LOCUS Exported 6560 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pDESTsmg SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6560) AUTHORS Goldstone RM, Moreland NJ, Bashiri G, Baker EN, Shaun Lott J TITLE A new Gateway vector and expression protocol for fast and efficient recombinant protein expression in Mycobacterium smegmatis. JOURNAL Protein Expr Purif. 2008 Jan . 57(1):81-7. PUBMED 17949993 REFERENCE 2 (bases 1 to 6560) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6560 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 61..80 /label=T7 /note="T7 promoter, forward primer" promoter 61..79 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" protein_bind 80..104 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." RBS 119..141 /note="efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)" CDS 154..171 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" protein_bind 187..311 /gene="mutant version of attR" /label=attR1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 336..366 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" CDS 420..1079 /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA" primer_bind complement(486..505) /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" CDS 1421..1726 /codon_start=1 /gene="ccdB" /product="CcdB, a bacterial toxin that poisons DNA gyrase" /label=ccdB /note="Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains." /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI" primer_bind 1605..1624 /label=ccdB-fwd /note="ccdB gene, forward primer" protein_bind complement(1767..1891) /gene="mutant version of attR" /label=attR2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" CDS 1941..1958 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(2011..2029) /label=T7 Term /note="T7 terminator, reverse primer" terminator 2025..2072 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" rep_origin 2109..2537 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2196..2215) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2406..2427 /label=F1ori-F /note="F1 origin, forward primer" rep_origin complement(4640..5228) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4720..4739) /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 ccggtgatgc cggccacgat gcgtccggcg tagaggatcg agatctcgat cccgcgaaat 61 taatacgact cactataggg gaattgtgag cggataacaa ttcccctcta gaaataattt 121 tgtttaactt taagaaggag atataccatg tcgcatcacc atcaccatca cctcgaatca 181 ccatcaacaa gtttgtacaa aaaagctgaa cgagaaacgt aaaatgatat aaatatcaat 241 atattaaatt agattttgca taaaaaacag actacataat actgtaaaac acaacatatc 301 cagtcactat ggcggccgca ttaggcaccc caggctttac actttatgct tccggctcgt 361 ataatgtgtg gattttgagt taggatccgt cgagattttc aggagctaag gaagctaaaa 421 tggagaaaaa aatcactgga tataccaccg ttgatatatc ccaatggcat cgtaaagaac 481 attttgaggc atttcagtca gttgctcaat gtacctataa ccagaccgtt cagctggata 541 ttacggcctt tttaaagacc gtaaagaaaa ataagcacaa gttttatccg gcctttattc 601 acattcttgc ccgcctgatg aatgctcatc cggaattccg tatggcaatg aaagacggtg 661 agctggtgat atgggatagt gttcaccctt gttacaccgt tttccatgag caaactgaaa 721 cgttttcatc gctctggagt gaataccacg acgatttccg gcagtttcta cacatatatt 781 cgcaagatgt ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg tttattgaga 841 atatgttttt cgtctcagcc aatccctggg tgagtttcac cagttttgat ttaaacgtgg 901 ccaatatgga caacttcttc gcccccgttt tcaccatggg caaatattat acgcaaggcg 961 acaaggtgct gatgccgctg gcgattcagg ttcatcatgc cgtttgtgat ggcttccatg 1021 tcggcagaat gcttaatgaa ttacaacagt actgcgatga gtggcagggc ggggcgtaaa 1081 gatctggatc cggcttacta aaagccagat aacagtatgc gtatttgcgc gctgattttt 1141 gcggtataag aatatatact gatatgtata cccgaagtat gtcaaaaaga ggtatgctat 1201 gaagcagcgt attacagtga cagttgacag cgacagctat cagttgctca aggcatatat 1261 gatgtcaata tctccggtct ggtaagcaca accatgcaga atgaagcccg tcgtctgcgt 1321 gccgaacgct ggaaagcgga aaatcaggaa gggatggctg aggtcgcccg gtttattgaa 1381 atgaacggct cttttgctga cgagaacagg ggctggtgaa atgcagttta aggtttacac 1441 ctataaaaga gagagccgtt atcgtctgtt tgtggatgta cagagtgata ttattgacac 1501 gcccgggcga cggatggtga tccccctggc cagtgcacgt ctgctgtcag ataaagtctc 1561 ccgtgaactt tacccggtgg tgcatatcgg ggatgaaagc tggcgcatga tgaccaccga 1621 tatggccagt gtgccggtct ccgttatcgg ggaagaagtg gctgatctca gccaccgcga 1681 aaatgacatc aaaaacgcca ttaacctgat gttctgggga atataaatgt caggctccct 1741 tatacacagc cagtctgcag gtcgaccata gtgactggat atgttgtgtt ttacagtatt 1801 atgtagtctg ttttttatgc aaaatctaat ttaatatatt gatatttata tcattttacg 1861 tttctcgttc agctttcttg tacaaagtgg ttgatgatcc gaattcgagc tccgtcgaca 1921 agcttgcggc cgcactcgag caccaccacc accaccactg agatccggct gctaacaaag 1981 cccgaaagga agctgagttg gctgctgcca ccgctgagca ataactagca taaccccttg 2041 gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata tccggattgg 2101 cgaatgggac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag 2161 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt 2221 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt 2281 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg 2341 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt 2401 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt 2461 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca 2521 aaaatttaac gcgaatttta acaaaatatc tagatttaaa gatctggtac cgcggccgcg 2581 ccacggatgc caccacaggc actacaacgg agttcgccac gtacatcacc acaaccaccg 2641 attctggcgg tgagctcccc gatattcagc ggaaatggct tggtatcgac caagattcgt 2701 agaaccccgt ctcgtctggc tggtattcaa aacgggcgca acgaaacacg caacgagaca 2761 ggcatggccc aaaccagaaa actagcgtct accaggactt ttacctgtcc gacccgttgc 2821 aacggaaccc cccacggaac ccccgcgaca cccgctcccc aattgcgtta gaacagcggt 2881 ggattgtcgg cttcgttgtg ggccttttga gccgcttcct gttctgccgc acgctctttc 2941 ctcgcccgat agccgagtcg cttaacggtg tccagatgca gcccgaaatg tttggccgtt 3001 tgcggccaag agtggccctc gtcgtcgtga taggcgcgga tgcgttcgcg gcgtgcagcc 3061 tgctcggcga gccactcgct gcgttcctgc gccacgagcc ggacgacgtg gcgttcggat 3121 agtccggtga ttcgagcgcc ttcggcggcg gtcacgcgcc gctttttgcg gacagtcggc 3181 tgccggttgt agccgtcgct gtagccgtcg ctcatagcaa tgcctccatg catggctgac 3241 gcggactttg cgcgccgcgc aactgtgctc gccgccgtgc gcgctgctgc gcccttccgc 3301 gagatggccg actggcgcgc actgagtgtg gcctcgtaga ccacgatccc gtccgcccaa 3361 atgcgcgact tggttgtgat ccaacgccaa atgctgttgg cgatggcgcg gacctcgctg 3421 tccggtagcg gtccgggaca cacgtcgttg cacggaaatt cggcgtttcg cgcgtggcac 3481 tcggcataga tcgcgcggcc gagtccgtcc acgttccggg tcggcaggta gatccgcatg 3541 agggcgggac gataggccca caacctgacg gaatcgaaca gtgcgcaatt ccgccctagc 3601 ggcgtcggag ccgctttgta cgtggtctgc tgacgccagc gcggcggtgg catgttcgcg 3661 ccgagctcgg cctcgatgtg gctgagtgtg tagagatctg agtggagcca ttccgtttcc 3721 caggcgatgt ggccggggtt tttggtcatg aggcctgagt aactgcggtc gccatcgacg 3781 gcgcgccgaa ggccttcggc gcacgccgcc atgtatgcga gcggcttacg ccgcgcgtat 3841 tcggtgcgtg gaacaggggc gttgagtgcc cacactgcgt gtgcgtggcc gttggcgcga 3901 ttgcccacga tcgcgttggg cagcggatgg gacccccggg cgctgagcgc tcggagcgct 3961 gcgtctggat ggtctacgtc cacgaccagc aggtttgcca gcgctgttgg gttcgcctcg 4021 atgtaccggc ggcctagggc cgacgcgcgg ctttggcggt agatcccctc gagcagatcg 4081 tcgcttgcca gcggccagta cggcagccag agctgctcaa attcgtcggc gacgtggctc 4141 acgcttggta gtagaccacg attaatcacc ggtgtatggt ccgacacgag ctccaagtca 4201 gatatttcgc tgaggggcca ccccacaact gcacactccc ccgctctccc gtcgagccct 4261 ggtggtggaa caccagcgac agccgagcac ccccaaccac ctgtaccaac caggaggaac 4321 acatgcgtcg tttcgaggac gtttccgggc cgctgagagc cgctgtggcg gccgtacacg 4381 ccgccttaga cccgttagac cccctgccgc ctgaatgcgc gggtacgagc cacacagcgc 4441 ccgaacttac ggagctggtg ggctcacgcg tgcggccgca cgcgttgcgc tcggtcgttc 4501 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag 4561 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa 4621 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc 4681 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc 4741 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg 4801 cctttctccc ctcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt 4861 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc 4921 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc 4981 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag 5041 agttcttgaa gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg 5101 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa 5161 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag 5221 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcatgga actagagcat 5281 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc 5341 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc 5401 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgacgtggc 5461 cgaccagccc gtcatcgtca acgcctgatc cgcggtgcgg acaggccgtg tcgcgaccgg 5521 ccgtgcggaa ttaagccggc ccgtaccctg tgaatagagg tccgctgtga cacaagaatc 5581 cctgttactt ctcgaccgta ttgattcgga tgattcctac gcgagcctgc ggaacgacca 5641 ggaattctgg gagccgctgg cccgccgagc cctggaggag ctcgggctgc cggtgccgcc 5701 ggtgctgcgg gtgcccggcg agagcaccaa ccccgtactg gtcggcgagc ccggcccggt 5761 gatcaagctg ttcggcgagc actggtgcgg tccggagagc ctcgcgtcgg agtcggaggc 5821 gtacgcggtc ctggcggacg ccccggtgcc ggtgccccgc ctcctcggcc gcggcgagct 5881 gcggcccggc accggagcct ggccgtggcc ctacctggtg atgagccgga tgaccggcac 5941 cacctggcgg tccgcgatgg acggcacgac cgaccggaac gcgctgctcg ccctggcccg 6001 cgaactcggc cgggtgctcg gccggctgca cagggtgccg ctgaccggga acaccgtgct 6061 caccccccat tccgaggtct tcccggaact gctgcgggaa cgccgcgcgg cgaccgtcga 6121 ggaccaccgc gggtggggct acctctcgcc ccggctgctg gaccgcctgg aggactggct 6181 gccggacgtg gacacgctgc tggccggccg cgaaccccgg ttcgtccacg gcgacctgca 6241 cgggaccaac atcttcgtgg acctggccgc gaccgaggtc accgggatcg tcgacttcac 6301 cgacgtctat gcgggagact cccgctacag cctggtgcaa ctgcatctca acgccttccg 6361 gggcgaccgc gagatcctgg ccgcgctgct cgacggggcg cagtggaagc ggaccgagga 6421 cttcgcccgc gaactgctcg ccttcacctt cctgcacgac ttcgaggtgt tcgaggagac 6481 cccgctggat ctctccggct tcaccgatcc ggaggaactg gcgcagttcc tctgggggcc 6541 gccggacacc gcccccggcg //