LOCUS pgRNA 4217 bp DNA circular UNA 08-JAN-2020 DEFINITION J23119-lacZ gRNA, sacB, pBR322 ori, Cm (aka pZZ45). ACCESSION urn.local...t-9du8wb2 VERSION urn.local...t-9du8wb2 KEYWORDS . SOURCE ORGANISM . FEATURES Location/Qualifiers modified_base 1..357 primer_bind 1..25 /standard_name="ZZ50" primer_bind 290..314 /standard_name="ZZ31" promoter 358..392 misc_feature 393..412 /standard_name="N20 Custom crRNA Sequence" misc_RNA 413..494 /standard_name="sgRNA" primer_bind 470..514 /standard_name="ZZ52" misc_feature complement(578..602) /note="Geneious type: primer_bind_reverse" /standard_name="ZZ58" misc_feature 703..1341 /note="Geneious type: marker" /note="Tn9" /product="ampicillin acetyl transferase" /db_xref="GI:15554330" /standard_name="cat" primer_bind 1065..1089 /standard_name="ZZ72" misc_feature complement(1149..1173) /note="Geneious type: primer_bind_reverse" /standard_name="ZZ71" primer_bind 1219..1243 /standard_name="ZZ64" misc_feature 1327..1366 /note="Geneious type: primer_bind_reverse" /standard_name="ZZ363" CDS complement(1415..2836) /codon_start=1 /gene="Bacillus subtilis sacB" /product="secreted levansucrase that renders bacterial growth sensitive to sucrose" /note="negative selection marker" /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGIS HITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHG YHIVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQT QEWSGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIF DGDGKTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEES LFNKAYYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIAS NTVTDEIERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLN KTGLVLKMDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLN IKGKKTSVVKDSILEQGQLTVNK" /standard_name="sacB" primer_bind complement(3258..3297) /standard_name="ZZ362" misc_feature 3268..3307 /note="Geneious type: primer_bind_reverse" /standard_name="ZZ365" misc_feature complement(3368..3397) /note="Geneious type: primer_bind_reverse" /standard_name="ZZ361" rep_origin 3437..4056 primer_bind 3527..3551 /standard_name="ZZ35" misc_feature complement(3626..3650) /note="Geneious type: primer_bind_reverse" /standard_name="ZZ34" ORIGIN 1 accatggctg gtgaccacgt cgtggaatgc cttcgaattc agcacctgca catgggacgt 61 cgacctgagg taattataac ccgggcccta tatatggatc caattgcaat gatcatcatg 121 acagatctgc gcgcgatcga tatcagcgct ttaaatttgc gcatgctagc tatagttcta 181 gaggtaccgg ttgttaacgt tagccggcta cgtatactcc ggaatattaa taggcctagg 241 atgcatatgg cggccgcctg cagctggcgc catcgatacg cgtacgtcgc gaccgcggac 301 atgtacagag ctcgagaagt actagtggcc acgtgggccg tgcaccttaa gcttggcttg 361 acagctagct cagtcctagg tataatgcta gcnnnnnnnn nnnnnnnnnn nngttttaga 421 gctagaaata gcaagttaaa ataaggctag tccgttatca acttgaaaaa gtggcaccga 481 gtcggtgctt ttttggcgcg cctacctgtg acggaagatc acttcgcaga ataaataaat 541 cctggtgtcc ctgttgatac cgggaagccc tgggccaact tttggcgaaa atgagacgtt 601 gatcggcacg taagaggttc caactttcac cataatgaaa taagatcact accgggcgta 661 ttttttgagt tgtcgagatt ttcaggagct aaggaagcta aaatggagaa aaaaatcact 721 ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga ggcatttcag 781 tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc ctttttaaag 841 accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct tgcccgcctg 901 atgaatgctc atccggaatt acgtatggca atgaaatacg gtgagctggt gatatgggat 961 agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc atcgctctgg 1021 agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga tgtggcgtgt 1081 tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt tttcgtctca 1141 gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat ggacaacttc 1201 ttcgcccccg ttttcaccat gggcaaatat tatacgcaag gcgacaaggt gctgatgccg 1261 ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag atgcttaatg 1321 aatacaacag tactgcgatg aatgggattc acctttatgt tgataagaaa taaaagaaaa 1381 tgccaatagg atatcggcat tttcttttgc gtttttattt gttaactgtt aattgtcctt 1441 gttcaaggat gctgtctttg acaacagatg ttttcttgcc tttgatgttc agcaggaagc 1501 ttggcgcaaa cgttgattgt ttgtctgcgt agaatcctct gtttgtcata tagcttgtaa 1561 tcacgacatt gtttcctttc gcttgaggta cagcgaagtg tgagtaagta aaggttacat 1621 cgttaggatc aagatccatt tttaacacaa ggccagtttt gttcagcggc ttgtatgggc 1681 cagttaaaga attagaaaca taaccaagca tgtaaatatc gttagacgta atgccgtcaa 1741 tcgtcatttt tgatccgcgg gagtcagtga acaggtacca tttgccgttc attttaaaga 1801 cgttcgcgcg ttcaatttca tctgttactg tgttagatgc aatcagcggt ttcatcactt 1861 ttttcagtgt gtaatcatcg tttagctcaa tcataccgag agcgccgttt gctaactcag 1921 ccgtgcgttt tttatcgctt tgcagaagtt tttgactttc ttgacggaag aatgatgtgc 1981 ttttgccata gtatgctttg ttaaataaag attcttcgcc ttggtagcca tcttcagttc 2041 cagtgtttgc ttcaaatact aagtatttgt ggcctttatc ttctacgtag tgaggatctc 2101 tcagcgtatg gttgtcgcct gagctgtagt tgccttcatc gatgaactgc tgtacatttt 2161 gatacgtttt tccgtcaccg tcaaagattg atttataatc ctctacaccg ttgatgttca 2221 aagagctgtc tgatgctgat acgttaactt gtgcagttgt cagtgtttgt ttgccgtaat 2281 gtttaccgga gaaatcagtg tagaataaac ggatttttcc gtcagatgta aatgtggctg 2341 aacctgacca ttcttgtgtt tggtctttta ggatagaatc atttgcatcg aatttgtcgc 2401 tgtctttaaa gacgcggcca gcgtttttcc agctgtcaat agaagtttcg ccgacttttt 2461 gatagaacat gtaaatcgat gtgtcatccg catttttagg atctccggct aatgcaaaga 2521 cgatgtggta gccgtgatag tttgcgacag tgccgtcagc gttttgtaat ggccagctgt 2581 cccaaacgtc caggcctttt gcagaagaga tatttttaat tgtggacgaa tcgaattcag 2641 gaacttgata tttttcattt ttttgctgtt cagggatttg cagcatatca tggcgtgtaa 2701 tatgggaaat gccgtatgtt tccttatatg gcttttggtt cgtttctttc gcaaacgctt 2761 gagttgcgcc tcctgccagc agtgcggtag taaaggttaa tactgttgct tgttttgcaa 2821 actttttgat gttcatcgtt catgtctcct tttttatgta ctgtgttagc ggtctgcttc 2881 ttccagccct cctgtttgaa gatggcaagt tagttacgca caataaaaaa agacctaaaa 2941 tatgtaaggg gtgacgccaa agtatacact ttgcccttta cacattttag gtcttgcctg 3001 ctttatcagt aacaaacccg cgcgatttac ttttcgacct cattctatta gactctcgtt 3061 tggattgcaa ctggtctatt ttcctctttt gtttgataga aaatcataaa aggatttgca 3121 gactacgggc ctaaagaact aaaaaatcta tctgtttctt ttcattctct gtatttttta 3181 tagtttctgt tgcatgggca taaagttgcc tttttaatca caattcagaa aatatcataa 3241 tatctcattt cactaaataa tagtgaacgg caggtatatg tgctgtcaga ccaagtttac 3301 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag 3361 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg 3421 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc 3481 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag 3541 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc 3601 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac 3661 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc 3721 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt 3781 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt 3841 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc 3901 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt 3961 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca 4021 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt 4081 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt 4141 attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag 4201 tcagtgagcg aggaagc //