LOCUS Exported 8631 bp ds-DNA circular SYN 10-AUG-2021 DEFINITION Optimized M. tuberculosis CRISPRi plasmid; L5 integrating.. ACCESSION . VERSION . KEYWORDS plRL2 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8631) AUTHORS Bosch B, DeJesus MA, Poulton NC, Zhang W, Engelhart CA, Zaveri A, Lavalette S, Ruecker N, Trujillo C, Wallach JB, Li S, Ehrt S, Chait BT, Schnappinger D, Rock JM TITLE Genome-wide gene expression tuning reveals diverse vulnerabilities of M. tuberculosis. JOURNAL Cell. 2021 Jul 16. pii: S0092-8674(21)00824-2. doi: 10.1016/j.cell.2021.06.033. PUBMED 34297925 REFERENCE 2 (bases 1 to 8631) AUTHORS . TITLE Direct Submission JOURNAL Exported Aug 10, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8631 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin complement(496..1084) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(576..595) /label=pBR322ori-F /note="pBR322 origin, forward primer" protein_bind 1267..1285 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 1297..1315 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" terminator 4795..4822 /label=rrnB T2 terminator /note="transcription terminator T2 from the E. coli rrnB gene" primer_bind complement(7588..7607) /label=pENTR-R /note="pENTR vectors, reverse primer" CDS 7599..8414 /codon_start=1 /gene="aph(3')-Ia" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin in bacteria or G418 (Geneticin(R)) in eukaryotes" /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG KPHAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA SDFDDERNGWPVEQVWKEMHNLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind complement(7672..7691) /label=Kan-R /note="Kanamycin resistance gene, reverse primer" ORIGIN 1 aataaacgaa aggctcagtc gaaagactgg gcctttcgtt tatctgttgt ttgaaaaaaa 61 aaagcgccgc aactgcggcg cttttttttt tgaattctct gaccagggaa aatagccctc 121 tgacctgggg atttgcgatc cctatcagtg atagatataa tctgggagga gacgattaat 181 gcgtctcggt ttttgtactc gaaagaagct acaaagataa ggcttcatgc cgaaatcaac 241 accctgtcat tttatggcag ggtgtttttt ttttgtcgac ttggggaccc tagaggtccc 301 cttttttatt tttttggatg aagagcatta tcaatggctc ttcacaggaa gcttaaaaaa 361 aaaagccccg cgattgcggg gctttttttt tttaatacgg ttatccacag aatcagggga 421 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc 481 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg 541 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg 601 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt 661 tctcccctcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt 721 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg 781 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact 841 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt 901 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct 961 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac 1021 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc 1081 tcaagaagat cctttgatct tttctacggg gtctgacgct catggaacaa taaacgaaag 1141 gctcagtcga aagactgggc ctttcgttta tctgttgttt gaaaaaaaaa agccccgcga 1201 ttgcggggct tttttttttc atatgtctga ccagggaaaa tagccctctg acctggggat 1261 ttgacttccc tatcagtgat agagataatc tgggagtccc tatcagtgat agagaacgcg 1321 gtatcgatat gtcggacctg gtgctgggcc tggccatcgg catcggctcg gtgggcgtgg 1381 gcatcctgaa caaggtgacc ggcgagatca tccacaagaa ctcgcgcatc ttcccggccg 1441 cccaggccga gaacaacctg gtgcgccgca ccaaccgcca gggccgccgc ctggcccgcc 1501 gcaagaagca ccgccgcgtg cgcctgaacc gcctgttcga ggagtcgggc ctgatcaccg 1561 acttcaccaa gatctcgatc aacctgaacc cgtaccagct gcgcgtgaag ggcctgaccg 1621 acgagctgtc gaacgaggag ctgttcatcg ccctgaagaa catggtgaag caccgcggca 1681 tctcgtacct ggacgacgcc tcggacgacg gcaactcgtc ggtgggcgac tacgcccaga 1741 tcgtgaagga gaactcgaag cagctggaga ccaagacccc gggccagatc cagctggagc 1801 gctaccagac ctacggccag ctgcgcggcg acttcaccgt ggagaaggac ggcaagaagc 1861 accgcctgat caacgtgttc ccgacctcgg cctaccgctc ggaggccctg cgcatcctgc 1921 agacccagca ggagttcaac ccgcagatca ccgacgagtt catcaaccgc tacctggaga 1981 tcctgaccgg caagcgcaag tactaccacg gcccgggcaa cgagaagtcg cgcaccgact 2041 acggccgcta ccgcacctcg ggcgagaccc tggacaacat cttcggcatc ctgatcggca 2101 agtgcacctt ctacccggac gagttccgcg ccgccaaggc ctcgtacacc gcccaggagt 2161 tcaacctgct gaacgacctg aacaacctga ccgtgccgac cgagaccaag aagctgtcga 2221 aggagcagaa gaaccagatc atcaactacg tgaagaacga gaaggccatg ggcccggcca 2281 agctgttcaa gtacatcgcc aagctgctgt cgtgcgacgt ggccgacatc aagggctacc 2341 gcatcgacaa gtcgggcaag gccgagatcc acaccttcga ggcctaccgc aagatgaaga 2401 ccctggagac cctggacatc gagcagatgg accgcgagac cctggacaag ctggcctacg 2461 tgctgaccct gaacaccgag cgcgagggca tccaggaggc cctggagcac gagttcgccg 2521 acggctcgtt ctcgcagaag caggtggacg agctggtgca gttccgcaag gccaactcgt 2581 cgatcttcgg caagggctgg cacaacttct cggtgaagct gatgatggag ctgatcccgg 2641 agctgtacga gacctcggag gagcagatga ccatcctgac ccgcctgggc aagcagaaga 2701 ccacctcgtc gtcgaacaag accaagtaca tcgacgagaa gctgctgacc gaggagatct 2761 acaacccggt ggtggccaag tcggtgcgcc aggccatcaa gatcgtgaac gccgccatca 2821 aggagtacgg cgacttcgac aacatcgtga tcgagatggc ccgcgagacc aacgaggacg 2881 acgagaagaa ggccatccag aagatccaga aggccaacaa ggacgagaag gacgccgcca 2941 tgctgaaggc cgccaaccag tacaacggca aggccgagct gccgcactcg gtgttccacg 3001 gccacaagca gctggccacc aagatccgcc tgtggcacca gcagggcgag cgctgcctgt 3061 acaccggcaa gaccatctcg atccacgacc tgatcaacaa ctcgaaccag ttcgaggtgg 3121 acgccatcct gccgctgtcg atcaccttcg acgactcgct ggccaacaag gtgctggtgt 3181 acgccaccgc caaccaggag aagggccagc gcaccccgta ccaggccctg gactcgatgg 3241 acgacgcctg gtcgttccgc gagctgaagg ccttcgtgcg cgagtcgaag accctgtcga 3301 acaagaagaa ggagtacctg ctgaccgagg aggacatctc gaagttcgac gtgcgcaaga 3361 agttcatcga gcgcaacctg gtggacaccc gctacgcctc gcgcgtggtg ctgaacgccc 3421 tgcaggagca cttccgcgcc cacaagatcg acaccaaggt gtcggtggtg cgcggccagt 3481 tcacctcgca gctgcgccgc cactggggca tcgagaagac ccgcgacacc taccaccacc 3541 acgccgtgga cgccctgatc atcgccgcct cgtcgcagct gaacctgtgg aagaagcaga 3601 agaacaccct ggtgtcgtac tcggaggacc agctgctgga catcgagacc ggcgagctga 3661 tctcggacga cgagtacaag gagtcggtgt tcaaggcccc gtaccagcac ttcgtggaca 3721 ccctgaagtc gaaggagttc gaggactcga tcctgttctc gtaccaggtg gactcgaagt 3781 tcaaccgcaa gatctcggac gccaccatct acgccacccg ccaggccaag gtgggcaagg 3841 acaaggccga cgagacctac gtgctgggca agatcaagga catctacacc caggacggct 3901 acgacgcctt catgaagatc tacaagaagg acaagtcgaa gttcctgatg taccgccacg 3961 acccgcagac cttcgagaag gtgatcgagc cgatcctgga gaactacccg aacaagcaga 4021 tcaacgacaa gggcaaggag gtgccgtgca acccgttcct gaagtacaag gaggagcacg 4081 gctacatccg caagtactcg aagaagggca acggcccgga gatcaagtcg ctgaagtact 4141 acgactcgaa gctgggcaac cacatcgaca tcaccccgaa ggactcgaac aacaaggtgg 4201 tgctgcagtc ggtgtcgccg tggcgcgccg acgtgtactt caacaagacc accggcaagt 4261 acgagatcct gggcctgaag tacgccgacc tgcagttcga caagggcacc ggcacctaca 4321 agatctcgca ggagaagtac aacgacatca agaagaagga gggcgtggac tcggactcgg 4381 agttcaagtt caccctgtac aagaacgacc tgctgctggt gaaggacacc gagaccaagg 4441 agcagcagct gttccgcttc ctgtcgcgca ccatgccgaa gcagaagcac tacgtggagc 4501 tgaagccgta cgacaagcag aagttcgagg gcggcgaggc cctgatcaag gtgctgggca 4561 acgtggccaa ctcgggccag tgcaagaagg gcctgggcaa gtcgaacatc tcgatctaca 4621 aggtgcgcac cgacgtgctg ggcaaccagc acatcatcaa gaacgagggc gacaagccga 4681 agctggactt ctaaatactc tggccatttt tctttaccgc ctttgagtga gctgataccg 4741 ctcgccgcag ccgaacgacc gagcgcaacg cgtgcggccg ctgattagct aagcagaagg 4801 ccatcctgac ggatggcctt tttctagaaa aaaaaaaagc gccgcaactg cggcgctttt 4861 ttttttttaa ttaagtgata gattgcgatg cgtgtgctcg gttgatataa tctgtcagat 4921 caggaggact tcgcatgtcg cggttggata agtccaaggt catcaactcg gcgctggagc 4981 tgctgaacga ggtcggtatc gagggactga ccacgcgcaa gttggcccaa aagctcggcg 5041 tggaacaacc gaccctctac tggcacgtga agaacaagcg cgcgctgctg gatgcgttgg 5101 ccatcgagat gctggatcgc catcacacgc acttttgccc cctggagggc gagagctggc 5161 aggattttct gcggaacaac gcgaagtcgt tccggtgcgc cctcctgagc catcgggatg 5221 gcgccaaagt ccacttgggc acccgcccga ccgagaagca atacgagacc ctggagaacc 5281 agctggcctt cctctgtcaa cagggttttt cgttggagaa cgccctgtat gccctgtccg 5341 ccgtgggaca cttcaccttg ggctgcgtgt tggaggatca ggaacatcag gtggcgaaag 5401 aggaacggga gacccccacg acggattcga tgcccccgtt gttgcgccag gcgatcgaac 5461 tgtttgacca ccagggcgcc gaacccgcct ttttgttcgg gctggagctg atcatctgcg 5521 ggctggaaaa gcaactgaag tgcgaaagcg gttcgtgata acgccgaaaa ccccgcttcg 5581 gcggggtttt gccgcggtct gcgttgtcgg gaagatttaa aataaaaaag gggacctcta 5641 gggtccccaa ttaaatagtt aacactagta ttaaaaaaaa aagccccgcg attgcggggc 5701 tttttttttt cgacggcgag ggttccgacc gctgcaactc ccggtgcaac cttgtcccgg 5761 tctattctct tcactgcacc agctccaatc tggtgtgaat gcccctcgtc tgttcgcgca 5821 ggcggggggc tctattcgtt tgtcagcatc gaaagtagcc agatcaggga tgcgttgcaa 5881 ccgcgtatgc ccaggtcaga agagtcgcac aagagttgca gacccctgga aagaaaaatg 5941 gccagagggc gaaaacaccc tctgaccagc ggagcgggcg acgggaatcg aacccgcgta 6001 gctagtttgg aagaatgggt gtctgccgac cacatatatg ggccggtcaa gataggtttt 6061 taccccctct cggctgcatc ctctaagtgg aaagaaattg caggtcgtag aagcgcgttg 6121 aagcctgaga gttgcacagg agttgcaacc cggtagcctt gttcacgacg agaggagacc 6181 tagttggcac gtcgcggatg gggatcgctg aagactcagc gcagcgggag gatccaagcc 6241 tcatacgtca acccgcagga cggtgtgagg tactacgcgc tgcagaccta cgacaacaag 6301 atggacgccg aagcctggct cgcgggcgag aagcggctca tcgagatgga gacctggacc 6361 cctccacagg accgggcgaa gaaggcagcc gccagcgcca tcacgctgga ggagtacacc 6421 cggaagtggc tcgtggagcg cgacctcgca gacggcacca gggatctgta cagcgggcac 6481 gcggagcgcc gcatctaccc ggtgctaggt gaagtggcgg tcacagagat gacgccagct 6541 ctggtgcgtg cgtggtgggc cgggatgggt aggaagcacc cgactgcccg ccggcatgcc 6601 tacaacgtcc tccgggcggt gatgaacaca gcggtcgagg acaagctgat cgcagagaac 6661 ccgtgccgga tcgagcagaa ggcagccgat gagcgcgacg tagaggcgct gacgcctgag 6721 gagctggaca tcgtcgccgc tgagatcttc gtgcactacc ggatcgcggc atacatcctg 6781 gcgtggacga gcctccggtt cggagagctg atcgagcttc gccgcaagga catcgtggac 6841 gacggcatga cgatgaagct ccgggtgcgc cgtggcgctt cccgcgtggg gaacaagatc 6901 gtcgttggca acgccaagac cgtccggtcg aagcgtcctg tgacggttcc gcctcacgtc 6961 gcggagatga tccgagcgca catgaaggac cgtacgaaga tgaacaaggg ccccgaggca 7021 ttcctggtga ccacgacgca gggcaaccgg ctgtcgaagt ccgcgttcac caagtcgctg 7081 aagcgtggct acgccaagat cggtcggccg gaactccgca tccacgacct ccgcgctgtc 7141 ggcgctacgt tcgccgctca ggcaggtgcg acgaccaagg agctgatggc ccgcctcggt 7201 cacacgactc ctaggatggc gatgaagtac cagatggcgt ctgaggcccg cgacgaggct 7261 atcgctgagg cgatgtccaa gctggccaag acctcctgaa acgcaaaaag cccccctccc 7321 aaggacactg agtcctaaag aggggggttt cttgtcagta cgcgaagaac cacgcctggc 7381 cgcgagcgcc agcaccgccg ctctgtgcgg agacctgggc accagccccg ccgccgccag 7441 gagcattgcc gttcccgcca gctagccacc tgacgtcggg ggggggggaa agccacgttg 7501 tgtctcaaaa tctctgatgt tacattgcac aagataaaaa tatatcatca tgaacaataa 7561 aactgtctgc ttacataaac agtaatacaa ggggtgttat gagccatatt caacgggaaa 7621 cgtcttgctc gaggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat 7681 gggctcgcga taatgtcggg caatcaggtg cgacaatcta tcgcttgtat gggaagcccc 7741 atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg 7801 agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta 7861 tccgtactcc tgatgatgca tggttactca ccactgcgat ccccgggaaa acagcattcc 7921 aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc 7981 tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat cgcgtatttc 8041 gcctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt gattttgatg 8101 acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataat cttttgccat 8161 tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg 8221 aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg 8281 atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt 8341 ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg 8401 atgagttttt ctaatcagaa ttggttaatt ggttgtaaca ctggcagagc attacgctga 8461 cttgacggga cggcggcttt gttgaataaa tcgaactttt gctgagttga aggatcagat 8521 cacgcatctt cccgacaacg cagaccgttc cgtggcaaag caaaagttca aaatcaccaa 8581 ctggtccacc tacaacaaag ctctcaccaa ccgtggctcc ctcacgatat c //