LOCUS Exported 6798 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION sacB plasmid with polylinker; streptomycin/spectinomycin resistance: used as a conditionally lethal gene in Anabaena sp. strain PCC 7120 to select for double recombinants and to entrap insertion seq.. ACCESSION . VERSION . KEYWORDS pRL277 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6798) AUTHORS Cai YP, Wolk CP TITLE Use of a conditionally lethal gene in Anabaena sp. strain PCC 7120 to select for double recombinants and to entrap insertion sequences. JOURNAL J Bacteriol. 1990 Jun;172(6):3138-45. PUBMED 2160938 REFERENCE 2 (bases 1 to 6798) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6798 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(19..38) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 179..201 /label=pGEX 3' /note="pGEX vectors, reverse primer" CDS complement(707..2128) /codon_start=1 /gene="Bacillus subtilis sacB" /product="secreted levansucrase that renders bacterial growth sensitive to sucrose" /label=SacB /note="negative selection marker" /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLEVWDSWPLQNADGTVANYHGYH IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV VKDSILEQGQLTVNK" promoter complement(2129..2574) /gene="sacR" /label=sacB promoter /note="sacB promoter and control region" CDS complement(3214..4005) /codon_start=1 /gene="aadA" /product="aminoglycoside adenylyltransferase (Murphy, 1985)" /label=SmR /note="confers resistance to spectinomycin and streptomycin" /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK" rep_origin 5977..6565 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6466..6485 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 6719..6736 /label=L4440 /note="L4440 vector, forward primer" misc_feature 6751..93 /label=bom /note="basis of mobility region from pBR322" ORIGIN 1 catatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agtatacact 61 ccgctatcgc tacgtgactg ggtcatggct gcgccccgac acccgccaac acccgctgac 121 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc 181 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag gcaggctggc 241 tgttttacgc gtatgacagg ctccggaaga cggttgttgc gcacgtattc ggtgaacgca 301 ctatggcgac gctggggcgt cttatgagcc tgctgtcacc ctttgacgtg gtgatatgga 361 tgacggatgg ctggccgctg tatgaatccc gcctgaaggg aaagctgcac gtaatcagca 421 agcgatatac gcagcgaatt gagcggcata acctgaatct gaggcagcac ctggcacggc 481 tgggacggaa gtcgctgtcg ttctcaaaat cggtggagct gcatgacaaa gtcatcgggc 541 attatctgaa cataaaacac tatcaataag ttggagtcat taccaaaagg ttaggaatac 601 ggttagccat ttgcctgctt ttatatagtt catatgggat tcacctttat gttgataaga 661 aataaaagaa aatgccaata ggatatcggc attttctttt gcgtttttat ttgttaactg 721 ttaattgtcc ttgttcaagg atgctgtctt tgacaacaga tgttttcttg cctttgatgt 781 tcagcaggaa gcttggcgca aacgttgatt gtttgtctgc gtagaatcct ctgtttgtca 841 tatagcttgt aatcacgaca ttgtttcctt tcgcttgagg tacagcgaag tgtgagtaag 901 taaaggttac atcgttagga tcaagatcca tttttaacac aaggccagtt ttgttcagcg 961 gcttgtatgg gccagttaaa gaattagaaa cataaccaag catgtaaata tcgttagacg 1021 taatgccgtc aatcgtcatt tttgatccgc gggagtcagt gaacaggtac catttgccgt 1081 tcattttaaa gacgttcgcg cgttcaattt catctgttac tgtgttagat gcaatcagcg 1141 gtttcatcac ttttttcagt gtgtaatcat cgtttagctc aatcataccg agagcgccgt 1201 ttgctaactc agccgtgcgt tttttatcgc tttgcagaag tttttgactt tcttgacgga 1261 agaatgatgt gcttttgcca tagtatgctt tgttaaataa agattcttcg ccttggtagc 1321 catcttcagt tccagtgttt gcttcaaata ctaagtattt gtggccttta tcttctacgt 1381 agtgaggatc tctcagcgta tggttgtcgc ctgagctgta gttgccttca tcgatgaact 1441 gctgtacatt ttgatacgtt tttccgtcac cgtcaaagat tgatttataa tcctctacac 1501 cgttgatgtt caaagagctg tctgatgctg atacgttaac ttgtgcagtt gtcagtgttt 1561 gtttgccgta atgtttaccg gagaaatcag tgtagaataa acggattttt ccgtcagatg 1621 taaatgtggc tgaacctgac cattcttgtg tttggtcttt taggatagaa tcatttgcat 1681 cgaatttgtc gctgtcttta aagacgcggc cagcgttttt ccagctgtca atagaagttt 1741 cgccgacttt ttgatagaac atgtaaatcg atgtgtcatc cgcattttta ggatctccgg 1801 ctaatgcaaa gacgatgtgg tagccgtgat agtttgcgac agtgccgtca gcgttttgta 1861 atggccagct gtcccaaacc tccaggcctt ttgcagaaga gatattttta attgtggacg 1921 aatcgaattc aggaacttga tatttttcat ttttttgctg ttcagggatt tgcagcatat 1981 catggcgtgt aatatgggaa atgccgtatg tttccttata tggcttttgg ttcgtttctt 2041 tcgcaaacgc ttgagttgcg cctcctgcca gcagtgcggt agtaaaggtt aatactgttg 2101 cttgttttgc aaactttttg atgttcatcg ttcatgtctc cttttttatg tactgtgtta 2161 gcggtctgct tcttccagcc ctcctgtttg aagatggcaa gttagttacg cacaataaaa 2221 aaagacctaa aatatgtaag gggtgacgcc aaagtataca ctttgccctt tacacatttt 2281 aggtcttgcc tgctttatca gtaacaaacc cgcgcgattt acttttcgac ctcattctat 2341 tagactctcg tttggattgc aactggtcta ttttcctctt ttgtttgata gaaaatcata 2401 aaaggatttg cagactacgg gcctaaagaa ctaaaaaatc tatctgtttc ttttcattct 2461 ctgtattttt tatagtttct gttgcatggg cataaagttg cctttttaat cacaattcag 2521 aaaatatcat aatatctcat ttcactaaat aatagtgaac ggcaggtata tgtgatgggt 2581 taaaaaggat cgatcctcta gctagagtcg acctgcatcc cttaacttac ttattaaata 2641 atttatagct attgaaaaga gataagaatt gttcaaagct aatattgttt aaatcgtcaa 2701 ttcctgcatg ttttaaggaa ttgttaaatt gattttttgt aaatattttc ttgtattctt 2761 tgttaaccca tttcataacg aaataattat acttttgttt atctttgtgt gatattcttg 2821 atttttttct acttaatctg ataagtgagc tattcacttt aggtttagga tgaaaaaaaa 2881 taaaaaaggg gacctctagg gtccccaatt aattagtaat ataatctatt aaaggtcatt 2941 caaaaggtca tccaccggat cagcttagta aagccctcgc tagattttaa tgcggatgtt 3001 gcgattactt cgccaactat tgcgataaca agaaaaagcc agcctttcat gatatatctc 3061 ccaatttgtg tagggcttat tatgcacgct taaaaataat aaaagcagac ttgacctgat 3121 agtttggctg tgagcaatta tgtgcttagt gcatctaacg cttgagttaa gccgcgccgc 3181 gaagcggcgt cggcttgaac gaattgttag acattatttg ccgactacct tggtgatctc 3241 gcctttcacg tagtggacaa attcttccaa ctgatctgcg cgcgaggcca agcgatcttc 3301 ttcttgtcca agataagcct gtctagcttc aagtatgacg ggctgatact gggccggcag 3361 gcgctccatt gcccagtcgg cagcgacatc cttcggcgcg attttgccgg ttactgcgct 3421 gtaccaaatg cgggacaacg taagcactac atttcgctca tcgccagccc agtcgggcgg 3481 cgagttccat agcgttaagg tttcatttag cgcctcaaat agatcctgtt caggaaccgg 3541 atcaaagagt tcctccgccg ctggacctac caaggcaacg ctatgttctc ttgcttttgt 3601 cagcaagata gccagatcaa tgtcgatcgt ggctggctcg aagatacctg caagaatgtc 3661 attgcgctgc cattctccaa attgcagttc gcgcttagct ggataacgcc acggaatgat 3721 gtcgtcgtgc acaacaatgg tgacttctac agcgcggaga atctcgctct ctccagggga 3781 agccgaagtt tccaaaaggt cgttgatcaa agctcgccgc gttgtttcat caagccttac 3841 ggtcaccgta accagcaaat caatatcact gtgtggcttc aggccgccat ccactgcgga 3901 gccgtacaaa tgtacggcca gcaacgtcgg ttcgagatgg cgctcgatga cgccaactac 3961 ctctgatagt tgagtcgata cttcggcgat caccgcttcc ctcatgatgt ttaactttgt 4021 tttagggcga ctgccctgct gcgtaacatc gttgctgctc cataacatca aacatcgacc 4081 cacggcgtaa cgcgcttgct gcttggatgc ccgaggcata gactgtaccc caaaaaaaca 4141 gtcataacaa gccatgaaaa ccgccactgc gccgttacca ccgctgcgtt cggtcaaggt 4201 tctggaccag ttgcgtgagc gcatacgcta cttgcattac agcttacgaa ccgaacaggc 4261 ttatgtccac tgggttcgtg ccttcatccg tttccacggt gtgcgtcacc cggcaacctt 4321 gggcagcagc gaagtcgagg catttctgtc ctggctggcg aacgagcgca aggtttcggt 4381 ctccacgcat cgtcaggcat tggcggcctt gctgttcttc tacggcaagg tgctgtgcac 4441 ggatctgccc tggcttcagg agatcggaag acctcggccg tcgcggcgct tgccggtggt 4501 gctgaccccg gatgaagtgg ttcgcatcct cggttttctg gaaggcgagc atcgtttgtt 4561 cgcccagctt ctgtatggaa cgggcatgcg gatcagtgag ggtttgcaac tgcgggtcaa 4621 ggatctggat ttcgatcacg gcacgatcat cgtgcgggag ggcaagggct ccaaggatcg 4681 ggccttgatg ttacccgaga gcttggcacc cagcctgcgc gagcagggga attgatccgg 4741 tggatgacct tttgaatgac ctttaataga ttatattact aattaattgg ggaccctaga 4801 ggtccccttt tttattttac tgcgatgagt ggcagggcgg ggcgtaattt ttttaaggca 4861 gttattggtg cccttaaacg cctggtgcta cgcctgaata agtgataata agcggatgaa 4921 tggcagaaat tcgatatcta gatctcgagc tggatacttc ccgtccgcca gggggacatg 4981 ccggcgatgc tgaaggtcgc gcgcattccc gatgaagagg ccggttaccg cctgtttgag 5041 gatatagtaa tctttctaaa tagctttgga ttggaggagt atggccacta atactaagtt 5101 cagctaataa aaaaatttgc taaagaactc cagctggatt tcactgatga gaatatcgtc 5161 ggagataaat ataataattc cacggactat agactatact agtatactcc gtctactgta 5221 cgatacactt ccgctcaggt ccttgtcctt taacgaggat tgttacgtac gctaatggcg 5281 tcaaaacaaa gactctagac ctaggcctta agatcgaaag agctggtagt tggcgcactg 5341 ttcgaagaac tgccgatgtc cagtaagatt cttactatgc tggttgaacc ggatgctggt 5401 aaagctactt gggttgctgc ttctacttat ggtaccgata caactactgg tgaggaagtt 5461 aaaggagctc gcgaaagctt gcatgcctgc agcaatggca acaacgttgc gcaaactatt 5521 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga 5581 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa 5641 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa 5701 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa 5761 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt 5821 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt 5881 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg 5941 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt 6001 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca 6061 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac 6121 tgtccttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac 6181 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct 6241 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg 6301 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca 6361 gcgtgagcat tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt 6421 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta 6481 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc 6541 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc 6601 cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa 6661 ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag 6721 cgagtcagtg agcgaggaag cggaagagcg cctgatgcgg tattttctcc ttacgcatct 6781 gtgcggtatt tcacaccg //