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 //