKTHTRHYELYRQKRLEQMGFSDVDSDNKPISFQQTFEAKRSNH
                     LAELQSKEEEVRQMFVQRVKEKEAELKESEKDLHAKFEKLKRDHAEEKRKLEESRKALE
                     EDYLDFQRRKQQLATAHHTLTLGKSKKK"
     misc_feature    2172..2177
                     /label=destroyed XbaI/SpeI
                     /note="color: #ccffff"
     RBS             2184..2206
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
                     /note="color: #a6acb3"
     CDS             2214..3857
                     /codon_start=1
                     /label=Peanut-Strep
                     /note="Peanut-Strep"
                     /note="This feature has 3 segments:
                        1: 2214 .. 3830 / #00ff00
                        2: 3831 .. 3854 / #ff9900 / Strep tag
                        3: 3855 .. 3857 / #00ff00"
                     /translation="MNSPRSNAVNGGSGGAISALPSTLAQLALRDKQQAASASASSATN
                     GSSGSESLVGVGGRPPNQPPSVPVAASGKLDTSGGGASNGDSNKLTHDLQEKEHQQAQK
                     PQKPPLPVRQKPMEIAGYVGFANLPNQVYRKAVKRGFEFTLMVVGASGLGKSTLINSMF
                     LSDIYNAEQYPGPSLRKKKTVAVEATKVMLKENGVNLTLTVVDTPGFGDAVDNSNCWVP
                     ILEYVDSKYEEYLTAESRVYRKTISDSRVHCCLYFIAPSGHGLLPLDIACMQSLSDKVN
                     LVPVIAKADTMTPDEVHLFKKQILNEIAQHKIKIYDFPATLEDAAEEAKTTQNLRSRVP
                     FAVVGANTIIEQDGKKVRGRRYPWGLVEVENLTHCDFIALRNMVIRTHLQDLKDVTNNV
                     HYENYRCRKLSELGLVDGKARLSNKNPLTQMEEEKREHEQKMKKMEAEMEQVFDMKVKE
                     KMQKLRDSELELARRHEERKKALELQIRELEEKRREFEREKKEWEDVNHVTLEELKRRS
                     LGANSSTDNVDGKKEKKKKGLFWSHPQFEK"
     terminator      3911..3958
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
                     /note="color: #ffffff"
     CDS             complement(4129..4920)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /note="color: #ccffcc"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        complement(4921..5012)
                     /gene="bla"
                     /label=AmpR promoter
                     /note="color: #ffffff; direction: LEFT"
     rep_origin      complement(5060..5798)
                     /direction=LEFT
                     /label=CloDF13 ori
                     /note="Plasmids containing the CloDF13 (CDF) origin of 
                     replication can be propagated in E. coli cells that contain
                     additional plasmids with compatible origins."
                     /note="color: #ffff00"
     CDS             complement(6008..7090)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /note="color: #993366"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(join(7091..7149,1..19))
                     /gene="lacI"
                     /label=lacI promoter
                     /note="color: #ffffff; direction: LEFT"
ORIGIN
        1 tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg
       61 agatctcgat cccgcgaaat taatacgact cactataggg gaattgtgag cggataacaa
      121 ttccccacta gtaataattt tgtttaactt taagaaggag atatacatat ggtgagcaag
      181 ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac
      241 ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc
      301 ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc
      361 ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc
      421 ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac
      481 ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc
      541 gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac
      601 aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg
      661 aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag
      721 cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc
      781 cagtccaagc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc
      841 gtgaccgccg ccgggatcac tctcggcatg gacgagctgt acaagggtac cggcagcggt
      901 atgtccgttg aggtggattt cgtcgacaag aaggaggtgc acctccgcac cctgaagcag
      961 tccggtcacg tgggattcga cagcttgcct gaccaactgg tcaacaagag tgtccagaat
     1021 ggattcgtct tcaacgtcat gtgtataggc gaaaccggac tgggcaagtc gacgctgatg
     1081 gacacgctgt tcaacaccag cttcgagtcc acgccgagtc cccacacttt gccgagcgtc
     1141 aagttgaagg cacacaccta cgagctgcag gagagcaatg tgcgactcaa gctcaccatc
     1201 tgcgacacgg ttggctacgg cgatcagatc aacaaggacg actcgttcaa ggcggtggtg
     1261 gactacatcg acgcacagtt cgagaactat ctgcaggagg agctgaagat caagcgatcg
     1321 ctggtgacat gccacgacag ccgcatccac atctgcctct actttatttg ccccactggg
     1381 cacggcctca aatcgctgga cctggtgtgc atgaagaagc tggacagcaa ggtgaacata
     1441 attccggtga ttgccaaggc ggataccatt tccaaggttg agctacagcg cttcaaggcg
     1501 aagatcattc aggagctaaa cgccaatgga gtgcacatct accagttccc caccgacgat
     1561 gagacggtgg ccgagaccaa caccagcatg aactcgcaca ttcccttcgc cgtggtgggc
     1621 agcacggagt ttattaaggt gggcaacaag ctcatccgcg ctcgccagta cccctggggc
     1681 acggtgcagg tggagaacga gacgcactgc gactttgtga agctgcgcga gatgctcatt
     1741 cgcaccaaca tggaggatat gcgcgaaaag acacacacgc gccactacga gctgtatcgc
     1801 cagaagaggc tggagcaaat gggcttcagc gatgtggaca gcgacaacaa gcccatctcg
     1861 ttccagcaga cgttcgaggc aaagcgctcc aaccatttgg ccgagctgca gtccaaggag
     1921 gaggaggttc gtcagatgtt cgtgcagcgc gttaaggaga aggaggccga gctgaaggag
     1981 agcgagaagg acctgcacgc caagttcgag aagctaaagc gggatcacgc cgaggagaag
     2041 cgcaagctgg aagagtcgcg caaggcgctc gaggaggact accttgactt ccagcggcgc
     2101 aagcagcagt tggccaccgc ccaccacacg ctcaccctcg gcaagagcaa gaagaagtag
     2161 gatcctaata gtctagtaat aattttgttt aactttaaga aggagatata catatgaata
     2221 gtcctcgctc gaacgcggtt aatggcggca gcggcggcgc catctccgct ctgcccagca
     2281 ctctggccca actggctctg cgcgacaagc agcaggctgc gtcggcatcg gcgtcctctg
     2341 ccacgaacgg cagcagcggc agcgagtcct tggtgggcgt cggtggacga ccgcccaatc
     2401 agccgcccag tgtgccagtg gccgccagtg gcaagctgga caccagcggc ggaggcgcct
     2461 ccaacggcga ctcgaacaaa ctgacccacg atctgcagga gaaggagcac cagcaggcgc
     2521 agaagccaca gaagccaccg ctcccggtgc gccagaagcc catggagatc gccggctatg
     2581 tgggcttcgc caacctgccc aatcaggtct atcgcaaggc cgtcaagcgg ggattcgagt
     2641 tcactctgat ggtggtgggc gccagtgggc tgggcaagtc gacgctgatc aactccatgt
     2701 tcctgtcgga catctacaac gcggaacagt atccgggccc ctcgctgcgc aaaaagaaga
     2761 ccgtggccgt ggaggccacc aaagtaatgc tcaaggagaa cggcgtcaat ctgacactca
     2821 cggtggtgga cacacccgga ttcggtgatg ccgttgacaa cagcaactgc tgggtgccca
     2881 tactggagta cgtggatagc aagtacgagg agtacctgac agccgagtcg cgggtgtacc
     2941 gaaaaaccat ttcggacagc cgggtgcact gctgcctcta cttcatagcg ccatcgggac
     3001 acggactcct gccgctggac attgcctgca tgcagagcct gtcggacaag gtaaatctgg
     3061 tgccggtgat cgccaaggcg gacaccatga cgcccgacga ggtgcatcta tttaagaagc
     3121 agatccttaa cgagatcgcc cagcacaaga tcaagatcta cgacttcccc gccacgctgg
     3181 aggatgcggc cgaggaggcc aaaaccacgc agaatttgcg cagccgagtg ccattcgcgg
     3241 tggtgggtgc caacaccatc atcgagcagg acggaaagaa ggtccggggt aggcgttatc
     3301 cctggggcct ggtggaggtg gagaacctga cgcattgcga ctttatagcg ctgcgcaaca
     3361 tggtcatacg cacgcacctt caggacctca aggacgtgac gaacaacgtc cactacgaga
     3421 actaccgctg ccggaagcta tccgagctgg gattggtgga cggcaaggcc aggctgtcca
     3481 acaagaaccc actcacccag atggaggagg aaaagcggga gcacgagcag aagatgaaga
     3541 agatggaggc cgagatggag caggtgtttg acatgaaggt caaggagaag atgcagaagc
     3601 tcagggactc ggagctggag ctggcgcggc gccacgagga gcgcaagaag gcgctggagc
     3661 tgcagatccg ggagctggag gagaagagac gcgagttcga gcgcgagaag aaggagtggg
     3721 aggatgtcaa ccacgtgacg ctcgaggagc tcaagcgacg cagcctgggg gccaacagca
     3781 gcacggacaa tgtggatggc aagaaggaga agaaaaagaa gggtctgttc tggagccacc
     3841 cgcagttcga aaaataagga tcctaatagt ctagagctag ccctaggctg ctgccaccgc
     3901 tgagcaataa ctagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgct
     3961 gaaacctcag gcatttgaga agcacacggt cacactgctt ccggtagtca ataaaccggt
     4021 aaaccagcaa tagacataag cggctattta acgaccctgc cctgaaccga cgaccgggtc
     4081 atcgtggccg gatcttgcgg cccctcggct tgaacgaatt gttagacatt atttgccgac
     4141 taccttggtg atctcgcctt tcacgtagtg gacaaattct tccaactgat ctgcgcgcga
     4201 ggccaagcga tcttcttctt gtccaagata agcctgtcta gcttcaagta tgacgggctg
     4261 atactgggcc ggcaggcgct ccattgccca gtcggcagcg acatccttcg gcgcgatttt
     4321 gccggttact gcgctgtacc aaatgcggga caacgtaagc actacatttc gctcatcgcc
     4381 agcccagtcg ggcggcgagt tccatagcgt taaggtttca tttagcgcct caaatagatc
     4441 ctgttcagga accggatcaa agagttcctc cgccgctgga cctaccaagg caacgctatg
     4501 ttctcttgct tttgtcagca agatagccag atcaatgtcg atcgtggctg gctcgaagat
     4561 acctgcaaga atgtcattgc gctgccattc tccaaattgc agttcgcgct tagctggata
     4621 acgccacgga atgatgtcgt cgtgcacaac aatggtgact tctacagcgc ggagaatctc
     4681 gctctctcca ggggaagccg aagtttccaa aaggtcgttg atcaaagctc gccgcgttgt
     4741 ttcatcaagc cttacggtca ccgtaaccag caaatcaata tcactgtgtg gcttcaggcc
     4801 gccatccact gcggagccgt acaaatgtac ggccagcaac gtcggttcga gatggcgctc
     4861 gatgacgcca actacctctg atagttgagt cgatacttcg gcgatcaccg cttccctcat
     4921 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     4981 catatttgaa tgtatttaga aaaataaaca aatagctagg tcactcggtc gctacgctcc
     5041 gggcgtgaga ctgcggcggg cgctgcggac acatacaaag ttacccacag attccgtgga
     5101 taagcagggg actaacatgt gaggcaaaac agcagggccg cgccggtggc gtttttccat
     5161 aggctccgcc ctcctgccag agttcacata aacagacgct tttccggtgc atctgtggga
     5221 gccgtgaggc tcaaccatga atctgacagt acgggcgaaa cccgacagga cttaaagatc
     5281 cccaccgttt ccggcgggtc gctccctctt gcgctctcct gttccgaccc tgccgtttac
     5341 cggatacctg ttccgccttt ctcccttacg ggaagtgtgg cgctttctca tagctcacac
     5401 actggtatct cggctcggtg taggtcgttc gctccaagct gggctgtaag caagaactcc
     5461 ccgttcagcc cgactgctgc gccttatccg gtaactgttc acttgagtcc aacccggaaa
     5521 agcacggtaa aacgccactg gcagcagcca ttggtaactg ggagttcgca gaggatttgt
     5581 ttagctaaac acgcggttgc tcttgaagtg tgcgccaaag tccggctaca ctggaaggac
     5641 agatttggtt gctgtgctct gcgaaagcca gttaccacgg ttaagcagtt ccccaactga
     5701 cttaaccttc gatcaaacca cctccccagg tggttttttc gtttacaggg caaaagatta
     5761 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctactgaa ccgctcttga
     5821 tttcagtgca atttatctct tcaaatgtag cacctgaagt cagccccata cgatataagt
     5881 tgtaattctc atgttagtca tgccccgcgc ccaccggaag gagctgactg ggttgaaggc
     5941 tctcaagggc atcggtcgag atcccggtgc ctaatgagtg agctaactta cattaattgc
     6001 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     6061 cggccaacgc gcggggagag gcggtttgcg tattgggcgc cagggtggtt tttcttttca
     6121 ccagtgagac gggcaacagc tgattgccct tcaccgcctg gccctgagag agttgcagca
     6181 agcggtccac gctggtttgc cccagcaggc gaaaatcctg tttgatggtg gttaacggcg
     6241 ggatataaca tgagctgtct tcggtatcgt cgtatcccac taccgagatg tccgcaccaa
     6301 cgcgcagccc ggactcggta atggcgcgca ttgcgcccag cgccatctga tcgttggcaa
     6361 ccagcatcgc agtgggaacg atgccctcat tcagcatttg catggtttgt tgaaaaccgg
     6421 acatggcact ccagtcgcct tcccgttccg ctatcggctg aatttgattg cgagtgagat
     6481 atttatgcca gccagccaga cgcagacgcg ccgagacaga acttaatggg cccgctaaca
     6541 gcgcgatttg ctggtgaccc aatgcgacca gatgctccac gcccagtcgc gtaccgtctt
     6601 catgggagaa aataatactg ttgatgggtg tctggtcaga gacatcaaga aataacgccg
     6661 gaacattagt gcaggcagct tccacagcaa tggcatcctg gtcatccagc ggatagttaa
     6721 tgatcagccc actgacgcgt tgcgcgagaa gattgtgcac cgccgcttta caggcttcga
     6781 cgccgcttcg ttctaccatc gacaccacca cgctggcacc cagttgatcg gcgcgagatt
     6841 taatcgccgc gacaatttgc gacggcgcgt gcagggccag actggaggtg gcaacgccaa
     6901 tcagcaacga ctgtttgccc gccagttgtt gtgccacgcg gttgggaatg taattcagct
     6961 ccgccatcgc cgcttccact ttttcccgcg ttttcgcaga aacgtggctg gcctggttca
     7021 ccacgcggga aacggtctga taagagacac cggcatactc tgcgacatcg tataacgtta
     7081 ctggtttcac attcaccacc ctgaattgac tctcttccgg gcgctatcat gccataccgc
     7141 gaaaggttt
//
