LOCUS Exported 3343 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Destination vector for Golden Gate assembly; 3' entry vector for Gateway cloning. ACCESSION . VERSION . KEYWORDS p3E_GGWDest+ SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 3343) AUTHORS Kirchmaier S, Lust K, Wittbrodt J TITLE Golden GATEway cloning--a combinatorial approach to generate fusion and recombination constructs. JOURNAL PLoS One. 2013 Oct 7;8(10):e76117. doi: 10.1371/journal.pone.0076117. PUBMED 24116091 REFERENCE 2 (bases 1 to 3343) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..3343 /organism="synthetic DNA construct" /mol_type="other DNA" terminator 268..295 /label=rrnB T2 terminator /note="transcription terminator T2 from the E. coli rrnB gene" terminator 387..473 /gene="Escherichia coli rrnB" /label=rrnB T1 terminator /note="transcription terminator T1 from the E. coli rrnB gene" primer_bind 537..553 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" regulatory 764..773 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS complement(786..1169) /codon_start=1 /gene="lacZ fragment" /product="LacZ-alpha fragment of beta-galactosidase" /label=lacZ-alpha /translation="MTMITPSYLGDTIEYSSYASNALGALPYGRPAGGREFTSDIEFPR PPWRPGACDVGPNSPYSESYYNSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEA RTDRPSQQLRSLNGEWTRPVAAH" primer_bind 950..966 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" promoter 973..991 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" regulatory 1022..1031 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" regulatory 1026..1035 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" promoter complement(1129..1147) /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind complement(1130..1147) /label=SP6 /note="SP6 promoter, forward primer" primer_bind complement(1165..1181) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1178..1200) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 1189..1205 /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(1213..1243) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 1258..1279 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind 1435..1451 /label=pBluescriptKS /note="For pBluescript vector" protein_bind complement(1452..1547) /gene="mutant version of attL" /label=attL3 /bound_moiety="LR Clonase(TM)" /note="recombination site for the Gateway(R) LR reaction" promoter complement(1585..1603) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(1608..1624) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1726..1745) /label=pENTR-R /note="pENTR vectors, reverse primer" CDS 1737..2546 /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="MSHIQRETSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind complement(1804..1823) /label=Kan-R /note="Kanamycin resistance gene, reverse primer" rep_origin 2693..3281 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 3182..3201 /label=pBR322ori-F /note="pBR322 origin, forward primer" ORIGIN 1 ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg agtgagctga 61 taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga 121 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca 181 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaata cgcgtaccgc 241 tagccaggaa gagtttgtag aaacgcaaaa aggccatccg tcaggatggc cttctgctta 301 gtttgatgcc tggcagttta tggcgggcgt cctgcccgcc accctccggg ccgttgcttc 361 acaacgttca aatccgctcc cggcggattt gtcctactca ggagagcgtt caccgacaaa 421 caacagataa aacgaaaggc ccagtcttcc gactgagcct ttcgttttat ttgatgcctg 481 gcagttccct actctcgcgt taacgctagc atggatgttt tcccagtcac gacgttgtaa 541 aacgacggcc agtcttaagc tcgggccctg cagctctaga gctcgaattc tacaggtcac 601 taataccatc taagtagttg gttcatagtg actgcatatg ttgtgtttta cagtattatg 661 tagtctgttt tttatgcaaa atctaattta atatattgat atttatatca ttttacgttt 721 ctcgttcaac tttcttgtac aaagtgaggg cgaattgggt accgccgcca tggcgagacc 781 cgcgcttaat gcgccgctac agggcgcgtc cattcgccat tcaggctgcg caactgttgg 841 gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct 901 gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg 961 gccagtgaat tgtaatacga ctcactatag ggcgaattgg gcccgacgtc gcatgctccc 1021 ggccgccatg gcggccgcgg gaattcgata tcactagtga attcgcggcc gcctgcaggt 1081 cgaccatatg ggagagctcc caacgcgttg gatgcatagc ttgagtattc tatagtgtca 1141 cctaaatagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct 1201 cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg 1261 agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct 1321 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg 1381 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gggtctctta aggctcgagg 1441 tcgacggtat ccaactttat tatacaaagt tggcattata aaaaagcatt gcttatcaat 1501 ttgttgcaac gaacaggtca ctatcagtca aaataaaatc attatttgga gctccatggt 1561 agcgttaacg cggccgcgat atcccctata gtgagtcgta ttacatggtc atagctgttt 1621 cctggcagct ctggcccgtg tctcaaaatc tctgatgtta cattgcacaa gataaaaata 1681 tatcatcatg aacaataaaa ctgtctgctt acataaacag taatacaagg ggtgttatga 1741 gccatattca acgggaaacg tcgaggccgc gattaaattc caacatggat gctgatttat 1801 atgggtataa atgggctcgc gataatgtcg ggcaatcagg tgcgacaatc tatcgcttgt 1861 atgggaagcc cgatgcgcca gagttgtttc tgaaacatgg caaaggtagc gttgccaatg 1921 atgttacaga tgagatggtc agactaaact ggctgacgga atttatgcct cttccgacca 1981 tcaagcattt tatccgtact cctgatgatg catggttact caccactgcg atccccggaa 2041 aaacagcatt ccaggtatta gaagaatatc ctgattcagg tgaaaatatt gttgatgcgc 2101 tggcagtgtt cctgcgccgg ttgcattcga ttcctgtttg taattgtcct tttaacagcg 2161 atcgcgtatt tcgtctcgct caggcgcaat cacgaatgaa taacggtttg gttgatgcga 2221 gtgattttga tgacgagcgt aatggctggc ctgttgaaca agtctggaaa gaaatgcata 2281 aacttttgcc attctcaccg gattcagtcg tcactcatgg tgatttctca cttgataacc 2341 ttatttttga cgaggggaaa ttaataggtt gtattgatgt tggacgagtc ggaatcgcag 2401 accgatacca ggatcttgcc atcctatgga actgcctcgg tgagttttct ccttcattac 2461 agaaacggct ttttcaaaaa tatggtattg ataatcctga tatgaataaa ttgcagtttc 2521 atttgatgct cgatgagttt ttctaatcag aattggttaa ttggttgtaa cactggcaga 2581 gcattacgct gacttgacgg gacggcgcaa gctcatgacc aaaatccctt aacgtgagtt 2641 acgcgtcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc 2701 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg 2761 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag 2821 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact 2881 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg 2941 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc 3001 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg 3061 aactgagata cctacagcgt gagcattgag aaagcgccac gcttcccgaa gggagaaagg 3121 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag 3181 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc 3241 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct 3301 ttttacggtt cctggccttt tgctggcctt ttgctcacat gtt //