LOCUS       pAI4_Cam                7586 bp ds-DNA     circular SYN 18-APR-2017
SOURCE      synthetic DNA construct
  ORGANISM  recombinant plasmid
REFERENCE   1  (bases 1 to 7586)
  AUTHORS   Morten Norholm Group
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, Apr 25, 2017 from SnapGene 3.3.3 to Vector NTI(R) 
            format
            http://www.snapgene.com
COMMENT     VNTDATE|803347200|
COMMENT     VNTDBDATE|803347200|
COMMENT     VNTNAME|pAI4_Cam|
COMMENT     VNTAUTHORNAME|Morten Norholm Group|
FEATURES             Location/Qualifiers
     terminator      476..523
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             657..1280
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
     misc_feature    1385..1389
                     /label=Doublicate
     terminator      1398..1492
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(1513..2172)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
     promoter        complement(2173..2275)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     rep_origin      complement(2338..2973)
                     /direction=LEFT
                     /label=ColA ori
                     /note="Plasmids containing the ColA origin of replication 
                     can be propagated in E. coli cells that contain additional 
                     plasmids with compatible origins."
     misc_RNA        complement(3222..3300)
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
     repeat_region   3472..3507
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   3538..3573
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   3604..3639
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   3670..3705
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   3736..3771
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     protein_bind    3839..3857
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3864..3882
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             3907..427
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
ORIGIN
        1 tctgccaagc aaatatgtga attttttata tttagctagt cattatgaaa agttgaaggg
       61 tagtccagaa gataacgaac aaaaacaatt gtttgtggag cagcataagc attatttaga
      121 tgagattatt gagcaaatca gtgaattttc taagcgtgtt attttagcag atgccaattt
      181 agataaagtt cttagtgcat ataacaaaca tagagacaaa ccaatacgtg aacaagcaga
      241 aaatattatt catttattta cgttgacgaa tcttggagct cccgctgctt ttaaatattt
      301 tgatacaaca attgatcgta aacgatatac gtctacaaaa gaagttttag atgccactct
      361 tatccatcaa tccatcactg gtctttatga aacacgcatt gatttgagtc agctaggagg
      421 tgactgagat cccatgttac cggtatccta ggctgctgcc accgctgagc aataactagc
      481 ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgctgaaac ctcaggcatt
      541 tgagaagcac acggtcacac tgcttccggt agtcaataaa ccggtattga cgccaacggg
      601 tcacacgggt acagtgctag cagaaacata tccatgaaat cccgccccga attcatatgt
      661 ctagattaga taaaagtaaa gtgattaaca gcgcattaga gctgcttaat gaggtcggaa
      721 tcgaaggttt aacaacccgt aaactcgccc agaagctagg tgtagagcag cctacattgt
      781 attggcatgt aaaaaataag cgggctttgc tcgacgcctt agccattgag atgttagata
      841 ggcaccatac tcacttttgc cctttagaag gggaaagctg gcaagatttt ttacgtaata
      901 aggctaaaag ttttagatgt gctttactaa gtcatcgcga tggagcaaaa gtacatttag
      961 gtacacggcc tacagaaaaa cagtatgaaa ctctcgaaaa tcaattagcc tttttatgcc
     1021 aacaaggttt ttcactagag aatgcattat atgcactcag cgctgtgggg cattttactt
     1081 taggttgcgt attggaagat caagagcatc aagtcgctaa agaagaaagg gaaacaccta
     1141 ctactgatag tatgccgcca ttattacgac aagctatcga attatttgat caccaaggtg
     1201 cagagccagc cttcttattc ggccttgaat tgatcatatg cggattagaa aaacaactta
     1261 aatgtgaaag tgggtcttaa gcaatagaca taagcggcta tttaacgacc ctgccctgaa
     1321 ccgacgacaa gctgacgacc gggtctccgc aagtggcact tttcggggaa atgtgcgcgg
     1381 aacccctatg tgcttggatt ctcaccaata aaaaacgccc ggcggcaacc gagcgttctg
     1441 aacaaatcca gatggagttc tgaggtcatt actggatcta tcaacaggag tccaagcgag
     1501 ctcgatatca aattacgccc cgccctgcca ctcatcgcag tactgttgta attcattaag
     1561 cattctgccg acatggaagc catcacagac ggcatgatga acctgaatcg ccagcggcat
     1621 cagcaccttg tcgccttgcg tataatattt gcccatggtg aaaacggggg cgaagaagtt
     1681 gtccatattg gccacgttta aatcaaaact ggtgaaactc acccagggat tggctgagac
     1741 gaaaaacata ttctcaataa accctttagg gaaataggcc aggttttcac cgtaacacgc
     1801 cacatcttgc gaatatatgt gtagaaactg ccggaaatcg tcgtggtatt cactccagag
     1861 cgatgaaaac gtttcagttt gctcatggaa aacggtgtaa caagggtgaa cactatccca
     1921 tatcaccagc tcaccgtctt tcattgccat acgaaattcc ggatgagcat tcatcaggcg
     1981 ggcaagaatg tgaataaagg ccggataaaa cttgtgctta tttttcttta cggtctttaa
     2041 aaaggccgta atatccagct gaacggtctg gttataggta cattgagcaa ctgactgaaa
     2101 tgcctcaaaa tgttctttac gatgccattg ggatatatca acggtggtat atccagtgat
     2161 ttttttctcc attttagctt ccttagctcc tgaaaatctc gataactcaa aaaatacgcc
     2221 cggtagtgat cttatttcat tatggtgaaa gttggaacct cttacgtgcc gatcaacgtc
     2281 tcattttcgc cagatatcga cgtctaagaa accatacaaa taggcatgct agcgcagaaa
     2341 cgtcctagaa gatgccagga ggatacttag cagagagaca ataaggccgg agcgaagccg
     2401 tttttccata ggctccgccc ccctgacgaa catcacgaaa tctgacgctc aaatcagtgg
     2461 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctgatgg ctccctcttg
     2521 cgctctcctg ttcccgtcct gcggcgtccg tgttgtggtg gaggctttac ccaaatcacc
     2581 acgtcccgtt ccgtgtagac agttcgctcc aagctgggct gtgtgcaaga accccccgtt
     2641 cagcccgact gctgcgcctt atccggtaac tatcatcttg agtccaaccc ggaaagacac
     2701 gacaaaacgc cactggcagc agccattggt aactgagaat tagtggattt agatatcgag
     2761 agtcttgaag tggtggccta acagaggcta cactgaaagg acagtatttg gtatctgcgc
     2821 tccactaaag ccagttacca ggttaagcag ttccccaact gacttaacct tcgatcaaac
     2881 cgcctcccca ggcggttttt tcgtttacag agcaggagat tacgacgatc gtaaaaggat
     2941 ctcaagaaga tcctttacgg attcccgaca ccatcactct agatttcagt gcaatttatc
     3001 tcttcaaatg tagcacctga agtcagcccc atacgatata agttgtaatt ctcatgttag
     3061 tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag ggcatcggtc
     3121 gagatcccgg tgcctaatga gtgagctaac ttacattaat tgcgttgcgc acaccgacta
     3181 gcgaaaaaac cccgccgaag cggggttttt tgcgaaaaaa agcaccgact cggtgccact
     3241 ttttcaagtt gataacggac tagccttatt ttaacttgct atgctgtttt gaatggttcc
     3301 gctagcactg tacctaggac tgagctagcc gtcaaaagca gcgtatatac catggccaca
     3361 attcagcaaa ttgtgaacat catcacgttc atctttccct ggttgccaat ggcccatttt
     3421 cctgtcagta acgagaaggt cgcgaattca ggcgcttttt agactggtcg tgttttagag
     3481 ctatgctgtt ttgaatggtc ccaaaacatg aactagcgat tagtcgctat gacttaagtt
     3541 ttagagctat gctgttttga atggtcccaa aacaaccaca ctagagaaca tactggctaa
     3601 atagttttag agctatgctg ttttgaatgg tcccaaaacg gttggactca agacgatagt
     3661 taccggatag ttttagagct atgctgtttt gaatggtccc aaaacggcga aacccgacag
     3721 gactataaag ataccgtttt agagctatgc tgttttgaat ggtcccaaaa caaaaaaaaa
     3781 ccccgcccct gacagggcgg ggtttttttt ttaattaaag tctggttata acctgaggtc
     3841 cctatcagtg atagagattg acatccctat cagtgataga gatactgagc acagaaggag
     3901 atatacatgg ataagaaata ctcaataggc ttagatatcg gcacaaatag cgtcggatgg
     3961 gcggtgatca ctgatgaata taaggttccg tctaaaaagt tcaaggttct gggaaataca
     4021 gaccgccaca gtatcaaaaa aaatcttata ggggctcttt tatttgacag tggagagaca
     4081 gcggaagcga ctcgtctcaa acggacagct cgtagaaggt atacacgtcg gaagaatcgt
     4141 atttgttatc tacaggagat tttttcaaat gagatggcga aagtagatga tagtttcttt
     4201 catcgacttg aagagtcttt tttggtggaa gaagacaaga agcatgaacg tcatcctatt
     4261 tttggaaata tagtagatga agttgcttat catgagaaat atccaactat ctatcatctg
     4321 cgaaaaaaat tggtagattc tactgataaa gcggatttgc gcttaatcta tttggcctta
     4381 gcgcatatga ttaagtttcg tggtcatttt ttgattgagg gagatttaaa tcctgataat
     4441 agtgatgtgg acaaactatt tatccagttg gtacaaacct acaatcaatt atttgaagaa
     4501 aaccctatta acgcaagtgg agtagatgct aaagcgattc tttctgcacg attgagtaaa
     4561 tcaagacgat tagaaaatct cattgctcag ctccccggtg agaagaaaaa tggcttattt
     4621 gggaatctca ttgctttgtc attgggtttg acccctaatt ttaaatcaaa ttttgatttg
     4681 gcagaagatg ctaaattaca gctttcaaaa gatacttacg atgatgattt agataattta
     4741 ttggcgcaaa ttggagatca atatgctgat ttgtttttgg cagctaagaa tttatcagat
     4801 gctattttac tttcagatat cctaagagta aatactgaaa taactaaggc tcccctatca
     4861 gcttcaatga ttaaacgcta cgatgaacat catcaagact tgactctttt aaaagcttta
     4921 gttcgacaac aacttccaga aaagtataaa gaaatctttt ttgatcaatc aaaaaacgga
     4981 tatgcaggtt atattgatgg gggagctagc caagaagaat tttataaatt tatcaaacca
     5041 attttagaaa aaatggatgg tactgaggaa ttattggtga aactaaatcg tgaagatttg
     5101 ctgcgcaagc aacggacctt tgacaacggc tctattcccc atcaaattca cttgggtgag
     5161 ctgcatgcta ttttgagaag acaagaagac ttttatccat ttttaaaaga caatcgtgag
     5221 aagattgaaa aaatcttgac ttttcgaatt ccttattatg ttggtccatt ggcgcgtggc
     5281 aatagtcgtt ttgcatggat gactcggaag tctgaagaaa caattacccc atggaatttt
     5341 gaagaagttg tcgataaagg tgcttcagct caatcattta ttgaacgcat gacaaacttt
     5401 gataaaaatc ttccaaatga aaaagtacta ccaaaacata gtttgcttta tgagtatttt
     5461 acggtttata acgaattgac aaaggtcaaa tatgttactg aaggaatgcg aaaaccagca
     5521 tttctttcag gtgaacagaa gaaagccatt gttgatttac tcttcaaaac aaatcgaaaa
     5581 gtaaccgtta agcaattaaa agaagattat ttcaaaaaaa tagaatgttt tgatagtgtt
     5641 gaaatttcag gagttgaaga tagatttaat gcttcattag gtacctacca tgatttgcta
     5701 aaaattatta aagataaaga ttttttggat aatgaagaaa atgaagatat cttagaggat
     5761 attgttttaa cattgacctt atttgaagat agggagatga ttgaggaaag acttaaaaca
     5821 tatgctcacc tctttgatga taaggtgatg aaacagctta aacgtcgccg ttatactggt
     5881 tggggacgtt tgtctcgaaa attgattaat ggtattaggg ataagcaatc tggcaaaaca
     5941 atattagatt ttttgaaatc agatggtttt gccaatcgca attttatgca gctgatccat
     6001 gatgatagtt tgacatttaa agaagacatt caaaaagcac aagtgtctgg acaaggcgat
     6061 agtttacatg aacatattgc aaatttagct ggtagccctg ctattaaaaa aggtatttta
     6121 cagactgtaa aagttgttga tgaattggtc aaagtaatgg ggcggcataa gccagaaaat
     6181 atcgttattg aaatggcacg tgaaaatcag acaactcaaa agggccagaa aaattcgcga
     6241 gagcgtatga aacgaatcga agaaggtatc aaagaattag gaagtcagat tcttaaagag
     6301 catcctgttg aaaatactca attgcaaaat gaaaagctct atctctatta tctccaaaat
     6361 ggaagagaca tgtatgtgga ccaagaatta gatattaatc gtttaagtga ttatgatgtc
     6421 gatcacattg ttccacaaag tttccttaaa gacgattcaa tagacaataa ggtcttaacg
     6481 cgttctgata aaaatcgtgg taaatcggat aacgttccaa gtgaagaagt agtcaaaaag
     6541 atgaaaaact attggagaca acttctaaac gccaagttaa tcactcaacg taagtttgat
     6601 aatttaacga aagctgaacg tggaggtttg agtgaacttg ataaagctgg ttttatcaaa
     6661 cgccaattgg ttgaaactcg ccaaatcact aagcatgtgg cacaaatttt ggatagtcgc
     6721 atgaatacta aatacgatga aaatgataaa cttattcgag aggttaaagt gattacctta
     6781 aaatctaaat tagtttctga cttccgaaaa gatttccaat tctataaagt acgtgagatt
     6841 aacaattacc atcatgccca tgatgcgtat ctaaatgccg tcgttggaac tgctttgatt
     6901 aagaaatatc caaaacttga atcggagttt gtctatggtg attataaagt ttatgatgtt
     6961 cgtaaaatga ttgctaagtc tgagcaagaa ataggcaaag caaccgcaaa atatttcttt
     7021 tactctaata tcatgaactt cttcaaaaca gaaattacac ttgcaaatgg agagattcgc
     7081 aaacgccctc taatcgaaac taatggggaa actggagaaa ttgtctggga taaagggcga
     7141 gattttgcca cagtgcgcaa agtattgtcc atgccccaag tcaatattgt caagaaaaca
     7201 gaagtacaga caggcggatt ctccaaggag tcaattttac caaaaagaaa ttcggacaag
     7261 cttattgctc gtaaaaaaga ctgggatcca aaaaaatatg gtggttttga tagtccaacg
     7321 gtagcttatt cagtcctagt ggttgctaag gtggaaaaag ggaaatcgaa gaagttaaaa
     7381 tccgttaaag agttactagg gatcacaatt atggaaagaa gttcctttga aaaaaatccg
     7441 attgactttt tagaagctaa aggatataag gaagttaaaa aagacttaat cattaaacta
     7501 cctaaatata gtctttttga gttagaaaac ggtcgtaaac ggatgctggc tagtgccgga
     7561 gaattacaaa aaggaaatga gctggc
//
