LOCUS Exported 7647 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS MIGS1.1 (JW493) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7647) AUTHORS de Felippes FF, Wang JW, Weigel D TITLE MIGS: MiRNA Induced Gene Silencing. JOURNAL Plant J. 2011 Dec 28. doi: 10.1111/j.1365-313X.2011.04896.x. PUBMED 22211571 REFERENCE 2 (bases 1 to 7647) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7647 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 542..564 /label=LB T-DNA repeat /note="left border repeat from nopaline C58 T-DNA (truncated)" terminator 588..840 /label=NOS terminator /note="nopaline synthase terminator and poly(A) signal" CDS complement(860..1411) /codon_start=1 /gene="Streptomyces hygroscopicus bar" /product="phosphinothricin acetyltransferase" /label=BlpR /note="confers resistance to bialophos or phosphinothricin" /translation="MSPERRPADIRRATEADMPAVCTIVNHYIETSTVNFRTEPQEPQE WTDDLVRLRERYPWLVAEVDGEVAGIAYAGPWKARNAYDWTAESTVYVSPRHQRTGLGS TLYTHLLKSLEAQGFKSVVAVIGLPNDPSVRMHEALGYAPRGMLRAAGFKHGNWHDVGF WQLDFSLPVPPRPVLPVTEI" promoter complement(1453..1632) /label=NOS promoter /note="nopaline synthase promoter" primer_bind 1869..1891 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 1883..1900 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 1884..1900 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 1910..1929 /label=T7 /note="T7 promoter, forward primer" promoter 1910..1928 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind 1953..1969 /label=pBluescriptKS /note="For pBluescript vector" primer_bind 1954..1970 /label=KS primer /note="common sequencing primer, one of multiple similar variants" promoter 2507..2852 /label=CaMV 35S promoter /note="strong constitutive promoter from cauliflower mosaic virus" primer_bind 2795..2815 /label=35S promoter /note="CaMV 35S promoter, forward primer" protein_bind 2899..3023 /gene="mutant version of attR" /label=attR1 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter 3048..3078 /label=lac UV5 promoter /note="E. coli lac promoter with an ""up"" mutation" CDS 3132..3791 /codon_start=1 /gene="cat" /product="chloramphenicol acetyltransferase" /label=CmR /note="confers resistance to chloramphenicol" /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA" primer_bind complement(3198..3217) /label=CAT-R /note="Chloramphenicol resistance gene, reverse primer" CDS 4133..4438 /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 4317..4336 /label=ccdB-fwd /note="ccdB gene, forward primer" protein_bind complement(4479..4603) /gene="mutant version of attR" /label=attR2 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" terminator 4610..5251 /label=E9 terminator /note="terminator and polyadenylation signal from the pea rbcS-E9 gene" promoter complement(5273..5291) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(5312..5328) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5312..5328) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(5325..5347) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 5336..5352 /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)." promoter complement(5360..5391) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5406..5427 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(5544..5561) /label=L4440 /note="L4440 vector, forward primer" misc_feature 5666..5690 /label=RB T-DNA repeat /note="right border repeat from nopaline C58 T-DNA" rep_origin complement(5781..6369) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5861..5880) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6540..7328) /codon_start=1 /gene="aadA" /product="aminoglycoside adenylyltransferase (Murphy, 1985)" /label=SmR /note="confers resistance to spectinomycin and streptomycin" /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP VILEARQAYLGQEDRLASRADQLEEFVHYVKGEITKVVGK" rep_origin 7619..407 /label=pSa ori /note="origin of replication from bacterial plasmid pSa" ORIGIN 1 tttttatccc cggaagcctg tggatagagg gtagttatcc acgtgaaacc gctaatgccc 61 cgcaaagcct tgattcacgg ggctttccgg cccgctccaa aaactatcca cgtgaaatcg 121 ctaatcaggg tacgtgaaat cgctaatcgg agtacgtgaa atcgctaata aggtcacgtg 181 aaatcgctaa tcaaaaaggc acgtgagaac gctaatagcc ctttcagatc aacagcttgc 241 aaacacccct cgctccggca agtagttaca gcaagtagta tgttcaatta gcttttcaat 301 tatgaatata tatatcaatt attggtcgcc cttggcttgt ggacaatgcg ctacgcgcac 361 cggctccgcc cgtggacaac cgcaagcggt tgcccaccgt cgagcgccag cgcctttgcc 421 cacaacccgg cggccggccg caacagatcg ttttataaat tttttttttt gaaaaagaaa 481 aagcccgaaa ggcggcaacc tctcgggctt ctggatttcc gatccccgga attagagatc 541 ttggcaggat atattgtggt gtaacgttat cagcttgcat gccggtcgat ctagtaacat 601 agatgacacc gcgcgcgata atttatccta gtttgcgcgc tatattttgt tttctatcgc 661 gtattaaatg tataattgcg ggactctaat cataaaaacc catctcataa ataacgtcat 721 gcattacatg ttaattatta catgcttaac gtaattcaac agaaattata tgataatcat 781 cgcaagaccg gcaacaggat tcaatcttaa gaaactttat tgccaaatgt ttgaacgatc 841 tgcttgactc taggggtcat cagatttcgg tgacgggcag gaccggacgg ggcggcaccg 901 gcaggctgaa gtccagctgc cagaaaccca cgtcatgcca gttcccgtgc ttgaagccgg 961 ccgcccgcag catgccgcgg ggggcatatc cgagcgcctc gtgcatgcgc acgctcgggt 1021 cgttgggcag cccgatgaca gcgaccacgc tcttgaagcc ctgtgcctcc agggacttca 1081 gcaggtgggt gtagagcgtg gagcccagtc ccgtccgctg gtggcggggg gagacgtaca 1141 cggtggactc ggccgtccag tcgtaggcgt tgcgtgcctt ccagggaccc gcgtaggcga 1201 tgccggcgac ctcgccgtcc acctcggcga cgagccaggg atagcgctcc cgcagacgga 1261 cgaggtcgtc cgtccactcc tgcggttcct gcggctcggt acggaagttg accgtgcttg 1321 tctcgatgta gtggttgacg atggtgcaga ccgccggcat gtccgcctcg gtggcacggc 1381 ggatgtcggc cgggcgtcgt tctgggctca tggtagatcc cctcgatcga gttgagagtg 1441 aatatgagac tctaattgga taccgagggg aatttatgga acgtcagtgg agcatttttg 1501 acaagaaata tttgctagct gatagtgacc ttaggcgact tttgaacgcg caataatggt 1561 ttctgacgta tgtgcttagc tcattaaact ccagaaaccc gcggctcagt ggctccttca 1621 acgttgcggt tctgtcagtt ccaaacgtaa aacggcttgt cccgcgtcat cggcgggggt 1681 cataacgtga ctcccttaat tctccgctca tgtatcgata acattaacgt ttacaatttc 1741 gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc 1801 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc 1861 agggttttcc cagtcacgac gttgtaaaac gacggccagt gagcgcgcgt aatacgactc 1921 actatagggc gaattgggta ccgggccccc cctcgaggtc gacggtatcg ataagcttga 1981 tatcgaattc ctgcagcccg ggggatccat tcggtcccca gattagcctt ttcaatttca 2041 gaaagaatgc taacccacag atggttagag aggcttacgc agcaggtctc atcaagacga 2101 tctacccgag caataatctc caggaaatca aataccttcc caagaaggtt aaagatgcag 2161 tcaaaagatt caggactaac tgcatcaaga acacagagaa agatatattt ctcaagatca 2221 gaagtactat tccagtatgg acgattcaag gcttgcttca caaaccaagg caagtaatag 2281 agattggagt ctctaaaaag gtagttccca ctgaatcaaa ggccatggag tcaaagattc 2341 aaatagagga cctaacagaa ctcgccgtaa agactggcga acagttcata cagagtctct 2401 tacgactcaa tgacaagaag aaaatcttcg tcaacatggt ggagcacgac acacttgtct 2461 actccaaaaa tatcaaagat acagtctcag aagaccaaag ggcaattgag acttttcaac 2521 aaagggtaat atccggaaac ctcctcggat tccattgccc agctatctgt cactttattg 2581 tgaagatagt ggaaaaggaa ggtggctcct acaaatgcca tcattgcgat aaaggaaagg 2641 ccatcgttga agatgcctct gccgacagtg gtcccaaaga tggaccccca cccacgagga 2701 gcatcgtgga aaaagaagac gttccaacca cgtcttcaaa gcaagtggat tgatgtgata 2761 tctccactga cgtaagggat gacgcacaat cccactatcc ttcgcaagac ccttcctcta 2821 tataaggaag ttcatttcat ttggagagaa cacgggggac gagctgtgat ttttctctac 2881 aagcgaatct agaggatcac aagtttgtac aaaaaagctg aacgagaaac gtaaaatgat 2941 ataaatatca atatattaaa ttagattttg cataaaaaac agactacata atactgtaaa 3001 acacaacata tccagtcata ttggcggccc cattaggcac cccaggcttt acactttatg 3061 cttccggctc gtataatgtg tggattttga gttaggagcc gtcgagattt tcaggagcta 3121 aggaagctaa aatggagaaa aaaatcactg gatataccac cgttgatata tcccaatggc 3181 atcgtaaaga acattttgag gcatttcagt cagttgctca atgtacctat aaccagaccg 3241 ttcagctgga tattacggcc tttttaaaga ccgtaaagaa aaataagcac aagttttatc 3301 cggcctttat tcacattctt gcccgcctga tgaatgctca tccggagttc cgtatggcaa 3361 tgaaagacgg tgagctggtg atatgggata gtgttcaccc ttgttacacc gttttccatg 3421 agcaaactga aacgttttca tcgctctgga gtgaatacca cgacgatttc cggcagtttc 3481 tacacatata ttcgcaagat gtggcgtgtt acggtgaaaa cctggcctat ttccctaaag 3541 ggtttattga gaatatgttt ttcgtctcag ccaatccctg ggtgagtttc accagttttg 3601 atttaaacgt ggccaatatg gacaacttct tcgcccccgt tttcaccatg ggcaaatatt 3661 atacgcaagg cgacaaggtg ctgatgccgc tggcgattca ggttcatcat gccgtttgtg 3721 atggcttcca tgtcggcaga atgcttaatg aattacaaca gtactgcgat gagtggcagg 3781 gcggggcgta aacgcgtgga gccggcttac taaaagccag ataacagtat gcgtatttgc 3841 gcgctgattt ttgcggtata agaatatata ctgatatgta tacccgaagt atgtcaaaaa 3901 gaggtatgct atgaagcagc gtattacagt gacagttgac agcgacagct atcagttgct 3961 caaggcatat atgatgtcaa tatctccggt ctggtaagca caaccatgca gaatgaagcc 4021 cgtcgtctgc gtgccgaacg ctggaaagcg gaaaatcagg aagggatggc tgaggtcgcc 4081 cggtttattg aaatgaacgg ctcttttgct gacgagaaca ggggctggtg aaatgcagtt 4141 taaggtttac acctataaaa gagagagccg ttatcgtctg tttgtggatg tacagagtga 4201 tattattgac acgcccggcc gacggatggt gatccccctg gccagtgcac gtctgctgtc 4261 agataaagtc tcccgtgaac tttacccggt ggtgcatatc ggggatgaaa gctggcgcat 4321 gatgaccacc gatatggcca gtgtgccggt ctccgttatc ggggaagaag tggctgatct 4381 cagccaccgc gaaaatgaca tcaaaaacgc cattaacctg atgttctggg gaatataaat 4441 gtcaggctcc cttatacaca gccagtctgc acctcgacca tagtgactgg atatgttgtg 4501 ttttacagta ttatgtagtc tgttttttat gcaaaatcta atttaatata ttgatattta 4561 tatcatttta cgtttctcgt tcagctttct tgtacaaagt ggtgatccta gctttcgttc 4621 gtatcatcgg tttcgacaac gttcgtcaag ttcaatgcat cagtttcatt gcgcacacac 4681 cagaatccta ctgagtttga gtattatggc attgggaaaa ctgtttttct tgtaccattt 4741 gttgtgcttg taatttactg tgttttttat tcggttttcg ctatcgaact gtgaaatgga 4801 aatggatgga gaagagttaa tgaatgatat ggtccttttg ttcattctca aattaatatt 4861 atttgttttt tctcttattt gttgtgtgtt gaatttgaaa ttataagaga tatgcaaaca 4921 ttttgttttg agtaaaaatg tgtcaaatcg tggcctctaa tgaccgaagt taatatgagg 4981 agtaaaacac ttgtagttgt accattatgc ttattcacta ggcaacaaat atattttcag 5041 acctagaaaa gctgcaaatg ttactgaata caagtatgtc ctcttgtgtt ttagacattt 5101 atgaactttc ctttatgtaa ttttccagaa tccttgtcag attctaatca ttgctttata 5161 attatagtta tactcatgga tttgtagttg agtatgaaaa tattttttaa tgcattttat 5221 gacttgccaa ttgattgaca acatgcatca atcggagctc cagcttttgt tccctttagt 5281 gagggttaat tccgagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt 5341 atccgctcac aattccacac aacatacgag ccggaaghca taaagtgtaa agcctggggt 5401 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg 5461 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg 5521 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg 5581 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat 5641 aacgcaggaa agaacatgaa ggccttgaca ggatatattg gcgggtaaac taagtcgctg 5701 tatgtgtttg tttgagatct catgtgagca aaaggccagc aaaaggccag gaaccgtaaa 5761 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat 5821 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc 5881 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc 5941 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt 6001 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac 6061 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg 6121 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca 6181 gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc 6241 gctctgctga agccagttac cttcggaaga agagttggta gctcttgatc cggcaaacaa 6301 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa 6361 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac 6421 tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta 6481 aattaaaaat gaagttttaa atcaatctaa agtatatatg tgtaacattg gtctagtgat 6541 tatttgccga ctaccttggt gatctcgcct ttcacgtagt gaacaaattc ttccaactga 6601 tctgcgcgcg aggccaagcg atcttcttgt ccaagataag cctgcctagc ttcaagtatg 6661 acgggctgat actgggccgg caggcgctcc attgcccagt cggcagcgac atccttcggc 6721 gcgattttgc cggttactgc gctgtaccaa atgcgggaca acgtaagcac tacatttcgc 6781 tcatcgccag cccagtcggg cggcgagttc catagcgtta aggtttcatt tagcgcctca 6841 aatagatcct gttcaggaac cggatcaaag agttcctccg ccgctggacc taccaaggca 6901 acgctatgtt ctcttgcttt tgtcagcaag atagccagat caatgtcgat cgtggctggc 6961 tcgaagatac ctgcaagaat gtcattgcgc tgccattctc caaattgcag ttcgcgctta 7021 gctggataac gccacggaat gatgtcgtcg tgcacaacaa tggtgacttc tacagcgcgg 7081 agaatctcgc tctctccagg ggaagccgaa gtttccaaaa ggtcgttgat caaagctcgc 7141 cgcgttgttt catcaagcct tacggtcacc gtaaccagca aatcaatatc actgtgtggc 7201 ttcaggccgc catccactgc ggagccgtac aaatgtacgg ccagcaacgt cggttcgaga 7261 tggcgctcga tgacgccaac tacctctgat agttgagtcg atacttcggc gatcaccgct 7321 tccctcataa caccccttgt attactgttt atgtaagcag acagttttat tgttcatgat 7381 gatatatttt tatcttgtgc aatgtaacat cagagatttt gagacacaac gtggctttgt 7441 tgaataaatc gaacttttgc tgagttgaag gatcagatca cgcatcttcc cgacaacgca 7501 gaccgttccg tggcaaagca aaagttcaaa atcaccaact ggtccaccta caacaaagct 7561 ctcatcaacc gtggctccct cactttctgg ctggatgatg gggcgattca ggcgatcccc 7621 atccaacagc ccgccgtcga gcgggct //