LOCUS Exported 7263 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Bacillus subtilis sgRNA expression vector; integrates into thrC. ACCESSION . VERSION . KEYWORDS pJMP3 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7263) AUTHORS Peters JM, Colavin A, Shi H, Czarny TL, Larson MH, Wong S, Hawkins JS, Lu CH, Koo BM, Marta E, Shiver AL, Whitehead EH, Weissman JS, Brown ED, Qi LS, Huang KC, Gross CA TITLE A Comprehensive, CRISPR-based Functional Analysis of Essential Genes in Bacteria. JOURNAL Cell. 2016 Jun 2;165(6):1493-506. doi: 10.1016/j.cell.2016.05.003. Epub 2016 May 26. PUBMED 27238023 REFERENCE 2 (bases 1 to 7263) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7263 /organism="synthetic DNA construct" /mol_type="other DNA" misc_RNA 858..933 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" CDS 954..983 /codon_start=1 /product="Myc (human c-Myc proto-oncogene) epitope tag" /label=Myc /translation="EQKLISEEDL" CDS 999..1016 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(1072..1089) /label=pBAD Reverse /note="For vectors with E. coli araBAD promoter, reverse primer" terminator 1242..1288 /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" rep_origin complement(5434..6022) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5514..5533) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6193..7053) /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" primer_bind 6816..6835 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7054..7158) /gene="bla" /label=AmpR promoter primer_bind 7226..7244 /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" ORIGIN 1 aattcatgta aaagatgagg ttggttcatt ctcgaaaatt acatctgtgt tctcagagcg 61 gggcgtgacg tttgaaaaaa tccttcagct gccaattaaa ggccatgatg agttagctga 121 aatcgtaatt gtcacacatc atacatcaga agctgatttc agtgatatcc tgcaaaacct 181 aaatgatttg gaagtcgttc aagaagtcaa aagcacatat cgtgtagaag ggaacggttg 241 gagctaatgt ggaaaggact tatccatcaa tataaagaat ttttacctgt aacagatcaa 301 acaccggcgc taactttaca tgaaggaaac acacctctta ttcacctgcc gaagctgtct 361 gagcagctcg gaattgagct tcatgtcaaa acggaaggcg tcaatcctac gggatcattt 421 aaagatcgcg gaatggttat ggctgtggca aaggcaaaag aagaaggcaa tgacacgatt 481 atgtgcgcgt caacaggtaa cacttccgct gctgcggcag catatgcagc ccgtgctaac 541 atgaaatgca ttgtcatcat cccgaacgga aaaattgcat ttggaaaact cgctcaagct 601 gtcatgtacg gagccgagat tatcgcaatt gacggaaact ttgacgatgc gcttaaaatt 661 gtccgttcca tctgtgagaa atcaccgatt gcccttgtca actcagtcaa cccttaccgc 721 attgaaggcc aaaaaactgc tgccttcgga tcctagaagc ttatcgaatt ccttattaac 781 gttgatataa tttaaatttt atttgacaaa aatgggctcg tgttgtacaa taaatgtaac 841 tttcagttta gcggtctgtt ttagagctag aaatagcaag ttaaaataag gctagtccgt 901 tatcaacttg aaaaagtggc accgagtcgg tgcttttttt gaagcttggg cccgaacaaa 961 aactcatctc agaagaggat ctgaatagcg ccgtcgacca tcatcatcat catcattgag 1021 tttaaacggt ctccagcttg gctgttttgg cggatgagag aagattttca gcctgataca 1081 gattaaatca gaacgcagaa gcggtctgat aaaacagaat ttgcctggcg gcagtagcgc 1141 ggtggtccca cctgacccca tgccgaactc agaagtgaaa cgccgtagcg ccgatggtag 1201 tgtggggtct ccccatgcga gagtagggaa ctgccaggca tcaaataaaa cgaaaggctc 1261 agtcgaaaga ctgggccttt cgttttatct gttgtttgtc ggtgaactgg atccttactc 1321 gagtctagac tgcaggcttc ctcgctcact gactcgaatt cctgcagccc tggcgaatgg 1381 cgattttcgt tcgtgaatac atgttataat aactataact aataacgtaa cgtgactggc 1441 aagagatatt tttaaaacaa tgaataggtt tacacttact ttagttttat ggaaatgaaa 1501 gatcatatca tatataatct agaataaaat taactaaaat aattattatc tagataaaaa 1561 atttagaagc caatgaaatc tataaataaa ctaaattaag tttatttaat taacaactat 1621 ggatataaaa taggtactaa tcaaaatagt gaggaggata tatttgaata catacgaaca 1681 aattaataaa gtgaaaaaaa tacttcggaa acatttaaaa aataacctta ttggtactta 1741 catgtttgga tcaggagttg agagtggact aaaaccaaat agtgatcttg actttttagt 1801 cgtcgtatct gaaccattga cagatcaaag taaagaaata cttatacaaa aaattagacc 1861 tatttcaaaa aaaataggag ataaaagcaa cttacgatat attgaattaa caattattat 1921 tcagcaagaa atggtaccgt ggaatcatcc tcccaaacaa gaatttattt atggagaatg 1981 gttacaagag ctttatgaac aaggatacat tcctcagaag gaattaaatt cagatttaac 2041 cataatgctt taccaagcaa aacgaaaaaa taaaagaata tacggaaatt atgacttaga 2101 ggaattacta cctgatattc cattttctga tgtgagaaga gccattatgg attcgtcaga 2161 ggaattaata gataattatc aggatgatga aaccaactct atattaactt tatgccgtat 2221 gattttaact atggacacgg gtaaaatcat accaaaagat attgcgggaa atgcagtggc 2281 tgaatcttct ccattagaac atagggagag aattttgtta gcagttcgta gttatcttgg 2341 agagaatatt gaatggacta atgaaaatgt aaatttaact ataaactatt taaataacag 2401 attaaaaaaa ttataaaaaa attgaaaaaa tggtggaaac acttttttca atttttttgt 2461 tttattattt aatatttggg aaatattcat tctaattggt aatcagattt tagaaaacaa 2521 taaacccttg cataggggga tctcgaggcc ttccgaaaat gcgcggcttt gaagctgagg 2581 gtgcggcggc aatcgtgcgc aatgaagtga ttgaaaatcc ggaaacaata gcgacagcca 2641 ttcgtatcgg aaacccggca agctgggaca aagctgtaaa ggcagccgag gaatccaatg 2701 ggaaaattga cgaagtcact gatgatgaaa tccttcacgc atatcagctg atcgcccgtg 2761 tagaaggcgt gtttgcagaa ccaggttctt gcgcgtctat cgcaggagtg ctgaaacagg 2821 tgaaatccgg agaaattccg aaaggcagca aggtcgtagc tgtgttaaca ggaaacggac 2881 tgaaagatcc gaacacagcg gtcgacattt cagaaatcaa gcctgtcaca ttgccgactg 2941 atgaagacag catccttgaa tatgtaaaag gagcggcccg tgtatgaacg aagccgacat 3001 gctgttctct gtcactgttc ccggaagcac agctaaccta ggccccggct ttgattcagt 3061 cggaatggcg ctcagcagat atttgaagct gaccgtcttt gaaagcgaca aatggtcttt 3121 tgaggctgaa acagaaacag tcgccggaat taattcgagc tcctttaact tcctcaccag 3181 tagttgtatc ggtaccataa gtagaagcag caacccaagt agctttacca gcatccggtt 3241 caaccagcat agtaagaatc ttactggaca tcggcagttc ttcgaacagt gcgccaacta 3301 ccagctcttt ctgcagttca ttcagggcac cggagaacct gcgtgcaatc catcttgttc 3361 aatcatgcga aacgatcctc atcctgtctc ttgatccatg gattacgcgt taacccgggc 3421 ccgcggatgc atatgatcag atcctttaac tctggcaacc ctcaaaattg aatgagacat 3481 gctacacctc cggataataa atatatataa acgtatatag atttcataaa gtctaacaca 3541 ctagacttat ttacttcgta attaagtcgt taaaccgtgt gctctacgac caaaactata 3601 aaacctttaa gaactttctt tttttacaag aaaaaagaaa ttagataaat ctctcatatc 3661 ttttattcaa taatcgcatc cgattgcagt ataaatttaa cgatcactca tcatgttcat 3721 atttatcaga gctcgtgcta taattatact aattttataa ggaggaaaaa atatgggcat 3781 ttttagtatt tttgtaatca gcacagttca ttatcaacca aacaaaaaat aagtggttat 3841 aatgaatcgt taataagcaa aattcatata accaaattaa agagggttat aatgaacgag 3901 aaaaatataa aacacagtca aaactttatt acttcaaaac ataatataga taaaataatg 3961 acaaatataa gattaaatga acatgataat atctttgaaa tcggctcagg aaaaggccat 4021 tttacccttg aattagtaaa gaggtgtaat ttcgtaactg ccattgaaat agaccataaa 4081 ttatgcaaaa ctacagaaaa taaacttgtt gatcacgata atttccaagt tttaaacaag 4141 gatatattgc agtttaaatt tcctaaaaac caatcctata aaatatatgg taatatacct 4201 tataacataa gtacggatat aatacgcaaa attgtttttg atagtatagc taatgagatt 4261 tatttaatcg tggaatacgg gtttgctaaa agattattaa atacaaaacg ctcattggca 4321 ttacttttaa tggcagaagt tgatatttct atattaagta tggttccaag agaatatttt 4381 catcctaaac ctaaagtgaa tagctcactt atcagattaa gtagaaaaaa atcaagaata 4441 tcacacaaag ataaacaaaa gtataattat ttcgttatga aatgggttaa caaagaatac 4501 aagaaaatat ttacaaaaaa tcaatttaac aattccttaa aacatgcagg aattgacgat 4561 ttaaacaata ttagctttga acaattctta tctcttttca atagctataa attatttaat 4621 aagtaagtta agggatgcat aaactgcatc ccttaacttg tttttcgtgt gcctattttt 4681 tgtgaatcga ttatgtcttt tgcgcagtcg gcttaaacca gttttcgctg gtgcgaaaaa 4741 agagtgtctt gtgacaccta aattcaaaat ctatcggtca gatttatacc gatttgattt 4801 tatatattct tgaataacat acgccgagtt atcacataaa agcgggaacc aatcatcaaa 4861 tttaaacttc attgcataat ccattaaact cttaaattct acgattcctt gttcatcaat 4921 aaactcaatc atttctttaa ttaatttata tctatctgtt gttgttttct ttaataattc 4981 atcaacatct acaccgccat aaactatcat atcttctttt tgatatttaa atttattagg 5041 atcttaaggc ctaggtctag agtctttgtt ttgacgccat tagcgtacgt aacaatcctc 5101 gttaaaggac aaggacctga gcggaagtgt atcgtacagt agacggagta tactagtata 5161 gtctatagtc cgtggaatta ttatatttat ctccgacgat attctcatca gtgaaatcca 5221 gctggagttc tttagcaaat ttttttatta gctgaactta gtattagtgg ccatactcct 5281 ccaatccaaa gctatttaga aagattacta tatcctcaaa caggcggtaa ccggcctctt 5341 catcgggaat gcgcgcgacc ttcagcatcg ccggcatgtc cccctggcgg acgggaagta 5401 tccagctcga ggtcgggccg cgttgctggc gtttttccat aggctccgcc cccctgacga 5461 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata 5521 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac 5581 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg 5641 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc 5701 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag 5761 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt 5821 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt 5881 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg atagctcttg 5941 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac 6001 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca 6061 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac 6121 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac 6181 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt 6241 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt 6301 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt 6361 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc 6421 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa 6481 tagtttgcgc aacgttgttg ccattgctgc aggcatcgtg gtgtcacgct cgtcgtttgg 6541 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt 6601 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc 6661 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt 6721 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg 6781 gcgaccgagt tgctcttgcc cggcgtcaac acgggataat accgcgccac atagcagaac 6841 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc 6901 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt 6961 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg 7021 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag 7081 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa 7141 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat 7201 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtcttcaaga 7261 att //